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Sample records for intercellular communication transmembrane

  1. Terbinafine inhibits gap junctional intercellular communication

    International Nuclear Information System (INIS)

    Lee, Ju Yeun; Yoon, Sei Mee; Choi, Eun Ju; Lee, Jinu

    2016-01-01

    Terbinafine is an antifungal agent that selectively inhibits fungal sterol synthesis by blocking squalene epoxidase. We evaluated the effect of terbinafine on gap junctional intercellular communication (GJIC). Fluorescence recovery after photobleaching (FRAP) and I-YFP GJIC assays revealed that terbinafine inhibits GJIC in a reversible and dose-dependent manner in FRT-Cx43 and LN215 cells. Treatment with terbinafine did not affect Cx43 phosphorylation status or intracellular Ca 2+ concentration, well-known action mechanisms of various GJIC blockers. While a structurally related chemical, naftifine, attenuated GJIC, epigallocatechin gallate, another potent squalene epoxidase inhibitor with a different structure, did not. These results suggest that terbinafine inhibits GJIC with a so far unknown mechanism of action. - Highlights: • In vitro pharmacological studies were performed on FRT-Cx43 and LN215 cells. • Terbinafine inhibits gap junctional intercellular communication in both cell lines. • The inhibitory effect of terbinafine is reversible and dose-dependent. • Treatment of terbinafine does not alter Cx43 phosphorylation or cytosolic Ca 2+ concentration. • Inhibition of squalene epoxidase is not involved in this new effect of terbinafine.

  2. Terbinafine inhibits gap junctional intercellular communication

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Ju Yeun, E-mail: whitewndus@naver.com [College of Pharmacy, Yonsei Institute of Pharmaceutical Sciences, Yonsei University, 85 Songdogwahak-ro, Yeonsu-gu, Incheon 21983 (Korea, Republic of); Yoon, Sei Mee, E-mail: sei_mee@naver.com [College of Pharmacy, Yonsei Institute of Pharmaceutical Sciences, Yonsei University, 85 Songdogwahak-ro, Yeonsu-gu, Incheon 21983 (Korea, Republic of); Department of Integrated OMICS for Biomedical Sciences, Yonsei University, 50 Yonsei-ro, Seodaemun-gu, Seoul 120-749 (Korea, Republic of); Choi, Eun Ju, E-mail: yureas@naver.com [College of Pharmacy, Yonsei Institute of Pharmaceutical Sciences, Yonsei University, 85 Songdogwahak-ro, Yeonsu-gu, Incheon 21983 (Korea, Republic of); Lee, Jinu, E-mail: jinulee@yonsei.ac.kr [College of Pharmacy, Yonsei Institute of Pharmaceutical Sciences, Yonsei University, 85 Songdogwahak-ro, Yeonsu-gu, Incheon 21983 (Korea, Republic of)

    2016-09-15

    Terbinafine is an antifungal agent that selectively inhibits fungal sterol synthesis by blocking squalene epoxidase. We evaluated the effect of terbinafine on gap junctional intercellular communication (GJIC). Fluorescence recovery after photobleaching (FRAP) and I-YFP GJIC assays revealed that terbinafine inhibits GJIC in a reversible and dose-dependent manner in FRT-Cx43 and LN215 cells. Treatment with terbinafine did not affect Cx43 phosphorylation status or intracellular Ca{sup 2+} concentration, well-known action mechanisms of various GJIC blockers. While a structurally related chemical, naftifine, attenuated GJIC, epigallocatechin gallate, another potent squalene epoxidase inhibitor with a different structure, did not. These results suggest that terbinafine inhibits GJIC with a so far unknown mechanism of action. - Highlights: • In vitro pharmacological studies were performed on FRT-Cx43 and LN215 cells. • Terbinafine inhibits gap junctional intercellular communication in both cell lines. • The inhibitory effect of terbinafine is reversible and dose-dependent. • Treatment of terbinafine does not alter Cx43 phosphorylation or cytosolic Ca{sup 2+} concentration. • Inhibition of squalene epoxidase is not involved in this new effect of terbinafine.

  3. Gap Junctional Intercellular Communication and Breast Cancer Metastasis to Bone

    National Research Council Canada - National Science Library

    Donahue, Henry

    2001-01-01

    .... We found that: 1) expressing the metastasis suppressing gene BRMS1 in diverse cancer cell lines, including breast and melanoma, restores homotypic gap junctional intercellular communication (GJIC); 2...

  4. Terbinafine inhibits gap junctional intercellular communication.

    Science.gov (United States)

    Lee, Ju Yeun; Yoon, Sei Mee; Choi, Eun Ju; Lee, Jinu

    2016-09-15

    Terbinafine is an antifungal agent that selectively inhibits fungal sterol synthesis by blocking squalene epoxidase. We evaluated the effect of terbinafine on gap junctional intercellular communication (GJIC). Fluorescence recovery after photobleaching (FRAP) and I-YFP GJIC assays revealed that terbinafine inhibits GJIC in a reversible and dose-dependent manner in FRT-Cx43 and LN215 cells. Treatment with terbinafine did not affect Cx43 phosphorylation status or intracellular Ca(2+) concentration, well-known action mechanisms of various GJIC blockers. While a structurally related chemical, naftifine, attenuated GJIC, epigallocatechin gallate, another potent squalene epoxidase inhibitor with a different structure, did not. These results suggest that terbinafine inhibits GJIC with a so far unknown mechanism of action. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Mobile Transcripts and Intercellular Communication in Plants.

    Science.gov (United States)

    Saplaoura, E; Kragler, F

    2016-01-01

    Phloem serves as a highway for mobile signals in plants. Apart from sugars and hormones, proteins and RNAs are transported via the phloem and contribute to the intercellular communication coordinating growth and development. Different classes of RNAs have been found mobile and in the phloem exudate such as viral RNAs, small interfering RNAs (siRNAs), microRNAs, transfer RNAs, and messenger RNAs (mRNAs). Their transport is considered to be mediated via ribonucleoprotein complexes formed between phloem RNA-binding proteins and mobile RNA molecules. Recent advances in the analysis of the mobile transcriptome indicate that thousands of transcripts move along the plant axis. Although potential RNA mobility motifs were identified, research is still in progress on the factors triggering siRNA and mRNA mobility. In this review, we discuss the approaches used to identify putative mobile mRNAs, the transport mechanism, and the significance of mRNA trafficking. © 2016 Elsevier Inc. All rights reserved.

  6. Intercellular Communication in Malignant Pleural Mesothelioma: Properties of Tunneling Nanotubes

    Directory of Open Access Journals (Sweden)

    Justin William Ady

    2014-10-01

    Full Text Available Malignant pleural mesothelioma is a particularly aggressive and locally invasive malignancy with a poor prognosis despite advances in understanding of cancer cell biology and development of new therapies. At the cellular level, cultured mesothelioma cells present a mesenchymal appearance and a strong capacity for local cellular invasion. One important but underexplored area of mesothelioma cell biology is intercellular communication. Our group has previously characterized in multiple histological subtypes of mesothelioma a unique cellular protrusion known as tunneling nanotubes (TnTs. TnTs are long, actin filament-based, narrow cytoplasmic extensions that are non-adherent when cultured in vitro and are capable of shuttling cellular cargo between connected cells. Our prior work confirmed the presence of nanotube structures in tumors resected from patients with human mesothelioma. In our current study, we quantified the number of TnTs/cell among various mesothelioma subtypes and normal mesothelial cells using confocal microscopic techniques. We also examined TnT length among adherent cells and cells in suspension. We further examined potential approaches to the in vivo study of TnTs in animal models of cancer. We have developed novel approaches to study TnTs in aggressive solid tumor malignancies and define fundamental characteristics of TnTs in malignant mesothelioma. There is mounting evidence that TnTs play an important role in intercellular communication in mesothelioma and thus merit further investigation of their role in vivo.

  7. Na,K-pump modulates intercellular communication in vascular wall

    DEFF Research Database (Denmark)

    Matchkov, Vladimir; Nilsson, Holger; Aalkjær, Christian

      Ouabain, a specific inhibitor of the Na,K-pump, has previously been shown to interfere with intercellular communication. Here we test the hypothesis that the communication between vascular smooth muscle cells (SMCs) is regulated through an interaction between the Na,K-pump and the Na...... were used as a model for electrical coupling of SMCs by measuring membrane capacitance (Cm). SMCs were uncoupled (evaluated by inhibition of vasomotion and desynchronization of calcium transients in vascular wall, or by reduction to half of Cm measured in paired A7r5 cells) when the Na,K-pump...... was inhibited either by a low concentration of ouabain or by ATP depletion. Uncoupling with ouabain was associated with a localized increase of intracellular calcium in discrete sites near the plasma membrane. Reduction of Na,K-pump activity by removal of extracellular potassium also uncoupled cells, but only...

  8. Inhibition of intercellular communication by airborne particulate matter

    Energy Technology Data Exchange (ETDEWEB)

    Heussen, G.A.H. (Landbouwhogeschool Wageningen (Netherlands). Dept. of Toxicology)

    1991-04-01

    To investigate the inhibition of gap junction mediated intercellular communication (IC) by extracts of airborne particulate matter (APM), V79 cells were incubated with extracts of APM and subsequently microinjected with the fluorescent dye Lucifer Yellow, after which the number of fluorescent (= communicating) cells was determined. To compare inhibitory effects on IC with mutagenicity, APM was also tested in the Salmonella microsome assay. Six different extracts were tested, two outdoor extracts representing a heavily polluted and a relatively clean sample, and four indoor extracts, taken either in livingrooms with or without wood combustion in an open fire place, or in a room with or without cigarette smoking. Non-cytotoxic doses of outdoor and indoor APM inhibited IC in V79 cells in dose- and time-dependent manner. Mutagenicity data and IC data were correlated. These results suggest that APM has tumor promoter activity in addition to mutagenic activity. (orig.).

  9. Na,K-pump modulates intercellular communication in vascular wall

    DEFF Research Database (Denmark)

    Matchkov, Vladimir

    were used as a model for electrical coupling of SMCs by measuring membrane capacitance (Cm). SMCs were uncoupled (evaluated by inhibition of vasomotion and desynchronization of calcium transients in vascular wall, or by reduction to half of Cm measured in paired A7r5 cells) when the Na......  Ouabain, a specific inhibitor of the Na,K-pump, has previously been shown to interfere with intercellular communication. Here we test the hypothesis that the communication between vascular smooth muscle cells (SMCs) is regulated through an interaction between the Na,K-pump and the Na......,Ca-exchanger leading to an increase in the intracellular calcium concentration in discrete areas near the plasma membrane. The intracellular calcium concentration in individual SMCs was imaged in cultured rat aortic SMCs (A7r5) and simultaneously with isometric force in rat mesenteric small arteries. Paired A7r5 cells...

  10. Xenobiotic Modulation of Human Mammary Epithelial Cell Gap Junctional Intercellular Communication and Growth

    National Research Council Canada - National Science Library

    Ruch, Randall

    1999-01-01

    .... These agents also inhibit gap junctional intercellular communication (GJIC). This inhibition may contribute to the enhancement of breast epithelial growth and breast cancer formation by xenobiotics...

  11. Inhibition of hepatocyte gap junctional intercellular communication by tumor promoters

    International Nuclear Information System (INIS)

    Ruch, R.J.

    1988-01-01

    The mechanisms by which tumor promoters enhance neoplasia are poorly understood. One effect common to most tumor promoters is their ability to inhibit the cell-to-cell exchange of small molecules and ions through gap junctions, i.e., gap junctional intercellular communication (IC). IC maybe necessary for normal growth control and the loss of IC may predispose cells to enhanced growth. In the present studies, the effects of liver tumor promoters and other agents on IC between rodent hepatocytes in primary culture has been studied. IC was detected between hepatocytes: (1) autoradiographically following the passage and incorporation of [5- 3 H]uridine nucleotides from pre-labeled donor hepatocytes to non-labeled, adjacent recipient hepatocytes and (2) by fluorescence microscopy after microinjection of fluorescent Lucifer Yellow CH dye into hepatocytes and visualizing dye spread into adjacent hepatocytes

  12. 7-Ketocholesterol modulates intercellular communication through gap-junction in bovine lens epithelial cells

    Directory of Open Access Journals (Sweden)

    Pereira Paulo

    2004-06-01

    Full Text Available Abstract Background Connexin43 (Cx43 is an integral membrane protein that forms intercellular channels called gap junctions. Intercellular communication in the eye lens relies on an extensive network of gap junctions essential for the maintenance of lens transparency. The association of Cx43 with cholesterol enriched lipid raft domains was recently demonstrated. The objective of this study is to assess if products of cholesterol oxidation (oxysterols affect gap junction intercellular communication (GJIC. Results Primary cultures of lens epithelial cells (LEC were incubated with 7-ketocholesterol (7-Keto, 25-hydroxycholesterol (25-OH or cholesterol and the subcellular distribution of Cx43 was evaluated by immunofluorescence confocal microscopy. The levels of Cx43 present in gap junction plaques were assessed by its insolubility in Triton X-100 and quantified by western blotting. The stability of Cx43 at the plasma membrane following incubation with oxysterols was evaluated by biotinylation of cell surface proteins. Gap junction intercellular communication was evaluated by transfer of the dye Lucifer yellow. The results obtained showed that 7-keto induces an accumulation of Cx43 at the plasma membrane and an increase in intercellular communication through gap junction. However, incubation with cholesterol or 25-OH did not lead to significant alterations on subcellular distribution of Cx43 nor in intercellular communication. Data further suggests that increased intercellular communication results from increased stability of Cx43 at the plasma membrane, presumably forming functional gap-junctions, as suggested by decreased solubility of Cx43 in 1% Triton X-100. The increased stability of Cx43 at the plasma membrane seems to be specific and not related to disruption of endocytic pathway, as demonstrated by dextran uptake. Conclusions Results demonstrate, for the first time, that 7-keto induces an increase in gap junction intercellular communication

  13. Quantification of gap junctional intercellular communication based on digital image analysis

    DEFF Research Database (Denmark)

    Hofgaard, Johannes P; Mollerup, Sarah; Holstein-Rathlou, Niels-Henrik

    2009-01-01

    Intercellular communication via gap junction channels can be quantified by several methods based on diffusion of fluorescent dyes or metabolites. Given the variation in intercellular coupling of cells, even under untreated control conditions, it is of essence to quantify the coupling between...... and in cells receiving dye by intercellular diffusion. The analysis performed is semiautomatic, and comparison with traditional cell counting shows that this method reliably determines the effect of uncoupling by several interventions. This new method of analysis yields a rapid and objective quantification...

  14. Is there a relationship between hypoxia, contact resistance, and intercellular communication

    International Nuclear Information System (INIS)

    Dertinger, H.; Guichard, M.; Malaise, E.P.

    1983-01-01

    This investigation addresses the shape of radiation survival curves of cells cultures as multicell spheroids. It is shown that spheroids of cells capable of intercellular communication by gap-junctions display survival curves lacking a radioresistant fraction of hypoxic cells. Compared to the corresponding monolayers, these spheriod survival curves exhibit a uniform increase in radioresistance due to the ''contact effect''. In contrast, biphasic survival curves indicative of hypoxic cells are obtained with non-communicating spheroids, however, without indication of a contact effect. Evidence is presented that this relationship between intercellular communication, hypoxia, and contact effect may possibly also hold for survival curves of solid tumors. (orig.)

  15. Coronary Heart Disease Alters Intercellular Communication by Modifying Microparticle-Mediated MicroRNA Transport

    Science.gov (United States)

    Finn, Nnenna A.; Eapen, Danny; Manocha, Pankaj; Kassem, Hatem Al; Lassegue, Bernard; Ghasemzadeh, Nima; Quyyumi, Arshed; Searles, Charles D.

    2013-01-01

    Coronary heart disease (CHD) is characterized by abnormal intercellular communication and circulating microRNAs (miRNAs) are likely involved in this process. Here, we show that CHD was associated with changes in the transport of circulating miRNA, particularly decreased miRNA enrichment in microparticles (MPs). Additionally, MPs from CHD patients were less efficient at transferring miRNA to cultured HUVECs, which correlated with their diminished capacity to bind developmental endothelial locus-1 (Del-1). In summary, CHD was associated with distinct changes in circulating miRNA transport and these changes may contribute to the abnormal intercellular communication that underlies CHD initiation and progression. PMID:24042051

  16. [Inhibition of gap junctional intercellular communication protects astrocytes from hypoxia/reoxygenation injury].

    Science.gov (United States)

    Tong, Xu-Hui; Gu, Yu-Chen; Jiao, Hao; Yu, Li; Dong, Shu-Ying

    2015-01-01

    To investigate the effects of inhibiting gap junctional intercellular communication on hypoxia/reoxygenation injury in astrocytes. Primary cultured cerebral cortical astrocytes of neonate rats were divided into normal control group, hypoxia reoxygenation injury group and 18-α-glycyrrhetinic acid and oleamide (gap junctional intercellular channel inhibitors) group. The gap junction intercellular communication was determined by Parachute assay. The viability of astrocyes was detected by MTT assay. The apoptosis of astrocytes were detected with annexin V/PI and Hoechst 33258 staining. Compared with the normal control group, the gap junctional function of astrocytes was increased significantly in ischemia/reperfusion group (Pastrocytes decreased significantly (Pastrocytes in18-α-glycyrrhetinic acid and oleamide group decreased significantly (Pastrocytes increased significantly (Pastrocytes.

  17. Chemopreventive agents attenuate rapid inhibition of gap junctional intercellular communication induced by environmental toxicants

    Czech Academy of Sciences Publication Activity Database

    Babica, Pavel; Čtveráčková, Lucie; Lenčešová, Zuzana; Trosko, J. E.; Upham, B. L.

    2016-01-01

    Roč. 68, č. 5 (2016), s. 827-837 ISSN 0163-5581 R&D Projects: GA MŠk LH12034 Institutional support: RVO:67985939 Keywords : gap junctional intercellular communication * chemopreventive agents * environmental toxicants Subject RIV: FR - Pharmacology ; Medidal Chemistry Impact factor: 2.447, year: 2016

  18. Connexin-based intercellular communication and astrocyte heterogeneity.

    Science.gov (United States)

    Theis, Martin; Giaume, Christian

    2012-12-03

    This review gives an overview of the current knowledge on connexin-mediated communication in astrocytes, covering gap junction and hemichannel functions mediated by connexins. Astroglia is the main brain cell type that expresses the largest amount of connexin and exhibits high level of gap junctional communication compared to neurons and oligodendrocytes. However, in certain developmental and regional situations, astrocytes are also coupled with oligodendrocytes and neurons. This heterotypic coupling is infrequent and minor in terms of extent of the coupling area, which does not mean that it is not important in terms of cell interaction. Here, we present an update on heterogeneity of connexin expression and function at the molecular, subcellular, cellular and networking levels. Interestingly, while astrocytes were initially considered as a homogenous population, there is now increasing evidence for morphological, developmental, molecular and physiological heterogeneity of astrocytes. Consequently, the specificity of gap junction channel- and hemichannel-mediated communication, which tends to synchronize cell populations, is also a parameter to take into account when neuroglial interactions are investigated. This article is part of a Special Issue entitled Electrical Synapses. Copyright © 2012 Elsevier B.V. All rights reserved.

  19. Effect of retinol and cigarette-smoke and condensate on dye-coupled intercellular communication between hamster tracheal epithelial cells

    NARCIS (Netherlands)

    Rutten, A.A.J.J.L.; Jongen, W.M.F.; Haan, L.H.J.de; Hendriksen, E.G.J.; Koeman, J.H.

    1988-01-01

    The dye-coupled intercellular communication across gap junctions in primary hamster tracheal epithelial cells has been studied in serum-free, hormone-supplemented medium. In the absence of vitamin A, non-cytotoxic concentrations of cigarette-smoke condensate (CSC) inhibited intercellular

  20. Regulation of the Host Antiviral State by Intercellular Communications

    Directory of Open Access Journals (Sweden)

    Sonia Assil

    2015-08-01

    Full Text Available Viruses usually induce a profound remodeling of host cells, including the usurpation of host machinery to support their replication and production of virions to invade new cells. Nonetheless, recognition of viruses by the host often triggers innate immune signaling, preventing viral spread and modulating the function of immune cells. It conventionally occurs through production of antiviral factors and cytokines by infected cells. Virtually all viruses have evolved mechanisms to blunt such responses. Importantly, it is becoming increasingly recognized that infected cells also transmit signals to regulate innate immunity in uninfected neighboring cells. These alternative pathways are notably mediated by vesicular secretion of various virus- and host-derived products (miRNAs, RNAs, and proteins and non-infectious viral particles. In this review, we focus on these newly-described modes of cell-to-cell communications and their impact on neighboring cell functions. The reception of these signals can have anti- and pro-viral impacts, as well as more complex effects in the host such as oncogenesis and inflammation. Therefore, these “broadcasting” functions, which might be tuned by an arms race involving selective evolution driven by either the host or the virus, constitute novel and original regulations of viral infection, either highly localized or systemic.

  1. A transwell assay that excludes exosomes for assessment of tunneling nanotube-mediated intercellular communication.

    Science.gov (United States)

    Thayanithy, Venugopal; O'Hare, Patrick; Wong, Phillip; Zhao, Xianda; Steer, Clifford J; Subramanian, Subbaya; Lou, Emil

    2017-11-13

    Tunneling nanotubes (TNTs) are naturally-occurring filamentous actin-based membranous extensions that form across a wide spectrum of mammalian cell types to facilitate long-range intercellular communication. Valid assays are needed to accurately assess the downstream effects of TNT-mediated transfer of cellular signals in vitro. We recently reported a modified transwell assay system designed to test the effects of intercellular transfer of a therapeutic oncolytic virus, and viral-activated drugs, between cells via TNTs. The objective of the current study was to demonstrate validation of this in vitro approach as a new method for effectively excluding diffusible forms of long- and close-range intercellular transfer of intracytoplasmic cargo, including exosomes/microvesicles and gap junctions in order to isolate TNT-selective cell communication. We designed several steps to effectively reduce or eliminate diffusion and long-range transfer via these extracellular vesicles, and used Nanoparticle Tracking Analysis to quantify exosomes following implementation of these steps. The experimental approach outlined here effectively reduced exosome trafficking by >95%; further use of heparin to block exosome uptake by putative recipient cells further impeded transfer of these extracellular vesicles. This validated assay incorporates several steps that can be taken to quantifiably control for extracellular vesicles in order to perform studies focused on TNT-selective communication.

  2. Novel approaches to mitigating bacterial biofilm formation and intercellular communication

    Science.gov (United States)

    Kasper, Stephen H.

    Long thought of as solitary single-cell organisms, it is now widely accepted that bacteria can act and cooperate as social organisms. Phenomena such as biofilm formation and quorum sensing (QS) are two intimately intertwined cooperative behaviors that significantly contribute to the pathogenesis of many bacteria. Biofilms are surface associated communities of bacteria encased in a secreted extracellular matrix, which provides several advantages over an individualized lifestyle, such as increased protection from antimicrobial agents as well as enhanced opportunity for the exchange of genetic material. Bacterial QS is a system of population-based communication through the production, sensing, and response to chemical signals, often controlling the expression of diverse virulence factors (e.g. toxins, proteases). Biofilm formation and QS are cooperative processes that are often leveraged as bacteria coordinate infection processes, and can therefore be novel targets for anti-infective treatments that differ from conventional antibiotic treatment. Our lab has previously identified a novel class of small molecules that inhibit biofilm formation and disrupt QS by the pathogenic bacterium Pseudomonas aeruginosa. These organosulfur-based compounds are either natural products or related derivatives of the tropical plant Petiveria alliacea. Because oral biofilm (e.g. dental plaque) is a major conduit of oral and systemic disease, and is also a site for horizontal transfer for genes encoding antibiotic resistance, there exists a need for novel strategies for inhibiting oral biofilm development. Therefore, a small library (˜50 compounds) of structural derivatives was developed and screened for their ability to inhibit biofilm formation by multiple orally associated bacteria. The screening effort uncovered several related compounds that inhibited oral biofilm development. To determine how natural product-based organosulfur compounds could be inducing QS inhibitory effects, an

  3. Intercellular communication via gap junctions affected by mechanical load in the bovine annulus fibrosus.

    Science.gov (United States)

    Desrochers, Jane; Duncan, Neil A

    2014-01-01

    Cells in the intervertebral disc, as in other connective tissues including tendon, ligament and bone, form interconnected cellular networks that are linked via functional gap junctions. These cellular networks may be necessary to affect a coordinated response to mechanical and environmental stimuli. Using confocal microscopy with fluorescence recovery after photobleaching methods, we explored the in situ strain environment of the outer annulus of an intact bovine disc and the effect of high-level flexion on gap junction signalling. The in situ strain environment in the extracellular matrix of the outer annulus under high flexion load was observed to be non-uniform with the extensive cellular processes remaining crimped sometimes at flexion angles greater than 25°. A significant transient disruption of intercellular communication via functional gap junctions was measured after 10 and 20 min under high flexion load. This study illustrates that in healthy annulus fibrosus tissue, high mechanical loads can impede the functioning of the gap junctions. Future studies will explore more complex loading conditions to determine whether losses in intercellular communication can be permanent and whether gap junctions in aged and degenerated tissues become more susceptible to load. The current research suggests that cellular structures such as gap junctions and intercellular networks, as well as other cell-cell and cell-matrix interconnections, need to be considered in computational models in order to fully understand how macroscale mechanical signals are transmitted across scales to the microscale and ultimately into a cellular biosynthetic response in collagenous tissues.

  4. INHIBITION OF GAP JUNCTIONAL INTERCELLULAR COMMUNICATION BY PERFLUORINATED COMPOUNDS IN RAT LIVER AND DOLPHIN KIDNEY EPITHELIAL CELL LINES IN VITRO AND SPRAGUE-DAWLEY RATS IN VIVO

    Science.gov (United States)

    Abstract Gap Junctional Intercellular Communication (GJIC) is the major pathway of intercellular signal transduction, and is, thus, important for normal cell growth and function. Recent studies have revealed a global distribution of some perfluorinated organic compounds e...

  5. Cytochrome P450-mediated metabolism of tumour promoters modifies the inhibition of intercellular communication: a modified assay for tumour promotion

    DEFF Research Database (Denmark)

    Vang, Ole; Wallin, H.; Doehmer, J.

    1993-01-01

    The role of metabolism of tumour promoters on the inhibition of intercellular communication was investigated in a modified V79 metabolic cooperation system. V79 cells, which stably express different rat cytochrome P450 enzymes (CYP1A1, CYP1A2 or CYP2B1), were used in the metabolic cooperation assay...... B1 and 4-nitrobiphenyl, did not inhibit metabolic cooperation in either V79 cells expressing or cells not expressing cytochrome P450. We conclude that cytochrome P450-associated metabolism plays an important role in the inhibition of gap junctional intercellular communication of some tumour...... promoters. The modified metabolic cooperation assay presented here is valuable for detecting some inhibitory chemicals which have been 'false negative' in previous assays for gap junctional intercellular communication. The assay also discloses that cytochrome P450 metabolism alters intercellular...

  6. Methylmercury inhibits gap junctional intercellular communication in primary cultures of rat proximal tubular cells

    Energy Technology Data Exchange (ETDEWEB)

    Yoshida, Minoru; Sumi, Yawara [Department of Chemistry, St. Marianna University School of Medicine, Kawasagi (Japan); Kujiraoka, Toru [Department of Physiology, St. Marianna University School of Medicine, Kawasagi (Japan); Hara, Masayuki [Department of Anatomy, St. Marianna University School of Medicine, Kawasagi (Japan); Nakazawa, Hirokazu [Department of Chemistry, Faculty of Sciences, Meisei University (Japan)

    1998-03-01

    Methylmercury (MeHg) causes renal injury in addition to central and peripheral neuropathy. To clarify the mechanism of nephrotoxicity by MeHg, we investigated the effect of this compound on intercellular communication through gap junction channels in primary cultures of rat renal proximal tubular cells. Twenty minutes after exposure to 30 {mu}M MeHg, gap junctional intercellular communication (GJIC), which was assessed by dye coupling, was markedly inhibited before appearance of cytotoxicity. When the medium containing MeHg was exchanged with MeHg-free medium, dye coupling recovered abruptly. However, the dye-coupling was abolished again 30 min after replacement with control medium, and the cells were damaged. Intracellular calcium concentration, [Ca{sup 2+}]{sub i}, which modulates the function of gap junctions, significantly increased following exposure of the cells to 30 {mu}M MeHg and returned to control level following replacement with MeHg-free medium. These results suggest that the inhibiting effect of MeHg on GJIC is related to the change in [Ca{sup 2+}]{sub i}, and may be involved in the pathogenesis of renal dysfunction. (orig.) With 5 figs., 23 refs.

  7. Cell walls as a stage for intercellular communication regulating shoot meristem development

    Directory of Open Access Journals (Sweden)

    Toshiaki eTameshige

    2015-05-01

    Full Text Available Aboveground organs of plants are ultimately derived/generated from the shoot apical meristem (SAM, which is the proliferative tissue located at the apex of the stem. The SAM contains a population of stem cells that provide new cells for organ/tissue formation. The SAM is composed of distinct cell layers and zones with different properties. Primordia of lateral organs develop at the periphery of the SAM. The shoot apex is a dynamic and complex tissue, and as such intercellular communications among cells, layers and zones play significant roles in the coordination of cell proliferation, growth and differentiation to achieve elaborate morphogenesis. Recent findings have highlighted the importance of a number of singling molecules acting in the cell wall space for the intercellular communication, including classic phytohormones and secretory peptides. Moreover, accumulating evidences reveal that cell wall properties and their modifying enzymes modulate hormone actions. In this review, we overview how behaviors of singling molecules and changes of cell wall properties are integrated for the shoot meristem regulation.

  8. Connexin 26-mediated gap junctional intercellular communication suppresses paracellular permeability of human intestinal epithelial cell monolayers

    International Nuclear Information System (INIS)

    Morita, Hidekazu; Katsuno, Tatsuro; Hoshimoto, Aihiro; Hirano, Noriaki; Saito, Yasushi; Suzuki, Yasuo

    2004-01-01

    In some cell types, gap junctional intercellular communication (GJIC) is associated with tight junctions. The present study was performed to determine the roles of GJIC in regulation of the barrier function of tight junctions. Caco-2 human colonic cells were used as a monolayer model, and barrier function was monitored by measuring mannitol permeability and transepithelial electrical resistance (TER). The monolayers were chemically disrupted by treatment with oleic acid and taurocholic acid. Western blotting analyses were performed to evaluate the protein levels of connexins, which are components of gap junctional intercellular channels. Cx26 expression was detected in preconfluent Caco-2 cells, and its level increased gradually after the monolayer reached confluency. These results prompted us to examine whether overexpression of Cx26 affects barrier function. Monolayers of Caco-2 cells stably expressing Cx26 showed significantly lower mannitol permeability and higher TER than mock transfectants when the monolayers were chemically disrupted. The levels of claudin-4, an important component of tight junctions, were significantly increased in the stable Cx26 transfectant. These results suggest that Cx26-mediated GJIC may play a crucial role in enhancing the barrier function of Caco-2 cell monolayers

  9. Intercellular signal communication among odontoblasts and trigeminal ganglion neurons via glutamate.

    Science.gov (United States)

    Nishiyama, A; Sato, M; Kimura, M; Katakura, A; Tazaki, M; Shibukawa, Y

    2016-11-01

    Various stimuli to the exposed surface of dentin induce changes in the hydrodynamic force inside the dentinal tubules resulting in dentinal pain. Recent evidences indicate that mechano-sensor channels, such as the transient receptor potential channels, in odontoblasts receive these hydrodynamic forces and trigger the release of ATP to the pulpal neurons, to generate dentinal pain. A recent study, however, has shown that odontoblasts also express glutamate receptors (GluRs). This implies that cells in the dental pulp tissue have the ability to release glutamate, which acts as a functional intercellular mediator to establish inter-odontoblast and odontoblast-trigeminal ganglion (TG) neuron signal communication. To investigate the intercellular signal communication, we applied mechanical stimulation to odontoblasts and measured the intracellular free Ca 2+ concentration ([Ca 2+ ] i ). During mechanical stimulation in the presence of extracellular Ca 2+ , we observed a transient [Ca 2+ ] i increase not only in single stimulated odontoblasts, but also in adjacent odontoblasts. We could not observe these responses in the absence of extracellular Ca 2+ . [Ca 2+ ] i increases in the neighboring odontoblasts during mechanical stimulation of single odontoblasts were inhibited by antagonists of metabotropic glutamate receptors (mGluRs) as well as glutamate-permeable anion channels. In the odontoblast-TG neuron coculture, we observed an increase in [Ca 2+ ] i in the stimulated odontoblasts and TG neurons, in response to direct mechanical stimulation of single odontoblasts. These [Ca 2+ ] i increases in the neighboring TG neurons were inhibited by antagonists for mGluRs. The [Ca 2+ ] i increases in the stimulated odontoblasts were also inhibited by mGluRs antagonists. We further confirmed that the odontoblasts express group I, II, and III mGluRs. However, we could not record any currents evoked from odontoblasts near the mechanically stimulated odontoblast, with or without

  10. Intercellular communication and cell proliferation in precision-cut rat liver slices : effect of medium composition and DDT

    NARCIS (Netherlands)

    Graaf, I.A.M.; Tajima, O.; Groten, J.P.; Wolterbeek, A.P.M.

    2000-01-01

    Gap junctional intercellular communication (GJIC) and cell proliferation were studied in control and 1,1'-bis(p-chlorophenyl)-2,2,2,-trichloroethane (DDT) treated precision-cut liver slices of rat by evaluating connexin 32 (Cx32) expression and 5-bromo-2'-deoxyuridine (BrdU) incorporation. In

  11. Tunneling Nanotubes and Gap Junctions–Their Role in Long-Range Intercellular Communication during Development, Health, and Disease Conditions

    Directory of Open Access Journals (Sweden)

    Jennifer Ariazi

    2017-10-01

    Full Text Available Cell-to-cell communication is essential for the organization, coordination, and development of cellular networks and multi-cellular systems. Intercellular communication is mediated by soluble factors (including growth factors, neurotransmitters, and cytokines/chemokines, gap junctions, exosomes and recently described tunneling nanotubes (TNTs. It is unknown whether a combination of these communication mechanisms such as TNTs and gap junctions may be important, but further research is required. TNTs are long cytoplasmic bridges that enable long-range, directed communication between connected cells. The proposed functions of TNTs are diverse and not well understood but have been shown to include the cell-to-cell transfer of vesicles, organelles, electrical stimuli and small molecules. However, the exact role of TNTs and gap junctions for intercellular communication and their impact on disease is still uncertain and thus, the subject of much debate. The combined data from numerous laboratories indicate that some TNT mediate a long-range gap junctional communication to coordinate metabolism and signaling, in relation to infectious, genetic, metabolic, cancer, and age-related diseases. This review aims to describe the current knowledge, challenges and future perspectives to characterize and explore this new intercellular communication system and to design TNT-based therapeutic strategies.

  12. Interaction between Na+/K+-pump and Na+/Ca2+-exchanger modulates intercellular communication.

    Science.gov (United States)

    Matchkov, Vladimir V; Gustafsson, Helena; Rahman, Awahan; Briggs Boedtkjer, Donna M; Gorintin, Sarah; Hansen, Anne Kirstine; Bouzinova, Elena V; Praetorius, Helle A; Aalkjaer, Christian; Nilsson, Holger

    2007-04-13

    Ouabain, a specific inhibitor of the Na(+)/K(+)-pump, has previously been shown to interfere with intercellular communication. Here we test the hypothesis that the communication between vascular smooth muscle cells is regulated through an interaction between the Na(+)/K(+)-pump and the Na(+)/Ca(2+)-exchanger leading to an increase in the intracellular calcium concentration ([Ca(2+)](i)) in discrete areas near the plasma membrane. [Ca(2+)](i) in smooth muscle cells was imaged in cultured rat aortic smooth muscle cell pairs (A7r5) and in rat mesenteric small artery segments simultaneously with force. In A7r5 coupling between cells was estimated by measuring membrane capacitance. Smooth muscle cells were uncoupled when the Na(+)/K(+)-pump was inhibited either by a low concentration of ouabain, which also caused a localized increase of [Ca(2+)](i) near the membrane, or by ATP depletion. Reduction of Na(+)/K(+)-pump activity by removal of extracellular potassium ([K(+)](o)) also uncoupled cells, but only after inhibition of K(ATP) channels. Inhibition of the Na(+)/Ca(2+)-exchange activity by SEA0400 or by a reduction of the equilibrium potential (making it more negative) also uncoupled the cells. Depletion of intracellular Na(+) and clamping of [Ca(2+)](i) at low concentrations prevented the uncoupling. The experiments suggest that the Na(+)/K(+)-pump may affect gap junction conductivity via localized changes in [Ca(2+)](i) through modulation of Na(+)/Ca(2+)-exchanger activity.

  13. Transient suppression of gap junctional intercellular communication after exposure to 100-nanosecond pulsed electric fields.

    Science.gov (United States)

    Steuer, Anna; Schmidt, Anke; Labohá, Petra; Babica, Pavel; Kolb, Juergen F

    2016-12-01

    Gap junctional intercellular communication (GJIC) is an important mechanism that is involved and affected in many diseases and injuries. So far, the effect of nanosecond pulsed electric fields (nsPEFs) on the communication between cells was not investigated. An in vitro approach is presented with rat liver epithelial WB-F344 cells grown and exposed in a monolayer. In order to observe sub-lethal effects, cells were exposed to pulsed electric fields with a duration of 100ns and amplitudes between 10 and 20kV/cm. GJIC strongly decreased within 15min after treatment but recovered within 24h. Gene expression of Cx43 was significantly decreased and associated with a reduced total amount of Cx43 protein. In addition, MAP kinases p38 and Erk1/2, involved in Cx43 phosphorylation, were activated and Cx43 became hyperphosphorylated. Immunofluorescent staining of Cx43 displayed the disassembly of gap junctions. Further, a reorganization of the actin cytoskeleton was observed whereas tight junction protein ZO-1 was not significantly affected. All effects were field- and time-dependent and most pronounced within 30 to 60min after treatment. A better understanding of a possible manipulation of GJIC by nsPEFs might eventually offer a possibility to develop and improve treatments. Copyright © 2016 Elsevier B.V. All rights reserved.

  14. Chlorpromazine reduces the intercellular communication via gap junctions in mammalian cells

    International Nuclear Information System (INIS)

    Orellana, Juan A.; Palacios-Prado, Nicolas; Saez, Juan C.

    2006-01-01

    In the work presented herein, we evaluated the effect of chlorpromazine (CPZ) on gap junctions expressed by two mammalian cell types; Gn-11 cells (cell line derived from mouse LHRH neurons) and rat cortical astrocytes maintained in culture. We also attempted to elucidate possible mechanisms of action of CPZ effects on gap junctions. CPZ, in concentrations comparable with doses used to treat human diseases, was found to reduce the intercellular communication via gap junctions as evaluated with measurements of dye coupling (Lucifer yellow). In both cell types, maximal inhibition of functional gap junctions was reached within about 1 h of treatment with CPZ, an recovery was almost complete at about 5 h after CPZ wash out. In both cell types, CPZ treatment increased the phosphorylation state of connexin43 (Cx43), a gap junction protein subunit. Moreover, CPZ reduced the reactivity of Cx43 (immunofluorescence) at cell interfaces and concomitantly increased its reactivity in intracellular vesicles, suggesting an increased retrieval from and/or reduced insertion into the plasma membrane. CPZ also caused cellular retraction reducing cell-cell contacts in a reversible manner. The reduction in contact area might destabilize existing gap junctions and abrogate formation of new ones. Moreover, the CPZ-induced reduction in gap junctional communication may depend on the connexins (Cxs) forming the junctions. If Cx43 were the only connexin expressed, MAPK-dependent phosphorylation of this connexin would induce closure of gap junction channels

  15. Proteome-metabolome profiling of ovarian cancer ascites reveals novel components involved in intercellular communication.

    Science.gov (United States)

    Shender, Victoria O; Pavlyukov, Marat S; Ziganshin, Rustam H; Arapidi, Georgij P; Kovalchuk, Sergey I; Anikanov, Nikolay A; Altukhov, Ilya A; Alexeev, Dmitry G; Butenko, Ivan O; Shavarda, Alexey L; Khomyakova, Elena B; Evtushenko, Evgeniy; Ashrafyan, Lev A; Antonova, Irina B; Kuznetcov, Igor N; Gorbachev, Alexey Yu; Shakhparonov, Mikhail I; Govorun, Vadim M

    2014-12-01

    Ovarian cancer ascites is a native medium for cancer cells that allows investigation of their secretome in a natural environment. This medium is of interest as a promising source of potential biomarkers, and also as a medium for cell-cell communication. The aim of this study was to elucidate specific features of the malignant ascites metabolome and proteome. In order to omit components of the systemic response to ascites formation, we compared malignant ascites with cirrhosis ascites. Metabolome analysis revealed 41 components that differed significantly between malignant and cirrhosis ascites. Most of the identified cancer-specific metabolites are known to be important signaling molecules. Proteomic analysis identified 2096 and 1855 proteins in the ovarian cancer and cirrhosis ascites, respectively; 424 proteins were specific for the malignant ascites. Functional analysis of the proteome demonstrated that the major differences between cirrhosis and malignant ascites were observed for the cluster of spliceosomal proteins. Additionally, we demonstrate that several splicing RNAs were exclusively detected in malignant ascites, where they probably existed within protein complexes. This result was confirmed in vitro using an ovarian cancer cell line. Identification of spliceosomal proteins and RNAs in an extracellular medium is of particular interest; the finding suggests that they might play a role in the communication between cancer cells. In addition, malignant ascites contains a high number of exosomes that are known to play an important role in signal transduction. Thus our study reveals the specific features of malignant ascites that are associated with its function as a medium of intercellular communication. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  16. Mammalian Tissue Response to Low Dose Ionizing Radiation: The Role of Oxidative Metabolism and Intercellular Communication

    Energy Technology Data Exchange (ETDEWEB)

    Azzam, Edouard I

    2013-01-16

    The objective of the project was to elucidate the mechanisms underlying the biological effects of low dose/low dose rate ionizing radiation in organs/tissues of irradiated mice that differ in their susceptibility to ionizing radiation, and in human cells grown under conditions that mimic the natural in vivo environment. The focus was on the effects of sparsely ionizing cesium-137 gamma rays and the role of oxidative metabolism and intercellular communication in these effects. Four Specific Aims were proposed. The integrated outcome of the experiments performed to investigate these aims has been significant towards developing a scientific basis to more accurately estimate human health risks from exposures to low doses ionizing radiation. By understanding the biochemical and molecular changes induced by low dose radiation, several novel markers associated with mitochondrial functions were identified, which has opened new avenues to investigate metabolic processes that may be affected by such exposure. In particular, a sensitive biomarker that is differentially modulated by low and high dose gamma rays was discovered.

  17. Extract from the Zooxanthellate Jellyfish Cotylorhiza tuberculata Modulates Gap Junction Intercellular Communication in Human Cell Cultures

    Directory of Open Access Journals (Sweden)

    Stefano Piraino

    2013-05-01

    Full Text Available On a global scale, jellyfish populations in coastal marine ecosystems exhibit increasing trends of abundance. High-density outbreaks may directly or indirectly affect human economical and recreational activities, as well as public health. As the interest in biology of marine jellyfish grows, a number of jellyfish metabolites with healthy potential, such as anticancer or antioxidant activities, is increasingly reported. In this study, the Mediterranean “fried egg jellyfish” Cotylorhiza tuberculata (Macri, 1778 has been targeted in the search forputative valuable bioactive compounds. A medusa extract was obtained, fractionated, characterized by HPLC, GC-MS and SDS-PAGE and assayed for its biological activity on breast cancer cells (MCF-7 and human epidermal keratinocytes (HEKa. The composition of the jellyfish extract included photosynthetic pigments, valuable ω-3 and ω-6 fatty acids, and polypeptides derived either from jellyfish tissues and their algal symbionts. Extract fractions showed antioxidant activity and the ability to affect cell viability and intercellular communication mediated by gap junctions (GJIC differentially in MCF-7and HEKa cells. A significantly higher cytotoxicity and GJIC enhancement in MCF-7 compared to HEKa cells was recorded. A putative action mechanism for the anticancer bioactivity through the modulation of GJIC has been hypothesized and its nutraceutical and pharmaceutical potential was discussed.

  18. TC-PTP directly interacts with connexin43 to regulate gap junction intercellular communication

    Science.gov (United States)

    Li, Hanjun; Spagnol, Gaelle; Naslavsky, Naava; Caplan, Steve; Sorgen, Paul L.

    2014-01-01

    ABSTRACT Protein kinases have long been reported to regulate connexins; however, little is known about the involvement of phosphatases in the modulation of intercellular communication through gap junctions and the subsequent downstream effects on cellular processes. Here, we identify an interaction between the T-cell protein tyrosine phosphatase (TC-PTP, officially known as PTPN2) and the carboxyl terminus of connexin43 (Cx43, officially known as GJA1). Two cell lines, normal rat kidney (NRK) cells endogenously expressing Cx43 and an NRK-derived cell line expressing v-Src with temperature-sensitive activity, were used to demonstrate that EGF and v-Src stimulation, respectively, induced TC-PTP to colocalize with Cx43 at the plasma membrane. Cell biology experiments using phospho-specific antibodies and biophysical assays demonstrated that the interaction is direct and that TC-PTP dephosphorylates Cx43 residues Y247 and Y265, but does not affect v-Src. Transfection of TC-PTP also indirectly led to the dephosphorylation of Cx43 S368, by inactivating PKCα and PKCδ, with no effect on the phosphorylation of S279 and S282 (MAPK-dependent phosphorylation sites). Dephosphorylation maintained Cx43 gap junctions at the plaque and partially reversed the channel closure caused by v-Src-mediated phosphorylation of Cx43. Understanding dephosphorylation, along with the well-documented roles of Cx43 phosphorylation, might eventually lead to methods to modulate the regulation of gap junction channels, with potential benefits for human health. PMID:24849651

  19. Estrogenic compounds inhibit gap junctional intercellular communication in mouse Leydig TM3 cells

    International Nuclear Information System (INIS)

    Iwase, Yumiko; Fukata, Hideki; Mori, Chisato

    2006-01-01

    Some estrogenic compounds are reported to cause testicular disorders in humans and/or experimental animals by direct action on Leydig cells. In carcinogenesis and normal development, gap junctional intercellular communication (GJIC) plays an essential role in maintaining homeostasis. In this study, we examine the effects of diethylstilbestrol (DES, a synthetic estrogen), 17β-estradiol (E 2 , a natural estrogen), and genistein (GEN, a phytoestrogen) on GJIC between mouse Leydig TM3 cells using Lucifer yellow microinjection. The three compounds tested produced GJIC inhibition in the TM3 cells after 24 h. Gradually, 10 μM DES began to inhibit GJIC for 24 h and this effect was observed until 72 h. On the other hand, both 20 μM E 2 and 25 μM GEN rapidly inhibited GJIC in 6 h and 2 h, respectively. The effects continued until 24 h, but weakened by 72 h. Furthermore, a combined effect at μM level between DES and E 2 on GJIC inhibition was observed, but not between GEN and E 2 . DES and E 2 showed GJIC inhibition at low dose levels (nearly physiological estrogen levels) after 72 h, but GEN did not. DES-induced GJIC inhibition at 10 pM and 10 μM was completely counteracted by ICI 182,780 (ICl), an estrogen receptor antagonist. On the other hand, the inhibitory effects on GJIC with E 2 (10 pM and 20 μM) and GEN (25 μM) were partially blocked by ICI or calphostin C, a protein kinase C (PKC) inhibitor, and were completely blocked by the combination of ICI and calphostin C. These results demonstrate that DES inhibits GJIC between Leydig cells via the estrogen receptor (ER), and that E 2 and GEN inhibit GJIC via ER and PKC. These estrogenic compounds may have different individual nongenotoxic mechanism including PKC pathway on testicular carcinogenesis or development

  20. Use of cultured cells with defects of citrulline metabolism in diagnosis and in the study of intercellular communication

    Energy Technology Data Exchange (ETDEWEB)

    Davidson, J S

    1985-01-01

    Citrullinemia and argininosuccinic aciduria are two disorders resulting from defects in two consecutive enzymes of the urea cycle, argininosuccinate synthetase and argininosuccinate lyase. Fibroblast cell lines were derived from patients with these disorders and the diagnoses, which had been made on the basis of amino acid levels in plasma and urine, were confirmed by demonstrating that the cell lines were unable to incorporate /sup 14/C-citrulline into protein. DNA from the argininosuccinate synthetase-deficient (ASS-) cells was analysed by restriction enzyme digestion and hybridisation to a cDNA probe which had been cloned from human argininosuccinate synthetase mRNA. No defect in the patient's DNA could be demonstrated, indicating that no major deletions in the argininosuccinate synthetase genes were present in this patient. Co-cultures of the ASS- and argininosuccinate lyase-deficient (ASL-) fibroblasts were able to incorporate /sup 14/C-citrulline into protein. Co-cultures of ASS- and ASL-cells were used as an assay system for measuring intercellular junctional communication. This allowed quantitation of the effects of pH and extra-cellular divalent cations on junctional communication. Tumor promoters such as phorbol esters and organochlorine pesticides have been reported to inhibit intercellular junctional communication in other systems, and this inhibitory activity may be related to the mechanism of tumor promotion. Retinoic acid and other retinoids also inhibited junctional communication, and the inhibitory effects of retinoic acid and TPA were additive. It is concluded that co-cultures of ASS- and ASL-cells constitute a useful system for providing quantitative measurements of intercellular junctional communication under a wide range of experimental conditions.

  1. The use of cultured cells with defects of citrulline metabolism in diagnosis and in the study of intercellular communication

    International Nuclear Information System (INIS)

    Davidson, J.S.

    1985-02-01

    Citrullinemia and argininosuccinic aciduria are two disorders resulting from defects in two consecutive enzymes of the urea cycle, argininosuccinate synthetase and argininosuccinate lyase. Fibroblast cell lines were derived from patients with these disorders and the diagnoses, which had been made on the basis of amino acid levels in plasma and urine, were confirmed by demonstrating that the cell lines were unable to incorporate 14 C-citrulline into protein. DNA from the argininosuccinate synthetase-deficient (ASS-) cells was analysed by restriction enzyme digestion and hybridisation to a cDNA probe which had been cloned from human argininosuccinate synthetase mRNA. No defect in the patient's DNA could be demonstrated, indicating that no major deletions in the argininosuccinate synthetase genes were present in this patient. Co-cultures of the ASS- and argininosuccinate lyase-deficient (ASL-) fibroblasts were able to incorporate 14 C-citrulline into protein. Co-cultures of ASS- and ASL-cells were used as an assay system for measuring intercellular junctional communication. This allowed quantitation of the effects of pH and extra-cellular divalent cations on junctional communication. Tumor promoters such as phorbol esters and organochlorine pesticides have been reported to inhibit intercellular junctional communication in other systems, and this inhibitory activity may be related to the mechanism of tumor promotion. Retinoic acid and other retinoids also inhibited junctional communication, and the inhibitory effects of retinoic acid and TPA were additive. It is concluded that co-cultures of ASS- and ASL-cells constitute a useful system for providing quantitative measurements of intercellular junctional communication under a wide range of experimental conditions

  2. Intercellular odontoblast communication via ATP mediated by pannexin-1 channel and phospholipase C-coupled receptor activation.

    Directory of Open Access Journals (Sweden)

    Masaki eSato

    2015-11-01

    Full Text Available Extracellular ATP released via pannexin-1 channels, in response to the activation of mechanosensitive-TRP channels during odontoblast mechanical stimulation, mediates intercellular communication among odontoblasts in dental pulp slice preparation dissected form rat incisor. Recently, odontoblast cell lines, such as mouse odontoblast lineage cells, have been widely used to investigate physiological/pathological cellular functions. To clarify whether the odontoblast cell lines also communicate with each other by diffusible chemical substance(s, we investigated the chemical intercellular communication among cells from mouse odontoblast cell lines following mechanical stimulation. A single cell was stimulated using a glass pipette filled with standard extracellular solution. We measured intracellular free Ca2+ concentration ([Ca2+]i by fura-2 in stimulated cells, as well as in cells located nearby. Direct mechanical stimulation to a single odontoblast increased [Ca2+]i, which showed sensitivity to capsazepine. In addition, we observed increases in [Ca2+]i not only in the mechanically stimulated odontoblast, but also in nearby odontoblasts. We could observe mechanical stimulation-induced increase in [Ca2+]i in a stimulated human embryo kidney (HEK 293 cell, but not in nearby HEK293 cells. The increase in [Ca2+]i in nearby odontoblasts, but not in the stimulated odontoblast, was inhibited by adenosine triphosphate (ATP release channel (pannexin-1 inhibitor in a concentration- and spatial-dependent manner. Moreover, in the presence of phospholipase C (PLC inhibitor, the increase in [Ca2+]i in nearby odontoblasts, following mechanical stimulation of a single odontoblast, was abolished. We could record some inward currents evoked from odontoblasts near the stimulated odontoblast, but the currents were observed in only 4.8% of the recorded odontoblasts. The results of this study showed that ATP is released via pannexin-1, from a mechanically stimulated

  3. Participation of intercellular communication and intracellular signal transduction in the radio-adaptive response of human fibroblastic cells

    International Nuclear Information System (INIS)

    Ishii, Keiichiro; Hoshi, Yuko; Iwasaki, Toshiyasu; Watanabe, Masami

    1997-01-01

    To investigate the radio-adaptive response of normal cells to low-dose radiation, we irradiated human embryonic cells with low-dose X-rays and examined the changes in sensitivity to subsequent high-dose X-irradiation. When the cells were irradiated by 200 cGy, the growth ratio of the viable cells five days after the irradiation decreased to 37% of that of the cells which received no X-irradiation. When the cells received a conditioning irradiation of 10 to 20 cGy four hours before the irradiation of 200 cGy, the growth ratio increased significantly to 45-53%, and a peak was reached at a conditioning dose of 13 cGy. Cells blocked off intercellular communication either in Ca 2+ ion-free medium or in TPA added medium during the conditioning irradiation of 13 cGy did not show the improvement of growth ratio. Addition of H-7, as an inhibitor of PKC, to the medium during the conditioning irradiation inhibited the induction of the radio-adaptive response. However, addition of either inhibitor of A kinase, H-89, or inhibitor of G kinase, H-8, failed to inhibit the induction of the radio-adaptive response. These results suggest that: (1) normal cells show an adaptive response to low-dose radiation, (2) intercellular communication may play a role in radio-adaptive responses, (3) the transduction of the signal induced in cells by low-dose X-irradiation via protein kinase C was involved in radio-adaptive responses, not via A kinase nor G kinase. (author)

  4. Modulatory effects of cAMP and PKC activation on gap junctional intercellular communication among thymic epithelial cells

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    Neves-dos-Santos Sandra

    2010-01-01

    Full Text Available Abstract Background We investigated the effects of the signaling molecules, cyclic AMP (cAMP and protein-kinase C (PKC, on gap junctional intercellular communication (GJIC between thymic epithelial cells (TEC. Results Treatment with 8-Br-cAMP, a cAMP analog; or forskolin, which stimulates cAMP production, resulted in an increase in dye transfer between adjacent TEC, inducing a three-fold enhancement in the mean fluorescence of coupled cells, ascertained by flow cytometry after calcein transfer. These treatments also increased Cx43 mRNA expression, and stimulated Cx43 protein accumulation in regions of intercellular contacts. VIP, adenosine, and epinephrine which may also signal through cyclic nucleotides were tested. The first two molecules did not mimic the effects of 8-Br-cAMP, however epinephrine was able to increase GJIC suggesting that this molecule functions as an endogenous inter-TEC GJIC modulators. Stimulation of PKC by phorbol-myristate-acetate inhibited inter-TEC GJIC. Importantly, both the enhancing and the decreasing effects, respectively induced by cAMP and PKC, were observed in both mouse and human TEC preparations. Lastly, experiments using mouse thymocyte/TEC heterocellular co-cultures suggested that the presence of thymocytes does not affect the degree of inter-TEC GJIC. Conclusions Overall, our data indicate that cAMP and PKC intracellular pathways are involved in the homeostatic control of the gap junction-mediated communication in the thymic epithelium, exerting respectively a positive and negative role upon cell coupling. This control is phylogenetically conserved in the thymus, since it was seen in both mouse and human TEC preparations. Lastly, our work provides new clues for a better understanding of how the thymic epithelial network can work as a physiological syncytium.

  5. Distinct moieties underlie biphasic H+ gating of connexin43 channels, producing a pH optimum for intercellular communication

    Science.gov (United States)

    Garciarena, Carolina D.; Malik, Akif; Swietach, Pawel; Moreno, Alonso P.; Vaughan-Jones, Richard D.

    2018-01-01

    Most mammalian cells can intercommunicate via connexin-assembled, gap-junctional channels. To regulate signal transmission, connexin (Cx) channel permeability must respond dynamically to physiological and pathophysiological stimuli. One key stimulus is intracellular pH (pHi), which is modulated by a tissue’s metabolic and perfusion status. Our understanding of the molecular mechanism of H+ gating of Cx43 channels—the major isoform in the heart and brain—is incomplete. To interrogate the effects of acidic and alkaline pHi on Cx43 channels, we combined voltage-clamp electrophysiology with pHi imaging and photolytic H+ uncaging, performed over a range of pHi values. We demonstrate that Cx43 channels expressed in HeLa or N2a cell pairs are gated biphasically by pHi via a process that consists of activation by H+ ions at alkaline pHi and inhibition at more acidic pHi. For Cx43 channel–mediated solute/ion transmission, the ensemble of these effects produces a pHi optimum, near resting pHi. By using Cx43 mutants, we demonstrate that alkaline gating involves cysteine residues of the C terminus and is independent of motifs previously implicated in acidic gating. Thus, we present a molecular mechanism by which cytoplasmic acid–base chemistry fine tunes intercellular communication and establishes conditions for the optimal transmission of solutes and signals in tissues, such as the heart and brain.—Garciarena, C. D., Malik, A., Swietach, P., Moreno, A. P., Vaughan-Jones, R. D. Distinct moieties underlie biphasic H+ gating of connexin43 channels, producing a pH optimum for intercellular communication. PMID:29183963

  6. Studies of Bystander Effect and Intercellular Communication in Human Epithelial Cell Cultures Irradiated with X-rays

    International Nuclear Information System (INIS)

    Romppanen, E.; Trott, K. R.; Musatonen, R.; Leszcznski, D.; Belyakov, O.

    2004-01-01

    The bystander effect is a phenomenon whereby biological consequences of irradiation are expressed in nonexposed cells in the vicinity of exposed cells. Two main pathways have been proposed to mediate the bystander effect: Gap Junction Intercellular Communication (GJIC) and medium borne soluble factors dependent mechanisms. The present study was designed to evaluate the relative contributions of gap junction intercellular communication and of soluble extracellular factors on the bystander effects of low dose X-ray irradiation. HaCaT human epithelial cell monolayers were exposed to X-ray using specially constructed shield, which cover 95% or 56% or 0% of the cells from the radiation. To evaluate whether the GJIC is involved in transmission of the bystander signal from irradiated to nonirradiated cells, irradiations were performed in presence or absence of GJIC inhibitor lindane. The cytochalasin B block technique was used to quantify fractions of micronucleated cells 48 hours after the irradiation. Our results suggest that more micronucleated cells are induced in partially shielded monolayers than expected according to back extrapolation of the data from open field irradiation. Treatment with lindane considerably reduced amount of the bystander damage. We demonstrated that fraction of micronucleated cells after X-rays irradiation of 5% of cells with 1 Gy was 0.07±0.08 (without lindane) and 0.05±0.004 (in presence of lindane). Irradiation of 100% of cells with the same dose resulted in 0.023±0.04 /without lindane) and 0.013±0.02 (in presence of lindane) fractions of micronucleated cells. Comparison with open field data showed that the fraction of micronucleated cells after irradiation of 5% of the cell culture was 5-10 times greater than the estimated fraction assuming no bystander effect. Irradiation of 44% of cells ded not demonstrate a pronounced bystander effect. (Author) 20 refs

  7. Gap junction intercellular communication mediated by connexin43 in astrocytes is essential for their resistance to oxidative stress.

    Science.gov (United States)

    Le, Hoa T; Sin, Wun Chey; Lozinsky, Shannon; Bechberger, John; Vega, José Luis; Guo, Xu Qiu; Sáez, Juan C; Naus, Christian C

    2014-01-17

    Oxidative stress induced by reactive oxygen species (ROS) is associated with various neurological disorders including aging, neurodegenerative diseases, as well as traumatic and ischemic insults. Astrocytes have an important role in the anti-oxidative defense in the brain. The gap junction protein connexin43 (Cx43) forms intercellular channels as well as hemichannels in astrocytes. In the present study, we investigated the contribution of Cx43 to astrocytic death induced by the ROS hydrogen peroxide (H2O2) and the mechanism by which Cx43 exerts its effects. Lack of Cx43 expression or blockage of Cx43 channels resulted in increased ROS-induced astrocytic death, supporting a cell protective effect of functional Cx43 channels. H2O2 transiently increased hemichannel activity, but reduced gap junction intercellular communication (GJIC). GJIC in wild-type astrocytes recovered after 7 h, but was absent in Cx43 knock-out astrocytes. Blockage of Cx43 hemichannels incompletely inhibited H2O2-induced hemichannel activity, indicating the presence of other hemichannel proteins. Panx1, which is predicted to be a major hemichannel contributor in astrocytes, did not appear to have any cell protective effect from H2O2 insults. Our data suggest that GJIC is important for Cx43-mediated ROS resistance. In contrast to hypoxia/reoxygenation, H2O2 treatment decreased the ratio of the hypophosphorylated isoform to total Cx43 level. Cx43 has been reported to promote astrocytic death induced by hypoxia/reoxygenation. We therefore speculate the increase in Cx43 dephosphorylation may account for the facilitation of astrocytic death. Our findings suggest that the role of Cx43 in response to cellular stress is dependent on the activation of signaling pathways leading to alteration of Cx43 phosphorylation states.

  8. Heavy-ion-induced bystander killing of human lung cancer cells. Role of gap junctional intercellular communication

    International Nuclear Information System (INIS)

    Harada, Kosaku; Nonaka, Tetsuo; Hamada, Nobuyuki; Sakurai, Hideyuki; Hasegawa, Masatoshi; Kobayashi, Yasuhiko; Nakano, Takashi; Funayama, Tomoo; Kakizaki, Takehiko

    2009-01-01

    The aim of the present study was to clarify the mechanisms of cell death induced by heavy-ion irradiation focusing on the bystander effect in human lung cancer A549 cells. In microbeam irradiation, each of 1, 5, and 25 cells under confluent cell conditions was irradiated with 1, 5, or 10 particles of carbon ions (220 MeV), and then the surviving fraction of the population was measured by a clonogenic assay in order to investigate the bystander effect of heavy-ions. In this experiment, the limited number of cells (0.0001-0.002%, 5-25 cells) under confluent cell conditions irradiated with 5 or 10 carbon ions resulted in an exaggerated 8-14% increase in cell death by clonogenic assay. However, these overshooting responses were not observed under exponentially growing cell conditions. Furthermore, these responses were inhibited in cells treated with an inhibitor of gap junctional intercellular communication (GJIC), whereas they were markedly enhanced by the addition of a stimulator of GJIC. The present results suggest that bystander cell killing by heavy-ions was induced mainly by direct cell-to-cell communication, such as GJIC, which might play important roles in bystander responses. (author)

  9. Stat3 is a positive regulator of gap junctional intercellular communication in cultured, human lung carcinoma cells

    Directory of Open Access Journals (Sweden)

    Geletu Mulu

    2012-12-01

    Full Text Available Abstract Background Neoplastic transformation of cultured cells by a number of oncogenes such as src suppresses gap junctional, intercellular communication (GJIC; however, the role of Src and its effector Signal transducer and activator of transcription-3 (Stat3 upon GJIC in non small cell lung cancer (NSCLC has not been defined. Immunohistochemical analysis revealed high Src activity in NSCLC biopsy samples compared to normal tissues. Here we explored the potential effect of Src and Stat3 upon GJIC, by assessing the levels of tyr418-phosphorylated Src and tyr705-phosphorylated Stat3, respectively, in a panel of NSCLC cell lines. Methods Gap junctional communication was examined by electroporating the fluorescent dye Lucifer yellow into cells grown on a transparent electrode, followed by observation of the migration of the dye to the adjacent, non-electroporated cells under fluorescence illumination. Results An inverse relationship between Src activity levels and GJIC was noted; in five lines with high Src activity GJIC was absent, while two lines with extensive GJIC (QU-DB and SK-LuCi6 had low Src levels, similar to a non-transformed, immortalised lung epithelial cell line. Interestingly, examination of the mechanism indicated that Stat3 inhibition in any of the NSCLC lines expressing high endogenous Src activity levels, or in cells where Src was exogenously transduced, did not restore GJIC. On the contrary, Stat3 downregulation in immortalised lung epithelial cells or in the NSCLC lines displaying extensive GJIC actually suppressed junctional permeability. Conclusions Our findings demonstrate that although Stat3 is generally growth promoting and in an activated form it can act as an oncogene, it is actually required for gap junctional communication both in nontransformed lung epithelial cells and in certain lung cancer lines that retain extensive GJIC.

  10. The B[a]P-increased intercellular communication via translocation of connexin-43 into gap junctions reduces apoptosis

    International Nuclear Information System (INIS)

    Tekpli, X.; Rivedal, E.; Gorria, M.; Landvik, N.E.; Rissel, M.; Dimanche-Boitrel, M.-T.; Baffet, G.; Holme, J.A.; Lagadic-Gossmann, D.

    2010-01-01

    Gap junctions are channels in plasma membrane composed of proteins called connexins. These channels are organized in special domains between cells, and provide for direct gap junctional intercellular communication (GJIC), allowing diffusion of signalling molecules < 1 kD. GJIC regulates cell homeostasis and notably the balance between proliferation, cell cycle arrest, cell survival and apoptosis. Here, we have investigated benzo[a]pyrene (B[a]P) effects on GJIC and on the subcellular localization of the major protein of gap junction: connexin-43 (Cx43). Our results showed that B[a]P increased GJIC between mouse hepatoma Hepa1c1c7 cells via translocation of Cx43 from Golgi apparatus and lipid rafts into gap junction plaques. Interestingly, inhibition of GJIC by chlordane or small interference RNA directed against Cx43 enhanced B[a]P-induced apoptosis in Hepa1c1c7 cells. The increased apoptosis caused by inhibition of GJIC appeared to be mediated by ERK/MAPK pathway. It is suggested that B[a]P could induce transfer of cell survival signal or dilute cell death signal via regulation of ERK/MAPK through GJIC.

  11. Intercellular Communication between Keratinocytes and Fibroblasts Induces Local Osteoclast Differentiation: a Mechanism Underlying Cholesteatoma-Induced Bone Destruction.

    Science.gov (United States)

    Iwamoto, Yoriko; Nishikawa, Keizo; Imai, Ryusuke; Furuya, Masayuki; Uenaka, Maki; Ohta, Yumi; Morihana, Tetsuo; Itoi-Ochi, Saori; Penninger, Josef M; Katayama, Ichiro; Inohara, Hidenori; Ishii, Masaru

    2016-06-01

    Bone homeostasis is maintained by a balance in activity between bone-resorbing osteoclasts and bone-forming osteoblasts. Shifting the balance toward bone resorption causes osteolytic bone diseases such as rheumatoid arthritis and periodontitis. Osteoclast differentiation is regulated by receptor activator of nuclear factor κB ligand (RANKL), which, under some pathological conditions, is produced by T and B lymphocytes and synoviocytes. However, the mechanism underlying bone destruction in other diseases is little understood. Bone destruction caused by cholesteatoma, an epidermal cyst in the middle ear resulting from hyperproliferation of keratinizing squamous epithelium, can lead to lethal complications. In this study, we succeeded in generating a model for cholesteatoma, epidermal cyst-like tissue, which has the potential for inducing osteoclastogenesis in mice. Furthermore, an in vitro coculture system composed of keratinocytes, fibroblasts, and osteoclast precursors was used to demonstrate that keratinocytes stimulate osteoclast differentiation through the induction of RANKL in fibroblasts. Thus, this study demonstrates that intercellular communication between keratinocytes and fibroblasts is involved in the differentiation and function of osteoclasts, which may provide the molecular basis of a new therapeutic strategy for cholesteatoma-induced bone destruction. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  12. Possible Mechanisms of Mercury Toxicity and Cancer Promotion: Involvement of Gap Junction Intercellular Communications and Inflammatory Cytokines

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    Roberto Zefferino

    2017-01-01

    Full Text Available A number of observations indicate that heavy metals are able to alter cellular metabolic pathways through induction of a prooxidative state. Nevertheless, the outcome of heavy metal-mediated effects in the development of human diseases is debated and needs further insights. Cancer is a well-established DNA mutation-linked disease; however, epigenetic events are perhaps more important and harmful than genetic alterations. Unfortunately, we do not have reliable screening methods to assess/validate the epigenetic (promoter effects of a physical or a chemical agent. We propose a mechanism of action whereby mercury acts as a possible promoter carcinogen. In the present contribution, we resume our previous studies on mercury tested at concentrations comparable with its occurrence as environmental pollutant. It is shown that Hg(II elicits a prooxidative state in keratinocytes linked to inhibition of gap junction-mediated intercellular communication and proinflammatory cytokine production. These combined effects may on one hand isolate cells from tissue-specific homeostasis promoting their proliferation and on the other hand tamper the immune system defense/surveillance checkmating the whole organism. Since Hg(II is not a mutagenic/genotoxic compound directly affecting gene expression, in a broader sense, mercury might be an example of an epigenetic tumor promoter or, further expanding this concept, a “metagenetic” effector.

  13. Biochemical alterations in inflammatory reactive chondrocytes: evidence for intercellular network communication

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    Eva Skiöldebrand

    2018-01-01

    Full Text Available Chondrocytes are effectively involved in the pathophysiological processes of inflammation in joints. They form cellular processes in the superficial layer of the articular cartilage and form gap junction coupled syncytium to facilitate cell-to-cell communication. However, very little is known about their physiological cellular identity and communication. The aim with the present work is to evaluate the physiological behavior after stimulation with the inflammatory inducers interleukin-1β and lipopolysaccharide. The cytoskeleton integrity and intracellular Ca2+ release were assessed as indicators of inflammatory state. Cytoskeleton integrity was analyzed through cartilage oligomeric matrix protein and actin labeling with an Alexa 488-conjugated phalloidin probe. Ca2+ responses were assessed through the Ca2+ sensitive fluorophore Fura-2/AM. Western blot analyses of several inflammatory markers were performed. The results show reorganization of the actin filaments. Glutamate, 5-hydoxytryptamine, and ATP evoked intracellular Ca2+ release changed from single peaks to oscillations after inflammatory induction in the chondrocytes. The expression of toll-like receptor 4, the glutamate transporters GLAST and GLT-1, and the matrix metalloproteinase-13 increased. This work demonstrates that chondrocytes are a key part in conditions that lead to inflammation in the cartilage. The inflammatory inducers modulate the cytoskeleton, the Ca2+ signaling, and several inflammatory parameters. In conclusion, our data show that the cellular responses to inflammatory insults from healthy and inflammatory chondrocytes resemble those previously observed in astrocyte and cardiac fibroblasts networks.

  14. Ochratoxim A alters cell adhesion and gap junction intercellular communication in MDCK cells

    International Nuclear Information System (INIS)

    Mally, Angela; Decker, Martina; Bekteshi, Michaela; Dekant, Wolfgang

    2006-01-01

    Ochratoxin A (OTA) is one of the most potent renal carcinogens studied to date, but the mechanism of tumor formation by ochratoxin A remains largely unknown. Cell adhesion and cell-cell communication participate in the regulation of signaling pathways involved in cell proliferation and growth control and it is therefore not surprising that modulation of cell-cell signaling has been implicated in cancer development. Several nephrotoxicants and renal carcinogens have been shown to alter cell-cell signaling by interference with gap junction intercell communication (GJIC) and/or cell adhesion, and the aim of this study was to determine if disruption of cell-cell interactions occurs in kidney epithelial cells in response to OTA treatment. MDCK cells were treated with OTA (0-50 μM) for up to 24 h and gap junction function was analyzed using the scrape-load/dye transfer assay. In addition, expression and intracellular localization of Cx43, E-cadherin and β-catenin were determined by immunoblot and immunofluorescence analysis. A clear decrease in the distance of dye transfer was evident following treatment with OTA at concentrations/incubation times which did not affect cell viability. Consistent with the functional inhibition of GJIC, treatment with OTA resulted in a dose-dependent decrease in Cx43 expression. In contrast to Cx43, OTA did not alter total amount of the adherens junction proteins E-cadherin and β-catenin. Moreover, Western blot analysis of Triton X-100 soluble and insoluble protein fractions did not indicate translocation of cell adhesion molecules from the membrane to the cytoplasm. However, a ∼78 kDa fragment of β-catenin was detected in the detergent soluble fraction, indicating proteolytic cleavage of β-catenin. Immunofluorescence analysis also revealed changes in the pattern of both β-catenin and E-cadherin labeling, suggesting that OTA may alter cell-adhesion. Taken together, these data support the hypothesis that disruption of cell

  15. A rapid co-culture stamping device for studying intercellular communication

    Science.gov (United States)

    Hassanzadeh-Barforoushi, Amin; Shemesh, Jonathan; Farbehi, Nona; Asadnia, Mohsen; Yeoh, Guan Heng; Harvey, Richard P.; Nordon, Robert E.; Warkiani, Majid Ebrahimi

    2016-10-01

    Regulation of tissue development and repair depends on communication between neighbouring cells. Recent advances in cell micro-contact printing and microfluidics have facilitated the in-vitro study of homotypic and heterotypic cell-cell interaction. Nonetheless, these techniques are still complicated to perform and as a result, are seldom used by biologists. We report here development of a temporarily sealed microfluidic stamping device which utilizes a novel valve design for patterning two adherent cell lines with well-defined interlacing configurations to study cell-cell interactions. We demonstrate post-stamping cell viability of >95%, the stamping of multiple adherent cell types, and the ability to control the seeded cell density. We also show viability, proliferation and migration of cultured cells, enabling analysis of co-culture boundary conditions on cell fate. We also developed an in-vitro model of endothelial and cardiac stem cell interactions, which are thought to regulate coronary repair after myocardial injury. The stamp is fabricated using microfabrication techniques, is operated with a lab pipettor and uses very low reagent volumes of 20 μl with cell injection efficiency of >70%. This easy-to-use device provides a general strategy for micro-patterning of multiple cell types and will be important for studying cell-cell interactions in a multitude of applications.

  16. Potential microRNA-mediated oncogenic intercellular communication revealed by pan-cancer analysis

    Science.gov (United States)

    Li, Yue; Zhang, Zhaolei

    2014-11-01

    Carcinogenesis consists of oncogenesis and metastasis, and intriguingly microRNAs (miRNAs) are involved in both processes. Although aberrant miRNA activities are prevalent in diverse tumor types, the exact mechanisms for how they regulate cancerous processes are not always clear. To this end, we performed a large-scale pan-cancer analysis via a novel probabilistic approach to infer recurrent miRNA-target interactions implicated in 12 cancer types using data from The Cancer Genome Atlas. We discovered ~20,000 recurrent miRNA regulations, which are enriched for cancer-related miRNAs/genes. Notably, miRNA 200 family (miR-200/141/429) is among the most prominent miRNA regulators, which is known to be involved in metastasis. Importantly, the recurrent miRNA regulatory network is not only enriched for cancer pathways but also for extracellular matrix (ECM) organization and ECM-receptor interactions. The results suggest an intriguing cancer mechanism involving miRNA-mediated cell-to-cell communication, which possibly involves delivery of tumorigenic miRNA messengers to adjacent cells via exosomes. Finally, survival analysis revealed 414 recurrent-prognostic associations, where both gene and miRNA involved in each interaction conferred significant prognostic power in one or more cancer types. Together, our comprehensive pan-cancer analysis provided not only biological insights into metastasis but also brought to bear the clinical relevance of the proposed recurrent miRNA-gene associations.

  17. Influence of Ionizing Radiation on Stromal-Epithelial Intercellular Communication in Esophageal Carcinogenesis

    Science.gov (United States)

    Patel, Zarana S.; Kalabis, Jiri; Rustgi, Anil K.; Cucinotta, Francis A.; Huff, Janice L.

    2010-01-01

    Esophageal cancer is the 6th leading cause of cancer death worldwide. Its development is associated with a variety of risk factors including tobacco use, heavy alcohol consumption, human papilloma virus infection, and certain dietary factors such as trace mineral and vitamin deficiencies. An association with ionizing radiation exposure is revealed by the high excess relative risk for squamous cell carcinoma of the esophagus observed in the survivors of the atomic bomb detonations in Japan. It is also seen as a secondary malignancy in patients who received radiotherapy for breast and thoracic cancers; additionally, patients with head/neck and oral squamous cell cancers are at increased risk for metachronous esophageal squamous cell cancers. This malignancy is rapidly fatal, mainly because it remains asymptomatic until late, advanced stages when the disease is rarely curable. The stromal microenvironment plays an essential role in the maintenance and modulation of normal epithelial cell growth and differentiation and cross talk between the epithelial and stromal compartments can influence many aspects of malignant progression, including tumor cell proliferation, migration, invasion and recruitment of new blood vessels. To test the hypothesis that radiation exposure plays a role in esophageal carcinogenesis via non-targeted mechanisms involving stromal-epithelial cell communication, we are studying radiation effects on hTERT-immortalized human esophageal epithelial cells and genetic variants grown in co-culture with human esophageal stromal fibroblasts (Okawa et al., Genes & Dev. 2007. 21: 2788-2803). We examined how radiation treatment of stromal fibroblasts affected epithelial migration and invasion, behaviors associated with cancer promotion and progression. Chemotactic and haptotactic migration of epithelial cells stimulated by conditioned media from irradiated fibroblasts was measured using assays conducted in Transwell cell culture chambers. Our results using

  18. Autophagy and gap junctional intercellular communication inhibition are involved in cadmium-induced apoptosis in rat liver cells

    Energy Technology Data Exchange (ETDEWEB)

    Zou, Hui [College of Veterinary Medicine, Yangzhou University, and Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009 (China); Zhuo, Liling [College of Life Science, Zaozhuang University, Zaozhuang, Shandong, 277160 (China); Han, Tao; Hu, Di; Yang, Xiaokang; Wang, Yi; Yuan, Yan; Gu, Jianhong; Bian, Jianchun; Liu, Xuezhong [College of Veterinary Medicine, Yangzhou University, and Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009 (China); Liu, Zongping, E-mail: liuzongping@yzu.edu.cn [College of Veterinary Medicine, Yangzhou University, and Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, Jiangsu, 225009 (China)

    2015-04-17

    Cadmium (Cd) is known to induce hepatotoxicity, yet the underlying mechanism of how this occurs is not fully understood. In this study, Cd-induced apoptosis was demonstrated in rat liver cells (BRL 3A) with apoptotic nuclear morphological changes and a decrease in cell index (CI) in a time- and concentration-dependent manner. The role of gap junctional intercellular communication (GJIC) and autophagy in Cd-induced apoptosis was investigated. Cd significantly induced GJIC inhibition as well as downregulation of connexin 43 (Cx43). The prototypical gap junction blocker carbenoxolone disodium (CBX) exacerbated the Cd-induced decrease in CI. Cd treatment was also found to cause autophagy, with an increase in mRNA expression of autophagy-related genes Atg-5, Atg-7, Beclin-1, and microtubule-associated protein light chain 3 (LC3) conversion from cytosolic LC3-I to membrane-bound LC3-II. The autophagic inducer rapamycin (RAP) prevented the Cd-induced CI decrease, while the autophagic inhibitor chloroquine (CQ) caused a further reduction in CI. In addition, CBX promoted Cd-induced autophagy, as well as changes in expression of Atg-5, Atg-7, Beclin-1 and LC3. CQ was found to block the Cd-induced decrease in Cx43 and GJIC inhibition, whereas RAP had opposite effect. These results demonstrate that autophagy plays a protective role during Cd-induced apoptosis in BRL 3A cells during 6 h of experiment, while autophagy exacerbates Cd-induced GJIC inhibition which has a negative effect on cellular fate. - Highlights: • GJIC and autophagy is crucial for biological processes. • Cd exposure causes GJIC inhibition and autophagy increase in BRL 3A cells. • Autophagy protects Cd induced BRL 3A cells apoptosis at an early stage. • Autophagy exacerbates Cd-induced GJIC inhibition. • GJIC plays an important role in autophagy induced cell death or survival.

  19. Low dose/low fluence ionizing radiation-induced biological effects: The role of intercellular communication and oxidative metabolism

    Science.gov (United States)

    Azzam, Edouard

    Mechanistic investigations have been considered critical to understanding the health risks of exposure to ionizing radiation. To gain greater insight in the biological effects of exposure to low dose/low fluence space radiations with different linear energy transfer (LET) properties, we examined short and long-term biological responses to energetic protons and high charge (Z) and high energy (E) ions (HZE particles) in human cells maintained in culture and in targeted and non-targeted tissues of irradiated rodents. Particular focus of the studies has been on mod-ulation of gene expression, proliferative capacity, induction of DNA damage and perturbations in oxidative metabolism. Exposure to mean doses of 1000 MeV/nucleon iron ions, by which a small to moderate proportion of cells in an exposed population is targeted through the nucleus by an HZE particle, induced stressful effects in the irradiated and non-irradiated cells in the population. Direct intercellular communication via gap-junctions was a primary mediator of the propagation of stressful effects from irradiated to non-irradiated cells. Compromised prolif-erative capacity, elevated level of DNA damage and oxidative stress evaluated by measurements of protein carbonylation, lipid peroxidation and activity of metabolic enzymes persisted in the progeny of irradiated and non-irradiated cells. In contrast, progeny of cells exposed to high or low doses from 150-1000 MeV protons retained the ability to form colonies and harbored similar levels of micronuclei, a surrogate form of DNA damage, as control, which correlated with normal reactive oxygen species (ROS) levels. Importantly, a significant increase in the spontaneous neoplastic transformation frequency was observed in progeny of bystander mouse embryo fibroblasts (MEFs) co-cultured with MEFs irradiated with energetic iron ions but not protons. Of particular significance, stressful effects were detected in non-targeted tissues of rats that received partial

  20. The antiarrhythmic peptide analog rotigaptide (ZP123) stimulates gap junction intercellular communication in human osteoblasts and prevents decrease in femoral trabecular bone strength in ovariectomized rats

    DEFF Research Database (Denmark)

    Jørgensen, Niklas Rye; Teilmann, Stefan Cuoni; Henriksen, Zanne

    2005-01-01

    Gap junctions play an important role in bone development and function, but the lack of pharmacological tools has hampered the gap junction research. The antiarrhythmic peptides stimulate gap junction communication between cardiomyocytes, but effects in noncardiac tissue are unknown. The purpose...... of this study was to examine whether antiarrhythmic peptides, which are small peptides increasing gap junctional conductivity, show specific binding to osteoblasts and investigate the effect of the stable analog rotigaptide (ZP123) on gap junctional intercellular communication in vitro and on bone mass...... and strength in vivo. Cell coupling and calcium signaling were assessed in vitro on human, primary, osteoblastic cells. In vivo effects of rotigaptide on bone strength and density were determined 4 wk after ovariectomy in rats treated with either vehicle, sc injection twice daily (300 nmol per kilogram body...

  1. When a transmembrane channel isn't, or how biophysics and biochemistry (mis)communicate.

    Science.gov (United States)

    Reviakine, Ilya

    2018-02-12

    Annexins are a family of soluble proteins that bind to acidic phospholipids such as phosphatidylserine in a calcium-dependent manner. The archetypical member of the annexin family is annexin A5. For many years, its function remained unknown despite the availability of a high-resolution structure. This, combined with the observations of specific ion conductance in annexin-bound membranes, fueled speculations about the possible membrane-spanning forms of annexins that functioned as ion channels. The channel hypothesis remained controversial and did not gather sufficient evidence to become accepted. Yet, it continues to draw attention as a framework for interpreting indirect (e.g., biochemical) data. The goal of the mini-review is to examine the data on annexin-lipid interactions from the last ~30 years from the point of view of the controversy between the two lines of inquiry: the well-characterized peripheral assembly of the annexins at membranes vs. their putative transmembrane insertion. In particular, the potential role of lipid rearrangements induced by annexin binding is highlighted. Copyright © 2018 Elsevier B.V. All rights reserved.

  2. Silver nanoparticles increase connexin43-mediated gap junctional intercellular communication in HaCaT cells through activation of reactive oxygen species and mitogen-activated protein kinase signal pathway

    DEFF Research Database (Denmark)

    Qin, Yu; Han, Limin; Yang, Di

    2018-01-01

    Silver nanoparticles (AgNPs) are widely used in health and consumer products that routinely contact skin. However, the biological effects and possible mechanisms of AgNPs on skin remain unclear. Gap junctional intercellular communication (GJIC) plays a critical role in multicellular organisms to ...

  3. Intercellular bridges in vertebrate gastrulation.

    Directory of Open Access Journals (Sweden)

    Luca Caneparo

    Full Text Available The developing zebrafish embryo has been the subject of many studies of regional patterning, stereotypical cell movements and changes in cell shape. To better study the morphological features of cells during gastrulation, we generated mosaic embryos expressing membrane attached Dendra2 to highlight cellular boundaries. We find that intercellular bridges join a significant fraction of epiblast cells in the zebrafish embryo, reaching several cell diameters in length and spanning across different regions of the developing embryos. These intercellular bridges are distinct from the cellular protrusions previously reported as extending from hypoblast cells (1-2 cellular diameters in length or epiblast cells (which were shorter. Most of the intercellular bridges were formed at pre-gastrula stages by the daughters of a dividing cell maintaining a membrane tether as they move apart after mitosis. These intercellular bridges persist during gastrulation and can mediate the transfer of proteins between distant cells. These findings reveal a surprising feature of the cellular landscape in zebrafish embryos and open new possibilities for cell-cell communication during gastrulation, with implications for modeling, cellular mechanics, and morphogenetic signaling.

  4. Evolution of Microbial Quorum Sensing to Human Global Quorum Sensing: An Insight into How Gap Junctional Intercellular Communication Might Be Linked to the Global Metabolic Disease Crisis.

    Science.gov (United States)

    Trosko, James E

    2016-06-15

    The first anaerobic organism extracted energy for survival and reproduction from its source of nutrients, with the genetic means to ensure protection of its individual genome but also its species survival. While it had a means to communicate with its community via simple secreted molecules ("quorum sensing"), the eventual shift to an aerobic environment led to multi-cellular metazoan organisms, with evolutionary-selected genes to form extracellular matrices, stem cells, stem cell niches, and a family of gap junction or "connexin" genes. These germinal and somatic stem cells responded to extracellular signals that triggered intra-cellular signaling to regulate specific genes out of the total genome. These extra-cellular induced intra-cellular signals also modulated gap junctional intercellular communication (GJIC) in order to regulate the new cellular functions of symmetrical and asymmetrical cell division, cell differentiation, modes of cell death, and senescence. Within the hierarchical and cybernetic concepts, differentiated by neurons organized in the brain of the Homo sapiens, the conscious mind led to language, abstract ideas, technology, myth-making, scientific reasoning, and moral decision-making, i.e., the creation of culture. Over thousands of years, this has created the current collision between biological and cultural evolution, leading to the global "metabolic disease" crisis.

  5. Evolution of Microbial Quorum Sensing to Human Global Quorum Sensing: An Insight into How Gap Junctional Intercellular Communication Might Be Linked to the Global Metabolic Disease Crisis

    Directory of Open Access Journals (Sweden)

    James E. Trosko

    2016-06-01

    Full Text Available The first anaerobic organism extracted energy for survival and reproduction from its source of nutrients, with the genetic means to ensure protection of its individual genome but also its species survival. While it had a means to communicate with its community via simple secreted molecules (“quorum sensing”, the eventual shift to an aerobic environment led to multi-cellular metazoan organisms, with evolutionary-selected genes to form extracellular matrices, stem cells, stem cell niches, and a family of gap junction or “connexin” genes. These germinal and somatic stem cells responded to extracellular signals that triggered intra-cellular signaling to regulate specific genes out of the total genome. These extra-cellular induced intra-cellular signals also modulated gap junctional intercellular communication (GJIC in order to regulate the new cellular functions of symmetrical and asymmetrical cell division, cell differentiation, modes of cell death, and senescence. Within the hierarchical and cybernetic concepts, differentiated by neurons organized in the brain of the Homo sapiens, the conscious mind led to language, abstract ideas, technology, myth-making, scientific reasoning, and moral decision–making, i.e., the creation of culture. Over thousands of years, this has created the current collision between biological and cultural evolution, leading to the global “metabolic disease” crisis.

  6. Nucleotide transmitters ATP and ADP mediate intercellular calcium wave communication via P2Y12/13 receptors among BV-2 microglia.

    Directory of Open Access Journals (Sweden)

    Pengchong Jiang

    Full Text Available Nerve injury is accompanied by a liberation of diverse nucleotides, some of which act as 'find/eat-me' signals in mediating neuron-glial interplay. Intercellular Ca2+ wave (ICW communication is the main approach by which glial cells interact and coordinate with each other to execute immune defense. However, the detailed mechanisms on how these nucleotides participate in ICW communication remain largely unclear. In the present work, we employed a mechanical stimulus to an individual BV-2 microglia to simulate localized injury. Remarkable ICW propagation was observed no matter whether calcium was in the environment or not. Apyrase (ATP/ADP-hydrolyzing enzyme, suramin (broad-spectrum P2 receptor antagonist, 2-APB (IP3 receptor blocker and thapsigargin (endoplasmic reticulum calcium pump inhibitor potently inhibited these ICWs, respectively, indicating the dependence of nucleotide signals and P2Y receptors. Then, we detected the involvement of five naturally occurring nucleotides (ATP, ADP, UTP, UDP and UDP-glucose by desensitizing receptors. Results showed that desensitization with ATP and ADP could block ICW propagation in a dose-dependent manner, whereas other nucleotides had little effect. Meanwhile, the expression of P2Y receptors in BV-2 microglia was identified and their contributions were analyzed, from which we suggested P2Y12/13 receptors activation mostly contributed to ICWs. Besides, we estimated that extracellular ATP and ADP concentration sensed by BV-2 microglia was about 0.3 μM during ICWs by analyzing calcium dynamic characteristics. Taken together, these results demonstrated that the nucleotides ATP and ADP were predominant signal transmitters in mechanical stimulation-induced ICW communication through acting on P2Y12/13 receptors in BV-2 microglia.

  7. Na,K-ATPase regulates intercellular communication in the vascular wall via cSrc kinase dependent connexin43 phosphorylation

    DEFF Research Database (Denmark)

    Hangaard, Lise; Bouzinova, Elena; Stæhr, Christian Albeck

    2017-01-01

    Communication between vascular smooth muscle cells (VSMCs) is dependent on gap junctions and is regulated by the Na-K-ATPase. The Na-K-ATPase is therefore important for synchronized VSMC oscillatory activity, i.e., vasomotion. The signaling between the Na-K-ATPase and gap junctions is unknown. We...... coupling in rat mesenteric small arteries in vitro. Phosphorylation of cSrc kinase and connexin43 (Cx43) were semiquantified by Western blotting. Micromole concentration of ouabain reduced the amplitude of norepinephrine-induced vasomotion and desynchronized Ca2+ transients in VSMC in the arterial wall...

  8. Inhibition of gap-junctional intercellular communication and activation of mitogen-activated protein kinases by cyanobacterial extracts--indications of novel tumor-promoting cyanotoxins?

    Science.gov (United States)

    Bláha, Ludĕk; Babica, Pavel; Hilscherová, Klára; Upham, Brad L

    2010-01-01

    Toxicity and liver tumor promotion of cyanotoxins microcystins have been extensively studied. However, recent studies document that other metabolites present in the complex cyanobacterial water blooms may also have adverse health effects. In this study we used rat liver epithelial stem-like cells (WB-F344) to examine the effects of cyanobacterial extracts on two established markers of tumor promotion, inhibition of gap-junctional intercellular communication (GJIC) and activation of mitogen-activated protein kinases (MAPKs) - ERK1/2. Extracts of cyanobacteria (laboratory cultures of Microcystis aeruginosa and Aphanizomenon flos-aquae and water blooms dominated by these species) inhibited GJIC and activated MAPKs in a dose-dependent manner (effective concentrations ranging 0.5-5mgd.w./mL). Effects were independent of the microcystin content and the strongest responses were elicited by the extracts of Aphanizomenon sp. Neither pure microcystin-LR nor cylindrospermopsin inhibited GJIC or activated MAPKs. Modulations of GJIC and MAPKs appeared to be specific to cyanobacterial extracts since extracts from green alga Chlamydomonas reinhardtii, heterotrophic bacterium Klebsiella terrigena, and isolated bacterial lipopolysaccharides had no comparable effects. Our study provides the first evidence on the existence of unknown cyanobacterial toxic metabolites that affect in vitro biomarkers of tumor promotion, i.e. inhibition of GJIC and activation of MAPKs.

  9. Antiproliferative Action of Conjugated Linoleic Acid on Human MCF-7 Breast Cancer Cells Mediated by Enhancement of Gap Junctional Intercellular Communication through Inactivation of NF-κB

    Directory of Open Access Journals (Sweden)

    Md. Abdur Rakib

    2013-01-01

    Full Text Available The major conjugated linoleic acid (CLA isomers, c9,t11-CLA and t10,c12-CLA, have anticancer effects; however, the exact mechanisms underlying these effects are unknown. Evidence suggests that reversal of reduced gap junctional intercellular communication (GJIC in cancer cells inhibits cell growth and induces cell death. Hence, we determined that CLA isomers enhance GJIC in human MCF-7 breast cancer cells and investigated the underlying molecular mechanisms. The CLA isomers significantly enhanced GJIC of MCF-7 cells at 40 μM concentration, whereas CLA inhibited cell growth and induced caspase-dependent apoptosis. CLA increased connexin43 (Cx43 expression both at the transcriptional and translational levels. CLA inhibited nuclear factor-κB (NF-κB activity and enhanced reactive oxygen species (ROS generation. No significant difference was observed in the efficacy of c9,t11-CLA and t10,c12-CLA. These results suggest that the anticancer effect of CLA is associated with upregulation of GJIC mediated by enhanced Cx43 expression through inactivation of NF-κB and generation of ROS in MCF-7 cells.

  10. Inhibition of gap junctional Intercellular communication in WB-F344 rat liver epithelial cells by triphenyltin chloride through MAPK and PI3-kinase pathways

    Directory of Open Access Journals (Sweden)

    Tsai Ming-Che

    2010-06-01

    Full Text Available Abstract Background Organotin compounds (OTCs have been widely used as stabilizers in the production of plastic, agricultural pesticides, antifoulant plaints and wood preservation. The toxicity of triphenyltin (TPT compounds was known for their embryotoxic, neurotoxic, genotoxic and immunotoxic effects in mammals. The carcinogenicity of TPT was not well understood and few studies had discussed the effects of OTCs on gap junctional intercellular communication (GJIC of cells. Method In the present study, the effects of triphenyltin chloride (TPTC on GJIC in WB-F344 rat liver epithelial cells were evaluated, using the scrape-loading dye transfer technique. Results TPTC inhibited GJIC after a 30-min exposure in a concentration- and time-dependent manner. Pre-incubation of cells with the protein kinase C (PKC inhibitor did not modify the response, but the specific MEK 1 inhibitor PD98059 and PI3K inhibitor LY294002 decreased substantially the inhibition of GJIC by TPTC. After WB-F344 cells were exposed to TPTC, phosphorylation of Cx43 increased as seen in Western blot analysis. Conclusions These results show that TPTC inhibits GJIC in WB-F344 rat liver epithelial cells by altering the Cx43 protein expression through both MAPK and PI3-kinase pathways.

  11. Neuropeptide Y, substance P, and human bone morphogenetic protein 2 stimulate human osteoblast osteogenic activity by enhancing gap junction intercellular communication

    International Nuclear Information System (INIS)

    Ma, W.H.; Liu, Y.J.; Wang, W.; Zhang, Y.Z.

    2015-01-01

    Bone homeostasis seems to be controlled by delicate and subtle “cross talk” between the nervous system and “osteo-neuromediators” that control bone remodeling. The purpose of this study was to evaluate the effect of interactions between neuropeptides and human bone morphogenetic protein 2 (hBMP2) on human osteoblasts. We also investigated the effects of neuropeptides and hBMP2 on gap junction intercellular communication (GJIC). Osteoblasts were treated with neuropeptide Y (NPY), substance P (SP), or hBMP2 at three concentrations. At various intervals after treatment, cell viability was measured by the MTT assay. In addition, cellular alkaline phosphatase (ALP) activity and osteocalcin were determined by colorimetric assay and radioimmunoassay, respectively. The effects of NPY, SP and hBMP on GJIC were determined by laser scanning confocal microscopy. The viability of cells treated with neuropeptides and hBMP2 increased significantly in a time-dependent manner, but was inversely associated with the concentration of the treatments. ALP activity and osteocalcin were both reduced in osteoblasts exposed to the combination of neuropeptides and hBMP2. The GJIC of osteoblasts was significantly increased by the neuropeptides and hBMP2. These results suggest that osteoblast activity is increased by neuropeptides and hBMP2 through increased GJIC. Identification of the GJIC-mediated signal transduction capable of modulating the cellular activities of bone cells represents a novel approach to studying the biology of skeletal innervation

  12. Tanshinone IIA increases the bystander effect of herpes simplex virus thymidine kinase/ganciclovir gene therapy via enhanced gap junctional intercellular communication.

    Directory of Open Access Journals (Sweden)

    Jianyong Xiao

    Full Text Available The bystander effect is an intriguing phenomenon by which adjacent cells become sensitized to drug treatment during gene therapy with herpes simplex virus thymidine kinase/ganciclovir (HSV-tk/GCV. This effect is reported to be mediated by gap junctional intercellular communication (GJIC, and therefore, we postulated that upregulation of genes that facilitate GJIC may enhance the HSV-tk/GCV bystander effect. Previous findings have shown Tanshinone IIA (Tan IIA, a chemical substance derived from a Chinese medicine herb, promotes the upregulation of the connexins Cx26 and Cx43 in B16 cells. Because gap junctions are formed by connexins, we hypothesized that Tan IIA might increase GJIC. Our results show that Tan IIA increased GJIC in B16 melanoma cells, leading to more efficient GCV-induced bystander killing in cells stably expressing HSV-tk. Additionally, in vivo experiments demonstrated that tumors in mice with 10% HSV-tk positive B16 cells and 90% wild-type B16 cells became smaller following treatment with the combination of GCV and Tan IIA as compared to GCV or Tan IIA alone. These data demonstrate that Tan IIA can augment the bystander effect of HSV-tk/GCV system through increased gap junction coupling, which adds strength to the promising strategy that develops connexins inducer to potentiate the effects of suicide gene therapy.

  13. Neuropeptide Y, substance P, and human bone morphogenetic protein 2 stimulate human osteoblast osteogenic activity by enhancing gap junction intercellular communication

    Energy Technology Data Exchange (ETDEWEB)

    Ma, W.H.; Liu, Y.J.; Wang, W.; Zhang, Y.Z. [The Third Hospital of Hebei Medical University, The Provincial Key Laboratory for Orthopedic Biomechanics of Hebei, Shijiazhuang, Hebei Province (China)

    2015-02-13

    Bone homeostasis seems to be controlled by delicate and subtle “cross talk” between the nervous system and “osteo-neuromediators” that control bone remodeling. The purpose of this study was to evaluate the effect of interactions between neuropeptides and human bone morphogenetic protein 2 (hBMP2) on human osteoblasts. We also investigated the effects of neuropeptides and hBMP2 on gap junction intercellular communication (GJIC). Osteoblasts were treated with neuropeptide Y (NPY), substance P (SP), or hBMP2 at three concentrations. At various intervals after treatment, cell viability was measured by the MTT assay. In addition, cellular alkaline phosphatase (ALP) activity and osteocalcin were determined by colorimetric assay and radioimmunoassay, respectively. The effects of NPY, SP and hBMP on GJIC were determined by laser scanning confocal microscopy. The viability of cells treated with neuropeptides and hBMP2 increased significantly in a time-dependent manner, but was inversely associated with the concentration of the treatments. ALP activity and osteocalcin were both reduced in osteoblasts exposed to the combination of neuropeptides and hBMP2. The GJIC of osteoblasts was significantly increased by the neuropeptides and hBMP2. These results suggest that osteoblast activity is increased by neuropeptides and hBMP2 through increased GJIC. Identification of the GJIC-mediated signal transduction capable of modulating the cellular activities of bone cells represents a novel approach to studying the biology of skeletal innervation.

  14. Intercellular communication in Helicobacter pylori: luxS is essential for the production of an extracellular signaling molecule.

    Science.gov (United States)

    Forsyth, M H; Cover, T L

    2000-06-01

    Individual bacteria of numerous species can communicate and coordinate their actions via the production, release, and detection of extracellular signaling molecules. In this study, we used the Vibrio harveyi luminescence bioassay to determine whether Helicobacter pylori produces such a factor. Cell-free conditioned media from H. pylori strains 60190 and 26695 each induced >100-fold-greater luminescence in V. harveyi than did sterile culture medium. The H. pylori signaling molecule had a molecular mass of 100-fold-greater luminescence in the V. harveyi bioassay than did conditioned medium from either mutant strain. Production of the signaling molecule was restored in an H. pylori luxS null mutant strain by complementation with a single intact copy of luxS placed in a heterologous site on the chromosome. In addition, Escherichia coli DH5alpha produced autoinducer activity following the introduction of an intact copy of luxS from H. pylori. Production of the signaling molecule by H. pylori was growth phase dependent, with maximal production occurring in the mid-exponential phase of growth. Transcription of H. pylori vacA also was growth phase dependent, but this phenomenon was not dependent on luxS activity. These data indicate that H. pylori produces an extracellular signaling molecule related to AI-2 from V. harveyi. We speculate that this signaling molecule may play a role in regulating H. pylori gene expression.

  15. Inhibition of Connexin 26/43 and Extracellular-Regulated Kinase Protein Plays a Critical Role in Melatonin Facilitated Gap Junctional Intercellular Communication in Hydrogen Peroxide-Treated HaCaT Keratinocyte Cells

    Directory of Open Access Journals (Sweden)

    Hyo-Jung Lee

    2012-01-01

    Full Text Available Though melatonin was known to regulate gap junctional intercellular communication (GJIC in chick astrocytes and mouse hepatocytes, the underlying mechanism by melatonin was not elucidated in hydrogen peroxide- (H2O2- treated HaCaT keratinocyte cells until now. In the current study, though melatonin at 2 mM and hydrogen peroxide (H2O2 at 300 μM showed weak cytotoxicity in HaCaT keratinocyte cells, melatonin significantly suppressed the formation of reactive oxygen species (ROS in H2O2-treated HaCaT cells compared to untreated controls. Also, the scrape-loading dye-transfer assay revealed that melatonin enhances the intercellular communication by introducing Lucifer Yellow into H2O2-treated cells. Furthermore, melatonin significantly enhanced the expression of connexin 26 (Cx26 and connexin 43 (Cx43 at mRNA and protein levels, but not that of connexin 30 (Cx30 in H2O2-treated HaCaT cells. Of note, melatonin attenuated the phosphorylation of extracellular signal-regulated protein kinases (ERKs more than p38 MAPK or JNK in H2O2-treated HaCaT cells. Conversely, ERK inhibitor PD98059 promoted the intercellular communication in H2O2-treated HaCaT cells. Furthermore, combined treatment of melatonin (200 μM and vitamin C (10 μg/mL significantly reduced ROS production in H2O2-treated HaCaT cells. Overall, these findings support the scientific evidences that melatonin facilitates gap junctional intercellular communication in H2O2-treated HaCaT keratinocyte cells via inhibition of connexin 26/43 and ERK as a potent chemopreventive agent.

  16. Dysfunction in gap junction intercellular communication induces aberrant behavior of the inner cell mass and frequent collapses of expanded blastocysts in mouse embryos.

    Science.gov (United States)

    Togashi, Kazue; Kumagai, Jin; Sato, Emiko; Shirasawa, Hiromitsu; Shimoda, Yuki; Makino, Kenichi; Sato, Wataru; Kumazawa, Yukiyo; Omori, Yasufumi; Terada, Yukihiro

    2015-06-01

    We investigated the role of gap junctions (GJs) in embryological differentiation, and observed the morphological behavior of the inner cell mass (ICM) by time-lapse movie observation (TLM) with gap junction inhibitors (GJis). ICR mouse embryos were exposed to two types of GJis in CZB medium: oleamide (0 to 50 μM) and 1-heptanol (0 to 10 mM). We compared the rate of blastocyst formation at embryonic day 4.5 (E4.5) with E5.5. We also observed and evaluated the times from the second cleavage to each embryonic developing stage by TLM. We investigated embryonic distribution of DNA, Nanog protein, and Connexin 43 protein with immunofluorescent staining. In the comparison of E4.5 with E5.5, inhibition of gap junction intercellular communication (GJIC) delayed embryonic blastocyst formation. The times from the second cleavage to blastocyst formation were significantly extended in the GJi-treated embryos (control vs with oleamide, 2224 ± 179 min vs 2354 ± 278 min, p = 0.013). Morphological differences were traced in control versus GJi-treated embryos until the hatching stage. Oleamide induced frequent severe collapses of expanded blastocysts (77.4 % versus 26.3 %, p = 0.0001) and aberrant ICM divisions connected to sticky strands (74.3 % versus 5.3 %, p = 0.0001). Immunofluorescent staining indicated Nanog-positive cells were distributed in each divided ICM. GJIC plays an important role in blastocyst formation, collapses of expanded blastocysts, and the ICM construction in mouse embryos.

  17. HYS-32, a novel analogue of combretastatin A-4, enhances connexin43 expression and gap junction intercellular communication in rat astrocytes.

    Science.gov (United States)

    Lin, Pei-Chun; Shen, Chien-Chang; Liao, Chih-Kai; Jow, Guey-Mei; Chiu, Chi-Ting; Chung, Tun-Hui; Wu, Jiahn-Chun

    2013-05-01

    HYS-32 [4-(3,4-dimethoxyphenyl)-3-(naphthalen-2-yl)-2(5H)-furanone] is a new analogue of the anti-tumor compound combretastatin A-4 containing a cis-stilbene moiety. In this study, we investigated its effects on Cx43 gap junction intercellular communication (GJIC) and the signaling pathway involved in rat primary astrocytes. Western blot analyses showed that HYS-32 dose- and time-dependently upregulated Cx43 expression. A confocal microscopic study and scrape-loading/dye transfer analyses demonstrated that HYS-32 (5μM) induced microtubule coiling, accumulation of Cx43 in gap junction plaques, and increased GJIC in astrocytes. The HYS-32-induced microtubule coiling and Cx43 accumulation in gap junction plaques was reversed when HYS-32 was removed. Treatment of astrocytes with cycloheximide resulted in time-dependent degradation of by co-treatment with HYS-32 by increasing the half-life of Cx43. Co-treatment with HYS-32 also prevented the LPS-induced downregulation of Cx43 and inhibition of GJIC in astrocytes. HYS-32 induced activation of PKC, ERK, and JNK, and co-treatment with the PKC inhibitor Go6976 or the ERK inhibitor PD98059, but not the JNK inhibitor SP600125, prevented the HYS-32-induced increase in Cx43 expression and GJIC. Go6976 suppressed the HYS-32-induced PKC phosphorylation and increase in phospho-ERK levels, while PD98059 did not prevent the HYS-32-induced increase in phospho-PKC levels, suggesting that PKC is an upstream effector of ERK. In conclusion, our results show that HYS-32 increases the half-life of Cx43 and enhances Cx43 expression and GJIC in astrocytes via a PKC-ERK signaling cascade. These novel biological effects of HYS-32 on astrocyte gap junctions support its potential for therapeutic use as a protective agent for the central nervous system. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. GAP junctional intercellular communication confers an enhanced bystander effect as well as a protective effect in HSV-TK/gancyclovir mediated cytotoxicity

    International Nuclear Information System (INIS)

    Wygoda, Marc R.; Rehemtulla, Alnawaz; Davis, Mary A.; Lawrence, Theodore S.

    1996-01-01

    Purpose/Objective: The 'Bystander Effect' is the phenomenon by which cells which are not transduced with the Herpes Simplex Virus - Thymidine Kinase (HSV-TK) gene are killed by the guanosine analog Gancyclovir (GCV) when they are nearby HSV-TK positive cells. Since currently available gene transfer technologies have a low efficiency (typically less than 10% of cells are transduced), a better understanding of the mechanism underlying the bystander effect is crucial for the success of the HSV-TK/GCV enzyme prodrug gene therapy approach. Gap junctions, which are specialized cell membrane channels allowing the passage of molecules < 1000 daltons in size directly from cell to cell, have been proposed as a possible mediator of the bystander effect. In particular, gap junctions could permit the transfer of toxic metabolites from the HSV-TK positive cells to the bystander cells. Our aim was to assess the role of gap junctional intercellular communication (GJIC) in the bystander effect. Materials and Methods: Co-culture of an HSV-TK transduced rat liver epithelial cell line (WB-TK), with either its communication competent parental cell line (WB), or an isogenic cell line differing by its communication incompetence (aB1). The ratios of effector cells (WB-TK) to bystander cells (WB or aB1), were (100(0)), (50(50)), (5(95)), or(0(100)) . These various mixed populations were exposed for 24 hours to 10μM GCV and thereafter placed at clonal density into selective media, allowing for growth of either the bystander (TK negative) cells or the effector (TK-positive) cells. Results: 1) When the effector/bystander ratio was (50(50)) the surviving fraction of the bystander cells was 3.5% and 37% for the WB and aB1, respectively. When the effector/bystander ratio was (5(95)), the surviving fraction was 51% and 65% for the WB and aB1, respectively. Either cell line (WB or aB1) cultured alone was unaffected by GCV. These results demonstrate that the bystander effect is much higher when

  19. [Effects of Chinese herbal compound for supplementing qi and activating blood circulation on actin, Cx43 expressions and gap junctional intercellular communication functions of myocardial cells in patients with Coxsackie virus B 3 viral myocarditis].

    Science.gov (United States)

    Zhang, Ming-xue; He, Wei; Gu, Ping

    2010-08-01

    To observe the effect of Chinese herbal compound for supplementing qi and activating blood circulation (CHC) on the gap junctional intercellular communication (GJIC) function of myocardial cells in patients with Coxsackie virus B 3 (CVB3) viral myocarditis. Expressions of actin and connexin43 (Cx43) in myocardial cells of patients arranged in three groups (the normal control group, the viral infected group and the CHC treated group) were detected by immunohistochemical method; the fluorescence photobleaching recovery rate of cells was detected by laser scanning confocal microscope. As compared with the viral infected group, the expressions of actin and Cx43 were increased and the GJIC function was improved in the CHC treated group. CHC could antagonize viral injury on skeleton protein, and repair the structure of gap junction channel to improve the GJIC function of myocardial cells after being attacked by CVB3.

  20. The Impact of the ‘Austrian’ Mutation of the Amyloid Precursor Protein Transmembrane Helix is Communicated to the Hinge Region

    DEFF Research Database (Denmark)

    Stelzer, Walter; Scharnagl, Christina; Leurs, Ulrike

    2016-01-01

    The transmembrane helix of the amyloid precursor protein is subject to proteolytic cleavages by γ-secretase at different sites resulting in Aβ peptides of different length and toxicity. A number of point mutations within this transmembrane helix alter the cleavage pattern thus enhancing production...... destabilizes amide hydrogen bonds in the hinge which connects dimerization and cleavage regions. Weaker intrahelical hydrogen bonds at the hinge may enhance helix bending and thereby affect recognition of the transmembrane substrate by the enzyme and/or presentation of its cleavage sites to the catalytic cleft....

  1. Inter-cellular transport of ran GTPase.

    Directory of Open Access Journals (Sweden)

    Deepak Khuperkar

    Full Text Available Ran, a member of the Ras-GTPase superfamily, has a well-established role in regulating the transport of macromolecules across the nuclear envelope (NE. Ran has also been implicated in mitosis, cell cycle progression, and NE formation. Over-expression of Ran is associated with various cancers, although the molecular mechanism underlying this phenomenon is unclear. Serendipitously, we found that Ran possesses the ability to move from cell-to-cell when transiently expressed in mammalian cells. Moreover, we show that the inter-cellular transport of Ran is GTP-dependent. Importantly, Ran displays a similar distribution pattern in the recipient cells as that in the donor cell and co-localizes with the Ran binding protein Nup358 (also called RanBP2. Interestingly, leptomycin B, an inhibitor of CRM1-mediated export, or siRNA mediated depletion of CRM1, significantly impaired the inter-cellular transport of Ran, suggesting a function for CRM1 in this process. These novel findings indicate a possible role for Ran beyond nucleo-cytoplasmic transport, with potential implications in inter-cellular communication and cancers.

  2. Disruption of gap junctional intercellular communication by antibiotic gentamicin is associated with aberrant localization of occludin, N-cadherin, connexin 43, and vimentin in SerW3 Sertoli cells in vitro.

    Science.gov (United States)

    Bekheet, Souad H M; Stahlmann, Ralf

    2009-09-01

    Spermatogenesis is a very complex process by which male germ cells differentiate into mature spermatozoa. The sophisticated communication network that controls spermatogenesis can be derailed so that dysfunction of one cell type propagates to all types as a cascade. This accounts for the particular vulnerability of the testis to environmental factors such as drugs and xenobiotics. Sertoli cells play an important role in protecting developing germ cells by forming a physiological barrier, limiting exposure to potentially toxic substrates, or conversely, facilitating uptake of xenobiotics within the testis. In this study, cells from the rat Sertoli line (SerW3) were incubated for 3, 6 and 9 subsequent days in serum free DMEM (SFDM) composed of DMEM supplemented with three different concentrations of antibiotic gentamicin (10, 30, and 100 μg). The effect of the three different concentrations of this antibiotic was determined on Sertoli cell-cell interaction through impaired expression of their constitutive tight junction proteins as early targets for different toxicants in vitro by immunochemistry analysis. The Sertoli SerW3 cell line illustrated the cytotoxicity of GS, as the intercellular junction proteins such as occludin, N-cadherin, connexin 43, and vimentin were delocalized from the membrane to the cytoplasmic compartment during exposure to the antibiotic. This study underlines the potential deleterious effects of the routine use of antibiotics during continuous cell culture.

  3. Endocochlear potential generation is associated with intercellular communication in the stria vascularis: structural analysis in the viable dominant spotting mouse mutant.

    Science.gov (United States)

    Carlisle, L; Steel, K; Forge, A

    1990-11-01

    Deafness in the viable dominant spotting mouse mutant is due to a primary defect of the stria vascularis which results in absence of the positive endocochlear potential in scala media. Endocochlear potentials were measured and the structure of stria vascularis of mutants with potentials close to zero was compared with that in normal littermate controls by use of morphometric methods. The stria vascularis was significantly thinner in mutants. Marginal cells were not significantly different from controls in terms of volume density or intramembrane particle density but the network density of tight junctions was significantly reduced in the mutants. A virtual absence of gap junctions between basal cells and marginal or intermediate cells was observed, but intramembrane particle density and junctional complexes between adjacent basal cells were not different from controls. The volume density of basal cells was significantly greater in mutants. Intermediate cells accounted for a significantly smaller volume density of the stria vascularis in mutants and had a lower density of intramembrane particles than controls. Melanocytes were not identified in the stria vascularis of mutants. These results suggest that communication between marginal, intermediate and basal cells might be important to the normal function of the stria vascularis.

  4. The functional response of bioactive titania-modified three-dimensional Ti-6Al-4V mesh structure toward providing a favorable pathway for intercellular communication and osteoincorporation.

    Science.gov (United States)

    Nune, K C; Misra, R D K; Li, S J; Hao, Y L; Zhang, W

    2016-10-01

    The objective of the study is to fundamentally elucidate the biological response of 3D printed mesh structures subjected to plasma electrolytic oxidation process through the study of osteoblast functions. The cellular activity of plasma electrolytic-oxidized mesh structure was explored in terms of cell-to-cell communication involving proliferation, synthesis of extracellular and intracellular proteins, and mineralization. Upon plasma electrolytic oxidation of the mesh structure, a thin layer of bioactive titania with pore size 1-3 µm was nucleated on the surface. The combination of microporous bioactive titania and interconnected porous architecture provided the desired pathway for supply of nutrients and oxygen to cells and tissue and a favorable osteogenic microenvironment for tissue on-growth and in-growth, in relation to the unmodified mesh structure. The formation of a confluent layer as envisaged via electron microscopy and quantitative assessment of the expression level of proteins (actin, vinculin, and fibronectin) point toward the determining role of surface-modified mesh structure in modulating osteoblasts functions. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2488-2501, 2016. © 2016 Wiley Periodicals, Inc.

  5. Short-range intercellular calcium signaling in bone

    DEFF Research Database (Denmark)

    Jørgensen, Niklas Rye

    2005-01-01

    into biological effects in bone. Intercellular calcium waves are increases in intracellular calcium concentration in single cells, subsequently propagating to adjacent cells, and can be a possible mechanism for the coupling of bone formation to bone resorption. The aim of the present studies was to investigate...... whether bone cells are capable of communicating via intercellular calcium signals, and determine by which mechanisms the cells propagate the signals. First, we found that osteoblastic cells can propagate intercellular calcium transients upon mechanical stimulation, and that there are two principally...... different mechanisms for this propagation. One mechanism involves the secretion of a nucleotide, possibly ATP, acting in an autocrine action to purinergic P2Y2 receptors on the neighboring cells, leading to intracellular IP3 generation and subsequent release of calcium from intracellular stores. The other...

  6. Short-range intercellular calcium signaling in bone

    DEFF Research Database (Denmark)

    Jørgensen, Niklas R

    2005-01-01

    The regulation of bone turnover is a complex and finely tuned process. Many factors regulate bone remodeling, including hormones, growth factors, cytokines etc. However, little is known about the signals coupling bone formation to bone resorption, and how mechanical forces are translated...... into biological effects in bone. Intercellular calcium waves are increases in intracellular calcium concentration in single cells, subsequently propagating to adjacent cells, and can be a possible mechanism for the coupling of bone formation to bone resorption. The aim of the present studies was to investigate...... whether bone cells are capable of communicating via intercellular calcium signals, and determine by which mechanisms the cells propagate the signals. First, we found that osteoblastic cells can propagate intercellular calcium transients upon mechanical stimulation, and that there are two principally...

  7. Intercellular protein-protein interactions at synapses.

    Science.gov (United States)

    Yang, Xiaofei; Hou, Dongmei; Jiang, Wei; Zhang, Chen

    2014-06-01

    Chemical synapses are asymmetric intercellular junctions through which neurons send nerve impulses to communicate with other neurons or excitable cells. The appropriate formation of synapses, both spatially and temporally, is essential for brain function and depends on the intercellular protein-protein interactions of cell adhesion molecules (CAMs) at synaptic clefts. The CAM proteins link pre- and post-synaptic sites, and play essential roles in promoting synapse formation and maturation, maintaining synapse number and type, accumulating neurotransmitter receptors and ion channels, controlling neuronal differentiation, and even regulating synaptic plasticity directly. Alteration of the interactions of CAMs leads to structural and functional impairments, which results in many neurological disorders, such as autism, Alzheimer's disease and schizophrenia. Therefore, it is crucial to understand the functions of CAMs during development and in the mature neural system, as well as in the pathogenesis of some neurological disorders. Here, we review the function of the major classes of CAMs, and how dysfunction of CAMs relates to several neurological disorders.

  8. Transmembrane Signaling Proteoglycans

    DEFF Research Database (Denmark)

    Couchman, John R

    2010-01-01

    Virtually all metazoan cells contain at least one and usually several types of transmembrane proteoglycans. These are varied in protein structure and type of polysaccharide, but the total number of vertebrate genes encoding transmembrane proteoglycan core proteins is less than 10. Some core prote...... proteins, including those of the syndecans, always possess covalently coupled glycosaminoglycans; others do not. Syndecan has a long evolutionary history, as it is present in invertebrates, but many other transmembrane proteoglycans are vertebrate inventions. The variety of proteins...... proteins has been obtained in mouse knockout experiments. Here some of the latest developments in the field are examined in hopes of stimulating further interest in this fascinating group of molecules. Expected final online publication date for the Annual Review of Cell and Developmental Biology Volume 26...

  9. Intercellular signalling in Stigmatella aurantiaca.

    Science.gov (United States)

    Plaga, W; Ulrich, S H

    1999-12-01

    The myxobacterium Stigmatella aurantiaca is a prokaryotic model used to study intercellular signalling and the genetic determination of morphogenesis. Signalling factors and genes required for the generation of the elaborate multicellular fruiting body are to be identified. Recently, the structure of stigmolone, which is the pheromone necessary for fruiting body formation, was elucidated, and genes involved in development were characterised. Progress has also been made in the genetic accessibility of S. aurantiaca.

  10. Understanding intercellular communication in the brain: Identified ...

    Indian Academy of Sciences (India)

    The transmission of information between nerve cells in the brain takes place at specialized sites of ... period of 10 days, and the giant chromosomes of larval salivary glands. ... extensive use of Drosophila in order to assay the effects of the genetic changes in ... the muscle responds only to a small percentage of the stimuli.

  11. Intercellular Ca2+ Waves: Mechanisms and Function

    Science.gov (United States)

    Sanderson, Michael J.

    2012-01-01

    Intercellular calcium (Ca2+) waves (ICWs) represent the propagation of increases in intracellular Ca2+ through a syncytium of cells and appear to be a fundamental mechanism for coordinating multicellular responses. ICWs occur in a wide diversity of cells and have been extensively studied in vitro. More recent studies focus on ICWs in vivo. ICWs are triggered by a variety of stimuli and involve the release of Ca2+ from internal stores. The propagation of ICWs predominately involves cell communication with internal messengers moving via gap junctions or extracellular messengers mediating paracrine signaling. ICWs appear to be important in both normal physiology as well as pathophysiological processes in a variety of organs and tissues including brain, liver, retina, cochlea, and vascular tissue. We review here the mechanisms of initiation and propagation of ICWs, the key intra- and extracellular messengers (inositol 1,4,5-trisphosphate and ATP) mediating ICWs, and the proposed physiological functions of ICWs. PMID:22811430

  12. The stability of the three transmembrane and the four transmembrane human vitamin K epoxide reductase models

    Science.gov (United States)

    Wu, Sangwook

    2016-04-01

    The three transmembrane and the four transmembrane helix models are suggested for human vitamin K epoxide reductase (VKOR). In this study, we investigate the stability of the human three transmembrane/four transmembrane VKOR models by employing a coarse-grained normal mode analysis and molecular dynamics simulation. Based on the analysis of the mobility of each transmembrane domain, we suggest that the three transmembrane human VKOR model is more stable than the four transmembrane human VKOR model.

  13. Methoxychlor and vinclozolin induce rapid changes in intercellular and intracellular signaling in liver progenitor cells

    Czech Academy of Sciences Publication Activity Database

    Babica, Pavel; Zurabian, R.; Kumar, E. R.; Chopra, R.; Mianecki, M. J.; Park, J.-S.; Jaša, Libor; Trosko, J. E.; Upham, B. L.

    2016-01-01

    Roč. 153, č. 1 (2016), s. 174-185 ISSN 1096-6080 R&D Projects: GA MŠk LH12034 Institutional support: RVO:67985939 Keywords : endocrine disrupters * gap junctional intercellular communication * resveratrol Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.081, year: 2016

  14. Biologically Inspired Intercellular Slot Synchronization

    Directory of Open Access Journals (Sweden)

    Alexander Tyrrell

    2009-01-01

    Full Text Available The present article develops a decentralized interbase station slot synchronization algorithm suitable for cellular mobile communication systems. The proposed cellular firefly synchronization (CelFSync algorithm is derived from the theory of pulse-coupled oscillators, common to describe synchronization phenomena in biological systems, such as the spontaneous synchronization of fireflies. In order to maintain synchronization among base stations (BSs, even when there is no direct link between adjacent BSs, some selected user terminals (UTs participate in the network synchronization process. Synchronization emerges by exchanging two distinct synchronization words, one transmitted by BSs and the other by active UTs, without any a priori assumption on the initial timing misalignments of BSs and UTs. In large-scale networks with inter-BS site distances up to a few kilometers, propagation delays severely affect the attainable timing accuracy of CelFSync. We show that by an appropriate combination of CelFSync with the timing advance procedure, which aligns uplink transmission of UTs to arrive simultaneously at the BS, a timing accuracy within a fraction of the inter-BS propagation delay is retained.

  15. Development of schizogenous intercellular spaces in plants

    Directory of Open Access Journals (Sweden)

    Kimitsune eIshizaki

    2015-07-01

    Full Text Available Gas exchange is essential for multicellular organisms. In contrast to the circulatory systems of animals, land plants have tissues with intercellular spaces (ICSs, called aerenchyma, that are critical for efficient gas exchange. Plants form ICSs by two different mechanisms: schizogeny, where localized cell separation creates spaces; and lysogeny, where cells die to create intercellular spaces. In schizogenous ICS formation, specific molecular mechanisms regulate the sites of cell separation and coordinate extensive reorganization of cell walls. Emerging evidence suggests the involvement of extracellular signaling, mediated by peptide ligands and leucine-rich repeat receptor-like kinases, in the regulation of cell wall remodeling during cell separation. Recent work on the liverwort Marchantia polymorpha has demonstrated a critical role for a plasma membrane-associated plant U-box E3 ubiquitin ligase in ICS formation. In this review, I discuss the mechanism of schizogenous ICS formation, focusing on the potential role of extracellular signaling in the regulation of cell separation.

  16. Extracellular ultrathin fibers sensitive to intracellular reactive oxygen species: Formation of intercellular membrane bridges

    Energy Technology Data Exchange (ETDEWEB)

    Jung, Se-Hui; Park, Jin-Young; Joo, Jung-Hoon; Kim, Young-Myeong; Ha, Kwon-Soo, E-mail: ksha@kangwon.ac.kr

    2011-07-15

    Membrane bridges are key cellular structures involved in intercellular communication; however, dynamics for their formation are not well understood. We demonstrated the formation and regulation of novel extracellular ultrathin fibers in NIH3T3 cells using confocal and atomic force microscopy. At adjacent regions of neighboring cells, phorbol 12-myristate 13-acetate (PMA) and glucose oxidase induced ultrathin fiber formation, which was prevented by Trolox, a reactive oxygen species (ROS) scavenger. The height of ROS-sensitive ultrathin fibers ranged from 2 to 4 nm. PMA-induced formation of ultrathin fibers was inhibited by cytochalasin D, but not by Taxol or colchicine, indicating that ultrathin fibers mainly comprise microfilaments. PMA-induced ultrathin fibers underwent dynamic structural changes, resulting in formation of intercellular membrane bridges. Thus, these fibers are formed by a mechanism(s) involving ROS and involved in formation of intercellular membrane bridges. Furthermore, ultrastructural imaging of ultrathin fibers may contribute to understanding the diverse mechanisms of cell-to-cell communication and the intercellular transfer of biomolecules, including proteins and cell organelles.

  17. The intercellular cell adhesion molecule-1 (icam-1) in lung cancer: implications for disease progression and prognosis.

    Science.gov (United States)

    Kotteas, Elias A; Boulas, Panagiotis; Gkiozos, Ioannis; Tsagkouli, Sofia; Tsoukalas, George; Syrigos, Konstantinos N

    2014-09-01

    The intercellular cell-adhesion molecule-1 (ICAM-1) is a transmembrane molecule and a distinguished member of the Immunoglobulin superfamily of proteins that participates in many important processes, including leukocyte endothelial transmigration, cell signaling, cell-cell interaction, cell polarity and tissue stability. ICAM-1and its soluble part are highly expressed in inflammatory conditions, chronic diseases and a number of malignancies. In the present article we present the implications of ICAM-1 in the progression and prognosis of one of the major global killers of our era: lung cancer. Copyright© 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  18. Role of protein dynamics in transmembrane receptor signalling

    DEFF Research Database (Denmark)

    Wang, Yong; Bugge, Katrine Østergaard; Kragelund, Birthe Brandt

    2018-01-01

    Cells are dependent on transmembrane receptors to communicate and transform chemical and physical signals into intracellular responses. Because receptors transport 'information', conformational changes and protein dynamics play a key mechanistic role. We here review examples where experiment...... to function. Because the receptors function in a heterogeneous environment and need to be able to switch between distinct functional states, they may be particularly sensitive to small perturbations that complicate studies linking dynamics to function....

  19. Serum deprivation induces glucose response and intercellular coupling in human pancreatic adenocarcinoma PANC-1 cells.

    Science.gov (United States)

    Hiram-Bab, Sahar; Shapira, Yuval; Gershengorn, Marvin C; Oron, Yoram

    2012-03-01

    This study aimed to investigate whether the previously described differentiating islet-like aggregates of human pancreatic adenocarcinoma cells (PANC-1) develop glucose response and exhibit intercellular communication. Fura 2-loaded PANC-1 cells in serum-free medium were assayed for changes in cytosolic free calcium ([Ca]i) induced by depolarization, tolbutamide inhibition of K(ATP) channels, or glucose. Dye transfer, assayed by confocal microscopy or by FACS, was used to detect intercellular communication. Changes in messenger RNA (mRNA) expression of genes of interest were assessed by quantitative real-time polymerase chain reaction. Proliferation was assayed by the MTT method. Serum-deprived PANC-1 cell aggregates developed [Ca]i response to KCl, tolbutamide, or glucose. These responses were accompanied by 5-fold increase in glucokinase mRNA level and, to a lesser extent, of mRNAs for K(ATP) and L-type calcium channels, as well as increase in mRNA levels of glucagon and somatostatin. Trypsin, a proteinase-activated receptor 2 agonist previously shown to enhance aggregation, modestly improved [Ca]i response to glucose. Glucose-induced coordinated [Ca]i oscillations and dye transfer demonstrated the emergence of intercellular communication. These findings suggest that PANC-1 cells, a pancreatic adenocarcinoma cell line, can be induced to express a differentiated phenotype in which cells exhibit response to glucose and form a functional syncytium similar to those observed in pancreatic islets.

  20. Plasmodesmata: channels for intercellular signaling during plant growth and development.

    Science.gov (United States)

    Sevilem, Iris; Yadav, Shri Ram; Helariutta, Ykä

    2015-01-01

    Plants have evolved strategies for short- and long-distance communication to coordinate plant development and to adapt to changing environmental conditions. Plasmodesmata (PD) are intercellular nanochannels that provide an effective pathway for both selective and nonselective movement of various molecules that function in diverse biological processes. Numerous non-cell-autonomous proteins (NCAP) and small RNAs have been identified that have crucial roles in cell fate determination and organ patterning during development. Both the density and aperture size of PD are developmentally regulated, allowing formation of spatial symplastic domains for establishment of tissue-specific developmental programs. The PD size exclusion limit (SEL) is controlled by reversible deposition of callose, as well as by some PD-associated proteins. Although a large number of PD-associated proteins have been identified, many of their functions remain unknown. Despite the fact that PD are primarily membranous structures, surprisingly very little is known about their lipid composition. Thus, future studies in PD biology will provide deeper insights into the high-resolution structure and tightly regulated functions of PD and the evolution of PD-mediated cell-to-cell communication in plants.

  1. Gap Junction Intercellular Communication in Bone Marrow Failure

    Science.gov (United States)

    2012-10-01

    traits are responsible for the differences in the severity of these diseases and their relation to skeletal malformations in BMF syndromes. Using a...mesenchymal transition (19-29), and are crucial in the establishment of electrical synapses in the central nervous system, heart , immune system and...developmental defective osteogenesis resulting in craniofacial and skeletal anomalies and associated with heart defects and neurological defects. ODDD

  2. Intrinsically disordered proteins aggregate at fungal cell-to-cell channels and regulate intercellular connectivity.

    Science.gov (United States)

    Lai, Julian; Koh, Chuan Hock; Tjota, Monika; Pieuchot, Laurent; Raman, Vignesh; Chandrababu, Karthik Balakrishna; Yang, Daiwen; Wong, Limsoon; Jedd, Gregory

    2012-09-25

    Like animals and plants, multicellular fungi possess cell-to-cell channels (septal pores) that allow intercellular communication and transport. Here, using a combination of MS of Woronin body-associated proteins and a bioinformatics approach that identifies related proteins based on composition and character, we identify 17 septal pore-associated (SPA) proteins that localize to the septal pore in rings and pore-centered foci. SPA proteins are not homologous at the primary sequence level but share overall physical properties with intrinsically disordered proteins. Some SPA proteins form aggregates at the septal pore, and in vitro assembly assays suggest aggregation through a nonamyloidal mechanism involving mainly α-helical and disordered structures. SPA loss-of-function phenotypes include excessive septation, septal pore degeneration, and uncontrolled Woronin body activation. Together, our data identify the septal pore as a complex subcellular compartment and focal point for the assembly of unstructured proteins controlling diverse aspects of intercellular connectivity.

  3. 1,4-Naphthoquinones: From Oxidative Damage to Cellular and Inter-Cellular Signaling

    Directory of Open Access Journals (Sweden)

    Lars-Oliver Klotz

    2014-09-01

    Full Text Available Naphthoquinones may cause oxidative stress in exposed cells and, therefore, affect redox signaling. Here, contributions of redox cycling and alkylating properties of quinones (both natural and synthetic, such as plumbagin, juglone, lawsone, menadione, methoxy-naphthoquinones, and others to cellular and inter-cellular signaling processes are discussed: (i naphthoquinone-induced Nrf2-dependent modulation of gene expression and its potentially beneficial outcome; (ii the modulation of receptor tyrosine kinases, such as the epidermal growth factor receptor by naphthoquinones, resulting in altered gap junctional intercellular communication. Generation of reactive oxygen species and modulation of redox signaling are properties of naphthoquinones that render them interesting leads for the development of novel compounds of potential use in various therapeutic settings.

  4. Biophysical Aspects of Transmembrane Signaling

    CERN Document Server

    Damjanovich, Sandor

    2005-01-01

    Transmembrane signaling is one of the most significant cell biological events in the life and death of cells in general and lymphocytes in particular. Until recently biochemists and biophysicists were not accustomed to thinking of these processes from the side of a high number of complex biochemical events and an equally high number of physical changes at molecular and cellular levels at the same time. Both types of researchers were convinced that their findings are the most decisive, having higher importance than the findings of the other scientist population. Both casts were wrong. Life, even at cellular level, has a number of interacting physical and biochemical mechanisms, which finally build up the creation of an "excited" cell that will respond to particular signals from the outer or inner world. This book handles both aspects of the signalling events, and in some cases tries to unify our concepts and help understand the signals that govern the life and death of our cells. Not only the understanding, bu...

  5. Intra- and Intercellular Quality Control Mechanisms of Mitochondria

    Directory of Open Access Journals (Sweden)

    Yoshimitsu Kiriyama

    2017-12-01

    Full Text Available Mitochondria function to generate ATP and also play important roles in cellular homeostasis, signaling, apoptosis, autophagy, and metabolism. The loss of mitochondrial function results in cell death and various types of diseases. Therefore, quality control of mitochondria via intra- and intercellular pathways is crucial. Intracellular quality control consists of biogenesis, fusion and fission, and degradation of mitochondria in the cell, whereas intercellular quality control involves tunneling nanotubes and extracellular vesicles. In this review, we outline the current knowledge on the intra- and intercellular quality control mechanisms of mitochondria.

  6. Intercellular adhesion molecules (ICAMs) and spermatogenesis

    Science.gov (United States)

    Xiao, Xiang; Mruk, Dolores D.; Cheng, C. Yan

    2013-01-01

    BACKGROUND During the seminiferous epithelial cycle, restructuring takes places at the Sertoli–Sertoli and Sertoli–germ cell interface to accommodate spermatogonia/spermatogonial stem cell renewal via mitosis, cell cycle progression and meiosis, spermiogenesis and spermiation since developing germ cells, in particular spermatids, move ‘up and down’ the seminiferous epithelium. Furthermore, preleptotene spermatocytes differentiated from type B spermatogonia residing at the basal compartment must traverse the blood–testis barrier (BTB) to enter the adluminal compartment to prepare for meiosis at Stage VIII of the epithelial cycle, a process also accompanied by the release of sperm at spermiation. These cellular events that take place at the opposite ends of the epithelium are co-ordinated by a functional axis designated the apical ectoplasmic specialization (ES)—BTB—basement membrane. However, the regulatory molecules that co-ordinate cellular events in this axis are not known. METHODS Literature was searched at http://www.pubmed.org and http://scholar.google.com to identify published findings regarding intercellular adhesion molecules (ICAMs) and the regulation of this axis. RESULTS Members of the ICAM family, namely ICAM-1 and ICAM-2, and the biologically active soluble ICAM-1 (sICAM-1) are the likely regulatory molecules that co-ordinate these events. sICAM-1 and ICAM-1 have antagonistic effects on the Sertoli cell tight junction-permeability barrier, involved in Sertoli cell BTB restructuring, whereas ICAM-2 is restricted to the apical ES, regulating spermatid adhesion during the epithelial cycle. Studies in other epithelia/endothelia on the role of the ICAM family in regulating cell movement are discussed and this information has been evaluated and integrated into studies of these proteins in the testis to create a hypothetical model, depicting how ICAMs regulate junction restructuring events during spermatogenesis. CONCLUSIONS ICAMs are crucial

  7. Modelling of intercellular synchronization in the Drosophila circadian clock

    International Nuclear Information System (INIS)

    Jun-Wei, Wang; Ai-Min, Chen; Jia-Jun, Zhang; Zhan-Jiang, Yuan; Tian-Shou, Zhou

    2009-01-01

    In circadian rhythm generation, intercellular signaling factors are shown to play a crucial role in both sustaining intrinsic cellular rhythmicity and acquiring collective behaviours across a population of circadian neurons. However, the physical mechanism behind their role remains to be fully understood. In this paper, we propose an indirectly coupled multicellular model for the synchronization of Drosophila circadian oscillators combining both intracellular and intercellular dynamics. By simulating different experimental conditions, we find that such an indirect coupling way can synchronize both heterogeneous self-sustained circadian neurons and heterogeneous mutational damped circadian neurons. Moreover, they can also be entrained to ambient light-dark (LD) cycles depending on intercellular signaling. (cross-disciplinary physics and related areas of science and technology)

  8. Phloroglucinol functions as an intracellular and intercellular chemical messenger influencing gene expression in Pseudomonas protegens.

    Science.gov (United States)

    Clifford, Jennifer C; Buchanan, Alex; Vining, Oliver; Kidarsa, Teresa A; Chang, Jeff H; McPhail, Kerry L; Loper, Joyce E

    2016-10-01

    Bacteria can be both highly communicative and highly competitive in natural habitats and antibiotics are thought to play a role in both of these processes. The soil bacterium Pseudomonas protegens Pf-5 produces a spectrum of antibiotics, two of which, pyoluteorin and 2,4-diacetylphloroglucinol (DAPG), function in intracellular and intercellular communication, both as autoinducers of their own production. Here, we demonstrate that phloroglucinol, an intermediate in DAPG biosynthesis, can serve as an intercellular signal influencing the expression of pyoluteorin biosynthesis genes, the production of pyoluteorin, and inhibition of Pythium ultimum, a phytopathogenic oomycete sensitive to pyoluteorin. Through analysis of RNAseq data sets, we show that phloroglucinol had broad effects on the transcriptome of Pf-5, significantly altering the transcription of more than two hundred genes. The effects of nanomolar versus micromolar concentrations of phloroglucinol differed both quantitatively and qualitatively, influencing the expression of distinct sets of genes or having opposite effects on transcript abundance of certain genes. Therefore, our results support the concept of hormesis, a phenomenon associated with signalling molecules that elicit distinct responses at different concentrations. Phloroglucinol is the first example of an intermediate of antibiotic biosynthesis that functions as a chemical messenger influencing gene expression in P. protegens. © 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.

  9. Exploring the role of lipids in intercellular conduits: breakthroughs in the pipeline

    Directory of Open Access Journals (Sweden)

    Elise eDelage

    2013-12-01

    Full Text Available It has been known for more than a century that most of the plant cells are connected to their neighbors through membranous pores perforating the cell wall, namely plasmodesmata (PDs. The recent discovery of tunneling nanotubes (TNTs, thin membrane bridges established between distant mammalian cells, suggests that intercellular communication mediated through cytoplasmic continuity could be a conserved feature of eukaryotic organisms. Although TNTs differ from PDs in their formation and architecture, both are characterized by a continuity of the plasma membrane between two cells, delimiting a nanotubular channel supported by actin-based cytoskeleton. Due to this unusual membrane organization, lipids are likely to play critical roles in the formation and stability of intercellular conduits like TNTs and PDs, but also in regulating the transfer through these structures. While it is crucial for a better understanding of those fascinating communication highways, the study of TNT lipid composition and dynamics turned out to be extremely challenging. The present review aims to give an overview of the recent findings in this context. We will also discuss some of the promising imaging approaches, which might be the key for future breakthroughs in the field and could also benefit the research on PDs.

  10. Laminin-332 alters connexin profile, dye coupling and intercellular Ca2+ waves in ciliated tracheal epithelial cells

    Directory of Open Access Journals (Sweden)

    Olsen Colin E

    2006-08-01

    Full Text Available Abstract Background Tracheal epithelial cells are anchored to a dynamic basement membrane that contains a variety of extracellular matrix proteins including collagens and laminins. During development, wound repair and disease of the airway epithelium, significant changes in extracellular matrix proteins may directly affect cell migration, differentiation and events mediated by intercellular communication. We hypothesized that alterations in cell matrix, specifically type I collagen and laminin α3β3γ2 (LM-332 proteins within the matrix, directly affect intercellular communication in ciliated rabbit tracheal epithelial cells (RTEC. Methods Functional coupling of RTEC was monitored by microinjection of the negatively charged fluorescent dyes, Lucifer Yellow and Alexa 350, into ciliated RTEC grown on either a LM-332/collagen or collagen matrix. Coupling of physiologically significant molecules was evaluated by the mechanism and extent of propagated intercellular Ca2+ waves. Expression of connexin (Cx mRNA and proteins were assayed by reverse transcriptase – polymerase chain reaction and immunocytochemistry, respectively. Results When compared to RTEC grown on collagen alone, RTEC grown on LM-332/collagen displayed a significant increase in dye transfer. Although mechanical stimulation of RTEC grown on either LM-332/collagen or collagen alone resulted in intercellular Ca2+ waves, the mechanism of transfer was dependent on matrix: RTEC grown on LM-332/collagen propagated Ca2+waves via extracellular purinergic signaling whereas RTEC grown on collagen used gap junctions. Comparison of RTEC grown on collagen or LM-332/collagen matrices revealed a reorganization of Cx26, Cx43 and Cx46 proteins. Conclusion Alterations in airway basement membrane proteins such as LM-332 can induce connexin reorganizations and result in altered cellular communication mechanisms that could contribute to airway tissue function.

  11. Phosphatidylinositol-bisphosphate regulates intercellular coupling in cardiac myocytes

    DEFF Research Database (Denmark)

    Hofgaard, Johannes P; Banach, Kathrin; Mollerup, Sarah

    2008-01-01

    that agonist-induced changes in PIP(2) can result in a reduction of the functional coupling of cardiomyocytes and, consequently, in changes in conduction velocity. Intercellular coupling was measured by Lucifer Yellow dye transfer in cultured neonatal rat cardiomyocytes. Conduction velocity was measured...

  12. Levels Of Serum Intercellular And Vascular Adhesion Molecules In ...

    African Journals Online (AJOL)

    The study evaluated the possible significant role of soluble intercellular and vascular adhesion molecule-1 (sICAM-1 and sVCAM-1), sE-selectin and interluekin-1β in development nephropathy in patients with insulin dependent diabetes mellitus (IDDM). This study included 60 patients with type 1 diabetes mellitus (IDDM) ...

  13. Identification of MarvelD3 as a tight junction-associated transmembrane protein of the occludin family

    Directory of Open Access Journals (Sweden)

    Balda Maria S

    2009-12-01

    Full Text Available Abstract Background Tight junctions are an intercellular adhesion complex of epithelial and endothelial cells, and form a paracellular barrier that restricts the diffusion of solutes on the basis of size and charge. Tight junctions are formed by multiprotein complexes containing cytosolic and transmembrane proteins. How these components work together to form functional tight junctions is still not well understood and will require a complete understanding of the molecular composition of the junction. Results Here we identify a new transmembrane component of tight junctions: MarvelD3, a four-span transmembrane protein. Its predicted transmembrane helices form a Marvel (MAL and related proteins for vesicle traffic and membrane link domain, a structural motif originally discovered in proteins involved in membrane apposition and fusion events, such as the tight junction proteins occludin and tricellulin. In mammals, MarvelD3 is expressed as two alternatively spliced isoforms. Both isoforms exhibit a broad tissue distribution and are expressed by different types of epithelial as well as endothelial cells. MarvelD3 co-localises with occludin at tight junctions in intestinal and corneal epithelial cells. RNA interference experiments in Caco-2 cells indicate that normal MarvelD3 expression is not required for the formation of functional tight junctions but depletion results in monolayers with increased transepithelial electrical resistance. Conclusions Our data indicate that MarvelD3 is a third member of the tight junction-associated occludin family of transmembrane proteins. Similar to occludin, normal expression of MarvelD3 is not essential for the formation of functional tight junctions. However, MarvelD3 functions as a determinant of epithelial paracellular permeability properties.

  14. Activation of L-type calcium channels is required for gap junction-mediated intercellular calcium signaling in osteoblastic cells

    DEFF Research Database (Denmark)

    Jørgensen, Niklas Rye; Teilmann, Stefan Cuoni; Henriksen, Zanne

    2003-01-01

    The propagation of mechanically induced intercellular calcium waves (ICW) among osteoblastic cells occurs both by activation of P2Y (purinergic) receptors by extracellular nucleotides, resulting in "fast" ICW, and by gap junctional communication in cells that express connexin43 (Cx43), resulting...... in "slow" ICW. Human osteoblastic cells transmit intercellular calcium signals by both of these mechanisms. In the current studies we have examined the mechanism of slow gap junction-dependent ICW in osteoblastic cells. In ROS rat osteoblastic cells, gap junction-dependent ICW were inhibited by removal...... of extracellular calcium, plasma membrane depolarization by high extracellular potassium, and the L-type voltage-operated calcium channel inhibitor, nifedipine. In contrast, all these treatments enhanced the spread of P2 receptor-mediated ICW in UMR rat osteoblastic cells. Using UMR cells transfected to express Cx...

  15. Drosophila wing imaginal discs respond to mechanical injury via slow InsP3R-mediated intercellular calcium waves

    Science.gov (United States)

    Restrepo, Simon; Basler, Konrad

    2016-08-01

    Calcium signalling is a highly versatile cellular communication system that modulates basic functions such as cell contractility, essential steps of animal development such as fertilization and higher-order processes such as memory. We probed the function of calcium signalling in Drosophila wing imaginal discs through a combination of ex vivo and in vivo imaging and genetic analysis. Here we discover that wing discs display slow, long-range intercellular calcium waves (ICWs) when mechanically stressed in vivo or cultured ex vivo. These slow imaginal disc intercellular calcium waves (SIDICs) are mediated by the inositol-3-phosphate receptor, the endoplasmic reticulum (ER) calcium pump SERCA and the key gap junction component Inx2. The knockdown of genes required for SIDIC formation and propagation negatively affects wing disc recovery after mechanical injury. Our results reveal a role for ICWs in wing disc homoeostasis and highlight the utility of the wing disc as a model for calcium signalling studies.

  16. Electrotonic potentials in Aloe vera L.: Effects of intercellular and external electrodes arrangement.

    Science.gov (United States)

    Volkov, Alexander G; Nyasani, Eunice K; Tuckett, Clayton; Scott, Jessenia M; Jackson, Mariah M Z; Greeman, Esther A; Greenidge, Ariane S; Cohen, Devin O; Volkova, Maia I; Shtessel, Yuri B

    2017-02-01

    Electrostimulation of plants can induce plant movements, activation of ion channels, ion transport, gene expression, enzymatic systems activation, electrical signaling, plant-cell damage, enhanced wound healing, and influence plant growth. Here we found that electrical networks in plant tissues have electrical differentiators. The amplitude of electrical responses decreases along a leaf and increases by decreasing the distance between polarizing Pt-electrodes. Intercellular Ag/AgCl electrodes inserted in a leaf and extracellular Ag/AgCl electrodes attached to the leaf surface were used to detect the electrotonic potential propagation along a leaf of Aloe vera. There is a difference in duration and amplitude of electrical potentials measured by electrodes inserted in a leaf and those attached to a leaf's surface. If the external reference electrode is located in the soil near the root, it changes the amplitude and duration of electrotonic potentials due to existence of additional resistance, capacitance, ion channels and ion pumps in the root. The information gained from this study can be used to elucidate extracellular and intercellular communication in the form of electrical signals within plants. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. Multicellular models of intercellular synchronization in circadian neural networks

    International Nuclear Information System (INIS)

    Henson, Michael A.

    2013-01-01

    The circadian clock generates 24 h rhythms that drive physiological and behavioral processes in a diverse range of organisms including microbes, plants, insects, and mammals. Recent experimental advances have produced improved understanding of the molecular mechanisms involved in circadian rhythm generation at the single cell level. However, the intercellular mechanisms that allow large populations of coupled pacemaker cells to synchronize and coordinate their rhythms remain poorly understood. The purpose of this article is to review recent progress in dynamic modeling of the circadian clock with a focus on multicellular models required to describe cell population synchronization. Mammalian systems are emphasized to illustrate the highly heterogeneous structure and rich dynamical behavior of multicellular circadian systems. Available multicellular models are characterized with respect to their single cell descriptions, intercellular coupling mechanisms, and network topologies. Examples drawn from our own research are used to demonstrate the advantages associated with integrating detailed single cell models within realistic multicellular networks for prediction of mammalian system dynamics. Mathematical modeling is shown to represent a powerful tool for understanding the intracellular and intercellular mechanisms utilized to robustly synchronize large populations of highly heterogeneous and sparsely coupled single cell oscillators. The article concludes with some possible directions for future research

  18. Evolution of altruism in spatial prisoner's dilemma: Intra- and inter-cellular interactions

    Science.gov (United States)

    Yokoi, Hiroki; Uehara, Takashi; Sakata, Tomoyuki; Naito, Hiromi; Morita, Satoru; Tainaka, Kei-ichi

    2014-12-01

    Iterated prisoner's dilemma game is carried out on lattice with “colony” structure. Each cell is regarded as a colony which contains plural players with an identical strategy. Both intra- and inter-cellular interactions are assumed. In the former a player plays with all other players in the same colony, while in the latter he plays with one player each from adjacent colonies. Spatial patterns among four typical strategies exhibit various dynamics and winners. Both theory and simulation reveal that All Cooperation (AC) wins, when the members of colony or the intensity of noise increases. This result explains the evolution of altruism in animal societies, even though errors easily occur in animal communications.

  19. ATP- and gap junction-dependent intercellular calcium signaling in osteoblastic cells

    DEFF Research Database (Denmark)

    Jorgensen, N R; Geist, S T; Civitelli, R

    1997-01-01

    mechanically induced calcium waves in two rat osteosarcoma cell lines that differ in the gap junction proteins they express, in their ability to pass microinjected dye from cell to cell, and in their expression of P2Y2 (P2U) purinergic receptors. ROS 17/2.8 cells, which express the gap junction protein......Many cells coordinate their activities by transmitting rises in intracellular calcium from cell to cell. In nonexcitable cells, there are currently two models for intercellular calcium wave propagation, both of which involve release of inositol trisphosphate (IP3)- sensitive intracellular calcium...... stores. In one model, IP3 traverses gap junctions and initiates the release of intracellular calcium stores in neighboring cells. Alternatively, calcium waves may be mediated not by gap junctional communication, but rather by autocrine activity of secreted ATP on P2 purinergic receptors. We studied...

  20. Membrane shape modulates transmembrane protein distribution.

    Science.gov (United States)

    Aimon, Sophie; Callan-Jones, Andrew; Berthaud, Alice; Pinot, Mathieu; Toombes, Gilman E S; Bassereau, Patricia

    2014-01-27

    Although membrane shape varies greatly throughout the cell, the contribution of membrane curvature to transmembrane protein targeting is unknown because of the numerous sorting mechanisms that take place concurrently in cells. To isolate the effect of membrane shape, we used cell-sized giant unilamellar vesicles (GUVs) containing either the potassium channel KvAP or the water channel AQP0 to form membrane nanotubes with controlled radii. Whereas the AQP0 concentrations in flat and curved membranes were indistinguishable, KvAP was enriched in the tubes, with greater enrichment in more highly curved membranes. Fluorescence recovery after photobleaching measurements showed that both proteins could freely diffuse through the neck between the tube and GUV, and the effect of each protein on membrane shape and stiffness was characterized using a thermodynamic sorting model. This study establishes the importance of membrane shape for targeting transmembrane proteins and provides a method for determining the effective shape and flexibility of membrane proteins. Copyright © 2014 Elsevier Inc. All rights reserved.

  1. Evolution of vertebrate interferon inducible transmembrane proteins

    Directory of Open Access Journals (Sweden)

    Hickford Danielle

    2012-04-01

    Full Text Available Abstract Background Interferon inducible transmembrane proteins (IFITMs have diverse roles, including the control of cell proliferation, promotion of homotypic cell adhesion, protection against viral infection, promotion of bone matrix maturation and mineralisation, and mediating germ cell development. Most IFITMs have been well characterised in human and mouse but little published data exists for other animals. This study characterised IFITMs in two distantly related marsupial species, the Australian tammar wallaby and the South American grey short-tailed opossum, and analysed the phylogeny of the IFITM family in vertebrates. Results Five IFITM paralogues were identified in both the tammar and opossum. As in eutherians, most marsupial IFITM genes exist within a cluster, contain two exons and encode proteins with two transmembrane domains. Only two IFITM genes, IFITM5 and IFITM10, have orthologues in both marsupials and eutherians. IFITM5 arose in bony fish and IFITM10 in tetrapods. The bone-specific expression of IFITM5 appears to be restricted to therian mammals, suggesting that its specialised role in bone production is a recent adaptation specific to mammals. IFITM10 is the most highly conserved IFITM, sharing at least 85% amino acid identity between birds, reptiles and mammals and suggesting an important role for this presently uncharacterised protein. Conclusions Like eutherians, marsupials also have multiple IFITM genes that exist in a gene cluster. The differing expression patterns for many of the paralogues, together with poor sequence conservation between species, suggests that IFITM genes have acquired many different roles during vertebrate evolution.

  2. Specificity of transmembrane protein palmitoylation in yeast.

    Directory of Open Access Journals (Sweden)

    Ayelén González Montoro

    Full Text Available Many proteins are modified after their synthesis, by the addition of a lipid molecule to one or more cysteine residues, through a thioester bond. This modification is called S-acylation, and more commonly palmitoylation. This reaction is carried out by a family of enzymes, called palmitoyltransferases (PATs, characterized by the presence of a conserved 50- aminoacids domain called "Asp-His-His-Cys- Cysteine Rich Domain" (DHHC-CRD. There are 7 members of this family in the yeast Saccharomyces cerevisiae, and each of these proteins is thought to be responsible for the palmitoylation of a subset of substrates. Substrate specificity of PATs, however, is not yet fully understood. Several yeast PATs seem to have overlapping specificity, and it has been proposed that the machinery responsible for palmitoylating peripheral membrane proteins in mammalian cells, lacks specificity altogether.Here we investigate the specificity of transmembrane protein palmitoylation in S. cerevisiae, which is carried out predominantly by two PATs, Swf1 and Pfa4. We show that palmitoylation of transmembrane substrates requires dedicated PATs, since other yeast PATs are mostly unable to perform Swf1 or Pfa4 functions, even when overexpressed. Furthermore, we find that Swf1 is highly specific for its substrates, as it is unable to substitute for other PATs. To identify where Swf1 specificity lies, we carried out a bioinformatics survey to identify amino acids responsible for the determination of specificity or Specificity Determination Positions (SDPs and showed experimentally, that mutation of the two best SDP candidates, A145 and K148, results in complete and partial loss of function, respectively. These residues are located within the conserved catalytic DHHC domain suggesting that it could also be involved in the determination of specificity. Finally, we show that modifying the position of the cysteines in Tlg1, a Swf1 substrate, results in lack of palmitoylation, as

  3. Proteomic analysis of exosomes from nasopharyngeal carcinoma cell identifies intercellular transfer of angiogenic proteins

    KAUST Repository

    Chan, Yuk-kit

    2015-04-01

    Exosomes, a group of secreted extracellular nanovesicles containing genetic materials and signaling molecules, play a critical role in intercellular communication. During tumorigenesis, exosomes have been demonstrated to promote tumor angiogenesis and metastasis while their biological functions in nasopharyngeal carcinoma (NPC) are poorly understood. In this study, we focused on the role of NPC-derived exosomes on angiogenesis. Exosomes derived from the NPC C666-1 cells and immortalized nasopharyngeal epithelial cells (NP69 and NP460) were isolated using ultracentrifugation. The molecular profile and biophysical characteristics of exosomes were verified by Western blotting, sucrose density gradient, and electron microscopy. We showed that the C666-1 exosomes (10 and 20 μg/ml) could significantly increase the tubulogenesis, migration and invasion of human umbilical vein endothelial cells (HUVECs) in a dose-dependent manner. Subsequently, an iTRAQ-based quantitative proteomics was used to identify the differentially expressed proteins in C666-1 exosomes. Among the 640 identified proteins, 51 and 89 proteins were considered as up- and down-regulated (≥ 1.5-fold variations) in C666-1 exosomes compared to the normal counterparts, respectively. As expected, pro-angiogenic proteins including intercellular adhesion molecule-1 (ICAM-1) and CD44 variant isoform 5 (CD44v5) are among the up-regulated proteins, whereas angio-suppressive protein, thrombospondin-1 (TSP-1) was down-regulated in C666-1 exosomes. Further confocal microscopic study and Western blotting clearly demonstrated that the alteration of ICAM-1, and TSP-1 expressions in recipient HUVECs are due to internalization of exosomes. Taken together, these data strongly indicated the critical roles of identified angiogenic proteins in the involvement of exosomes-induced angiogenesis, which could potentially be developed as therapeutic targets in future. This article is protected by copyright. All rights reserved.

  4. Methoxychlor and Vinclozolin Induce Rapid Changes in Intercellular and Intracellular Signaling in Liver Progenitor Cells.

    Science.gov (United States)

    Babica, Pavel; Zurabian, Rimma; Kumar, Esha R; Chopra, Rajus; Mianecki, Maxwell J; Park, Joon-Suk; Jaša, Libor; Trosko, James E; Upham, Brad L

    2016-09-01

    Methoxychlor (MXC) and vinclozolin (VIN) are well-recognized endocrine disrupting chemicals known to alter epigenetic regulations and transgenerational inheritance; however, non-endocrine disruption endpoints are also important. Thus, we determined the effects of MXC and VIN on the dysregulation of gap junctional intercellular communication (GJIC) and activation of mitogen-activated protein kinases (MAPKs) in WB-F344 rat liver epithelial cells. Both chemicals induced a rapid dysregulation of GJIC at non-cytotoxic doses, with 30 min EC50 values for GJIC inhibition being 10 µM for MXC and 126 µM for VIN. MXC inhibited GJIC for at least 24 h, while VIN effects were transient and GJIC recovered after 4 h. VIN induced rapid hyperphosphorylation and internalization of gap junction protein connexin43, and both chemicals also activated MAPK ERK1/2 and p38. Effects on GJIC were not prevented by MEK1/2 inhibitor, but by an inhibitor of phosphatidylcholine-specific phospholipase C (PC-PLC), resveratrol, and in the case of VIN, also, by a p38 inhibitor. Estrogen (ER) and androgen receptor (AR) modulators (estradiol, ICI 182,780, HPTE, testosterone, flutamide, VIN M2) did not attenuate MXC or VIN effects on GJIC. Our data also indicate that the effects were elicited by the parental compounds of MXC and VIN. Our study provides new evidence that MXC and VIN dysregulate GJIC via mechanisms involving rapid activation of PC-PLC occurring independently of ER- or AR-dependent genomic signaling. Such alterations of rapid intercellular and intracellular signaling events involved in regulations of gene expression, tissue development, function and homeostasis, could also contribute to transgenerational epigenetic effects of endocrine disruptors. © The Author 2016. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  5. Regulation of intercellular tight junctions by zonula occludens toxin and its eukaryotic analogue zonulin.

    Science.gov (United States)

    Fasano, A

    2000-01-01

    The intestinal epithelium represents the largest interface between the external environment and the internal host milieu and constitutes the major barrier through which molecules can either be absorbed or secreted. There is now substantial evidence that tight junctions (tj) play a major role in regulating epithelial permeability by influencing paracellular flow of fluid and solutes. Tj are one of the hallmarks of absorptive and secretory epithelia. Evidence now exists that tj are dynamic rather than static structures and readily adapt to a variety of developmental, physiological, and pathological circumstances. These adaptive mechanisms are still incompletely understood. Activation of PKC either by Zonula occludens toxin (Zot) or by phorbol esters increases paracellular permeability. Alteration of epithelial tj is a recently described property for infectious agents. Clostridium difficile toxin A and B and influenza and vesicular stomatitis viruses have been shown to loosen tj in tissue culture monolayers. Unlike what occurs after the Zot stimulus, these changes appear to be irreversible and are associated with destruction of the tj complex. On the basis of this observation, we postulated that Zot may mimic the effect of a functionally and immunologically related endogenous modulator of epithelial tj. We were able to identify an intestinal Zot analogue, which we named zonulin. It is conceivable that the zonulins participate in the physiological regulation of intercellular tj not only in the small intestine, but also throughout a wide range of extraintestinal epithelia as well as the ubiquitous vascular endothelium, including the blood-brain barrier. Disregulation of this hypothetical zonulin model may contribute to disease states that involve disordered intercellular communication, including developmental and intestinal disorders, tissue inflammation, malignant transformation, and metastasis.

  6. Proteomic analysis of exosomes from nasopharyngeal carcinoma cell identifies intercellular transfer of angiogenic proteins

    KAUST Repository

    Chan, Yuk-kit; Zhang, Huoming; Liu, Pei; Tsao, George Sai-wah; Li Lung, Maria; Mak, Nai-ki; Ngok-shun Wong, Ricky; Ying-kit Yue, Patrick

    2015-01-01

    Exosomes, a group of secreted extracellular nanovesicles containing genetic materials and signaling molecules, play a critical role in intercellular communication. During tumorigenesis, exosomes have been demonstrated to promote tumor angiogenesis and metastasis while their biological functions in nasopharyngeal carcinoma (NPC) are poorly understood. In this study, we focused on the role of NPC-derived exosomes on angiogenesis. Exosomes derived from the NPC C666-1 cells and immortalized nasopharyngeal epithelial cells (NP69 and NP460) were isolated using ultracentrifugation. The molecular profile and biophysical characteristics of exosomes were verified by Western blotting, sucrose density gradient, and electron microscopy. We showed that the C666-1 exosomes (10 and 20 μg/ml) could significantly increase the tubulogenesis, migration and invasion of human umbilical vein endothelial cells (HUVECs) in a dose-dependent manner. Subsequently, an iTRAQ-based quantitative proteomics was used to identify the differentially expressed proteins in C666-1 exosomes. Among the 640 identified proteins, 51 and 89 proteins were considered as up- and down-regulated (≥ 1.5-fold variations) in C666-1 exosomes compared to the normal counterparts, respectively. As expected, pro-angiogenic proteins including intercellular adhesion molecule-1 (ICAM-1) and CD44 variant isoform 5 (CD44v5) are among the up-regulated proteins, whereas angio-suppressive protein, thrombospondin-1 (TSP-1) was down-regulated in C666-1 exosomes. Further confocal microscopic study and Western blotting clearly demonstrated that the alteration of ICAM-1, and TSP-1 expressions in recipient HUVECs are due to internalization of exosomes. Taken together, these data strongly indicated the critical roles of identified angiogenic proteins in the involvement of exosomes-induced angiogenesis, which could potentially be developed as therapeutic targets in future. This article is protected by copyright. All rights reserved.

  7. Identification and characterization of RBM44 as a novel intercellular bridge protein.

    Directory of Open Access Journals (Sweden)

    Tokuko Iwamori

    2011-02-01

    Full Text Available Intercellular bridges are evolutionarily conserved structures that connect differentiating germ cells. We previously reported the identification of TEX14 as the first essential intercellular bridge protein, the demonstration that intercellular bridges are required for male fertility, and the finding that intercellular bridges utilize components of the cytokinesis machinery to form. Herein, we report the identification of RNA binding motif protein 44 (RBM44 as a novel germ cell intercellular bridge protein. RBM44 was identified by proteomic analysis after intercellular bridge enrichment using TEX14 as a marker protein. RBM44 is highly conserved between mouse and human and contains an RNA recognition motif of unknown function. RBM44 mRNA is enriched in testis, and immunofluorescence confirms that RBM44 is an intercellular bridge component. However, RBM44 only partially localizes to TEX14-positive intercellular bridges. RBM44 is expressed most highly in pachytene and secondary spermatocytes, but disappears abruptly in spermatids. We discovered that RBM44 interacts with itself and TEX14 using yeast two-hybrid, mammalian two-hybrid, and immunoprecipitation. To define the in vivo function of RBM44, we generated a targeted deletion of Rbm44 in mice. Rbm44 null male mice produce somewhat increased sperm, and show enhanced fertility of unknown etiology. Thus, although RBM44 localizes to intercellular bridges during meiosis, RBM44 is not required for fertility in contrast to TEX14.

  8. COMMUNICATIONS

    CERN Multimedia

    L. Taylor and D. Barney

    2010-01-01

    CMS Centres, Outreach and the 7 TeV Media Event The new CMS Communications group is now established and is addressing three areas that are critical to CMS as it enters the physics operations phase: - Communications Infrastructure, including almost 50 CMS Centres Worldwide, videoconferencing systems, and CERN meeting rooms - Information systems, including the internal and external Web sites as well as the document preparation and management systems - Outreach and Education activities, including working with print, radio and TV media, visits to CMS, and exhibitions. The group has been active in many areas, with the highest priority being accorded to needs of CMS operations and preparations for the major media event planned for 7 TeV collisions. Unfortunately the CMS Centre@CERN suffered a major setback when, on 21st December, a cooling water pipe froze and burst on the floor above the CMS Centre main room. Water poured through the ceiling, flooding the floor and soaking some of the consoles, before e...

  9. COMMUNICATIONS

    CERN Multimedia

    A. Petrilli

    2013-01-01

    The organisation of the Open Days at the end of September was the single biggest effort of the CMS Communications Group this year. We would like to thank all volunteers for their hard work to show our Point 5 facilities and explain science and technology to the general public. During two days more than 5,000 people visited the CMS detector underground and profited from the surface activities, which included an exhibition on CMS, a workshop on superconductivity, and an activity for our younger visitors involving wooden Kapla blocks. The Communications Group took advantage of the preparations to produce new CMS posters that can be reused at other venues. Event display images have been produced not just for this occasion but also for other exhibits, education purposes, publications etc. During the Open Days, Gilles Jobin, 2012 winner of CERN Collide@CERN prize, performed his Quantum show in Point 5, with the light installation of German artist Julius von Bismarck. Image 3: CERN Open Days at CMS wel...

  10. Control of phospholipid flip-flop by transmembrane peptides

    International Nuclear Information System (INIS)

    Kaihara, Masanori; Nakao, Hiroyuki; Yokoyama, Hirokazu; Endo, Hitoshi; Ishihama, Yasushi; Handa, Tetsurou; Nakano, Minoru

    2013-01-01

    Highlights: ► Phospholipid flip-flop in transmembrane peptide-containing vesicles was investigated. ► Peptides that contained polar residues in the center of the transmembrane region promoted phospholipid flip-flop. ► A bioinformatics approach revealed the presence of polar residues in the transmembrane region of ER membrane proteins. ► Polar residues in ER membrane proteins possibly provide flippase-like activity. - Abstract: We designed three types of transmembrane model peptides whose sequence originates from a frequently used model peptide KALP23, and we investigated their effects on phospholipid flip-flop. Time-resolved small-angle neutron scattering and a dithionite fluorescent quenching assay demonstrated that TMP-L, which has a fully hydrophobic transmembrane region, did not enhance phospholipid flip-flop, whereas TMP-K and TMP-E, which have Lys and Glu, respectively, in the center of their transmembrane regions, enhanced phospholipid flip-flop. Introduction of polar residues in the membrane-spanning helices is considered to produce a locally polar region and enable the lipid head group to interact with the polar side-chain inside the bilayers, thereby reducing the activation energy for the flip-flop. A bioinformatics approach revealed that acidic and basic residues account for 4.5% of the central region of the transmembrane domain in human ER membrane proteins. Therefore, polar residues in ER membrane proteins are considered to provide flippase-like activity

  11. On the involvement of host proteins in Cowpea mosaic virus intercellular spread

    NARCIS (Netherlands)

    Hollander, den P.W.

    2014-01-01

    Abstract of thesis Paulus den Hollander entitled “On the involvement of host proteins in Cowpea mosaic virus intercellular spread”.

    Defence: 18th of November 13.30 h

    Abstract

    Intercellular spread of Cowpea mosaic virus (CPMV) occurs via movement

  12. Proteins mediating intra- and intercellular transport of lipids and lipid-modified proteins

    NARCIS (Netherlands)

    Neumann, S.

    2008-01-01

    Proteins mediating intra- and intercellular transport of lipids and lipid-modified proteins In this thesis, I studied the intra- and intercellular transport of lipidic molecules, in particular glycosphingolipids and lipid-modified proteins. The first part focuses on the intracellular transport of

  13. Compression induced intercellular shaping for some geometric cellular lattices

    Directory of Open Access Journals (Sweden)

    Adonai Gimenez Calbo

    2001-03-01

    Full Text Available The wall perimeter fraction, which contact neighboring cells, was named compression ratio (alpha. A zero compression ratio indicates maximum intercellular (air volume (vG, v/v and neglectable contact among cells, while alpha=1 indicates complete adherence between neighboring cells and no vG in the lattice. The maximum intercellular air volume (beta, v/v, when alpha=0, was 0.593 for triangular, 0.2146 for square and 0,0931 for hexagonal lattices. The equation alpha=1- (vG/beta½ was derived to relate alpha, beta and vG in the studied lattices. The relation (P S=p/alpha between cell turgor (P S and the tissue aggregating pressure (p, defined as the compression to keep in place a layer of cells, was demonstrated using the compression ratio concept. Intercellular deformations of Ipomea batatas L. roots obtained with pressure chamber were used to test alpha, vG, p and P S as a function of compression. Volumetric and transversal elastic extensibilities and the lamella media tearing forces were obtained and alpha constancy was considered as a criteria of cellular shape stability.A fração do perímetro da parede celular em contato com células vizinha foi denominada razão de compressão (alfa. Razão de compressão zero indica volume intercelular (vG, v/v máximo e contato neglível entre as células, enquanto alfa=1 ocorre quando há completa aderência com as células vizinhas (vG=0. O volume (gasoso intercelular máximo (beta, v/v, quando alfa=0, foi 0,593, 0,2146 e 0,0931 para látices triangulares, quadradas e hexagonais. A equação derivada para relacionar alfa, beta and vG nas látices estudadas foi alfa=1- (vG/beta½. A razão de compressão foi em seguida empregada para estabelecer a relação P S=p/alfa entre a pressão de turgescência (P S e a pressão de agregação (p, definida com a compressão para manter uma camada de células no seu lugar. As deformações intercelulares de batata-doce obtidas com procedimentos de c

  14. Analysis of trafficking, stability and function of human connexin 26 gap junction channels with deafness-causing mutations in the fourth transmembrane helix.

    Directory of Open Access Journals (Sweden)

    Cinzia Ambrosi

    Full Text Available Human Connexin26 gene mutations cause hearing loss. These hereditary mutations are the leading cause of childhood deafness worldwide. Mutations in gap junction proteins (connexins can impair intercellular communication by eliminating protein synthesis, mis-trafficking, or inducing channels that fail to dock or have aberrant function. We previously identified a new class of mutants that form non-functional gap junction channels and hemichannels (connexons by disrupting packing and inter-helix interactions. Here we analyzed fourteen point mutations in the fourth transmembrane helix of connexin26 (Cx26 that cause non-syndromic hearing loss. Eight mutations caused mis-trafficking (K188R, F191L, V198M, S199F, G200R, I203K, L205P, T208P. Of the remaining six that formed gap junctions in mammalian cells, M195T and A197S formed stable hemichannels after isolation with a baculovirus/Sf9 protein purification system, while C202F, I203T, L205V and N206S formed hemichannels with varying degrees of instability. The function of all six gap junction-forming mutants was further assessed through measurement of dye coupling in mammalian cells and junctional conductance in paired Xenopus oocytes. Dye coupling between cell pairs was reduced by varying degrees for all six mutants. In homotypic oocyte pairings, only A197S induced measurable conductance. In heterotypic pairings with wild-type Cx26, five of the six mutants formed functional gap junction channels, albeit with reduced efficiency. None of the mutants displayed significant alterations in sensitivity to transjunctional voltage or induced conductive hemichannels in single oocytes. Intra-hemichannel interactions between mutant and wild-type proteins were assessed in rescue experiments using baculovirus expression in Sf9 insect cells. Of the four unstable mutations (C202F, I203T, L205V, N206S only C202F and N206S formed stable hemichannels when co-expressed with wild-type Cx26. Stable M195T hemichannels

  15. Shaping intercellular channels of plasmodesmata: the structure-to-function missing link.

    Science.gov (United States)

    Nicolas, William J; Grison, Magali S; Bayer, Emmanuelle M

    2017-12-18

    Plasmodesmata (PD) are a hallmark of the plant kingdom and a cornerstone of plant biology and physiology, forming the conduits for the cell-to-cell transfer of proteins, RNA and various metabolites, including hormones. They connect the cytosols and endomembranes of cells, which allows enhanced cell-to-cell communication and synchronization. Because of their unique position as intercellular gateways, they are at the frontline of plant defence and signalling and constitute the battleground for virus replication and spreading. The membranous organization of PD is remarkable, where a tightly furled strand of endoplasmic reticulum comes into close apposition with the plasma membrane, the two connected by spoke-like elements. The role of these structural features is, to date, still not completely understood. Recent data on PD seem to point in an unexpected direction, establishing a close parallel between PD and membrane contact sites and defining plasmodesmal membranes as microdomains. However, the implications of this new viewpoint are not fully understood. Aided by available phylogenetic data, this review attempts to reassess the function of the different elements comprising the PD and the relevance of membrane lipid composition and biophysics in defining specialized microdomains of PD, critical for their function. © The Author(s) 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  16. Intercellular Resistance to BRAF Inhibition Can Be Mediated by Extracellular Vesicle–Associated PDGFRβ

    Directory of Open Access Journals (Sweden)

    Laura J. Vella

    2017-11-01

    Full Text Available Treatment of BRAF mutant melanoma with kinase inhibitors has been associated with rapid tumor regression; however, this clinical benefit is short-lived, and most patients relapse. A number of studies suggest that the extracellular environment promotes BRAF inhibitor resistance and tumor progression. Extracellular vesicles, such as exosomes, are functional mediators in the extracellular environment. They are small vesicles known to carry a concentrated group of functional cargo and serve as intercellular communicators not only locally but also systemically. Increasingly, it is reported that extracellular vesicles facilitate the development of drug resistance in cancer; however, their role in BRAF inhibitor resistance in melanoma is unclear. Here we investigated if extracellular vesicles from BRAF inhibitor–resistant melanoma could influence drug sensitivity in recipient melanoma cells. We demonstrate that the resistance driver, PDGFRβ, can be transferred to recipient melanoma cells via extracellular vesicles, resulting in a dose-dependent activation of PI3K/AKT signaling and escape from MAPK pathway BRAF inhibition. These data suggest that the BRAF inhibitor–sensitive phenotype of metastatic melanoma can be altered by delivery of PDGFRβ by extracellular vesicles derived from neighboring drug-resistant melanoma cells.

  17. Intercellular and intracellular signaling pathways mediating ionizing radiation-induced bystander effects

    International Nuclear Information System (INIS)

    Hamada, Nobuyuki; Hara, Takamitsu; Kobayashi, Yasuhiko; Matsumoto, Hideki

    2007-01-01

    A rapidly growing body of experimental evidence indicates that ionizing radiation induces biological effects in non-irradiated bystander cells that have received signals from adjacent or distant irradiated cells. This phenomenon, which has been termed the ionizing radiation-induced bystander effect, challenges the long-standing paradigm that radiation traversal through the nucleus of a cell is a prerequisite to elicit genetic damage or a biological response. Bystander effects have been observed in a number of experimental systems, and cells whose nucleus or cytoplasm is irradiated exert bystander responses. Bystander cells manifest a multitude of biological consequences, such as genetic and epigenetic changes, alterations in gene expression, activation of signal transduction pathways, and delayed effects in their progeny. Several mediating mechanisms have been proposed. These involve gap junction-mediated intercellular communication, secreted soluble factors, oxidative metabolism, plasma membrane-bound lipid rafts, and calcium fluxes. This paper reviews briefly the current knowledge of the bystander effect with a focus on proposed mechanisms. The potential benefit of bystander effects to cancer radiotherapy will also be discussed. (author)

  18. Urinary Exosomes: The Potential for Biomarker Utility, Intercellular Signaling and Therapeutics in Urological Malignancy.

    Science.gov (United States)

    Franzen, Carrie A; Blackwell, Robert H; Foreman, Kimberly E; Kuo, Paul C; Flanigan, Robert C; Gupta, Gopal N

    2016-05-01

    Exosomes are small secreted vesicles that contain proteins, mRNA and miRNA with the potential to alter signaling pathways in recipient cells. While exosome research has flourished, few publications have specifically considered the role of genitourinary cancer shed exosomes in urine, their implication in disease progression and their usefulness as noninvasive biomarkers. In this review we examined the current literature on the role of exosomes in intercellular communication and as biomarkers, and their potential as delivery vehicles for therapeutic applications in bladder, prostate and renal cancer. We searched PubMed® and Google® with the key words prostate cancer, bladder cancer, kidney cancer, exosomes, microvesicles and urine. Relevant articles, including original research studies and reviews, were selected based on contents. A review of this literature was generated. Cancer exosomes can be isolated from urine using various techniques. Cancer cells have been found to secrete more exosomes than normal cells. These exosomes have a role in cellular communication by interacting with and depositing their cargo in target cells. Bladder, prostate and renal cancer exosomes have been shown to enhance migration, invasion and angiogenesis. These exosomes have also been shown to increase proliferation, confer drug resistance and promote immune evasion. Urinary exosomes can be isolated from bladder, kidney and prostate cancer. They serve as a potential reservoir for biomarker identification. Exosomes also have potential for therapeutics as siRNA or pharmacological agents can be loaded into exosomes. Copyright © 2016 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

  19. PDBTM: Protein Data Bank of transmembrane proteins after 8 years.

    Science.gov (United States)

    Kozma, Dániel; Simon, István; Tusnády, Gábor E

    2013-01-01

    The PDBTM database (available at http://pdbtm.enzim.hu), the first comprehensive and up-to-date transmembrane protein selection of the Protein Data Bank, was launched in 2004. The database was created and has been continuously updated by the TMDET algorithm that is able to distinguish between transmembrane and non-transmembrane proteins using their 3D atomic coordinates only. The TMDET algorithm can locate the spatial positions of transmembrane proteins in lipid bilayer as well. During the last 8 years not only the size of the PDBTM database has been steadily growing from ∼400 to 1700 entries but also new structural elements have been identified, in addition to the well-known α-helical bundle and β-barrel structures. Numerous 'exotic' transmembrane protein structures have been solved since the first release, which has made it necessary to define these new structural elements, such as membrane loops or interfacial helices in the database. This article reports the new features of the PDBTM database that have been added since its first release, and our current efforts to keep the database up-to-date and easy to use so that it may continue to serve as a fundamental resource for the scientific community.

  20. Topology of transmembrane channel-like gene 1 protein.

    Science.gov (United States)

    Labay, Valentina; Weichert, Rachel M; Makishima, Tomoko; Griffith, Andrew J

    2010-10-05

    Mutations of transmembrane channel-like gene 1 (TMC1) cause hearing loss in humans and mice. TMC1 is the founding member of a family of genes encoding proteins of unknown function that are predicted to contain multiple transmembrane domains. The goal of our study was to define the topology of mouse TMC1 expressed heterologously in tissue culture cells. TMC1 was retained in the endoplasmic reticulum (ER) membrane of five tissue culture cell lines that we tested. We used anti-TMC1 and anti-HA antibodies to probe the topologic orientation of three native epitopes and seven HA epitope tags along full-length TMC1 after selective or complete permeabilization of transfected cells with digitonin or Triton X-100, respectively. TMC1 was present within the ER as an integral membrane protein containing six transmembrane domains and cytosolic N- and C-termini. There is a large cytoplasmic loop, between the fourth and fifth transmembrane domains, with two highly conserved hydrophobic regions that might associate with or penetrate, but do not span, the plasma membrane. Our study is the first to demonstrate that TMC1 is a transmembrane protein. The topologic organization revealed by this study shares some features with that of the shaker-TRP superfamily of ion channels.

  1. The predominant mechanism of intercellular calcium wave propagation changes during long-term culture of human osteoblast-like cells

    DEFF Research Database (Denmark)

    Henriksen, Zanne; Hiken, Jeffrey F; Steinberg, Thomas H

    2006-01-01

    cells still responded to addition of ATP, but P2Y desensitization did not inhibit ICW propagation. Our data indicate that the relative role of P2Y-mediated and gap junction-mediated ICW changes during osteoblast differentiation in vitro. In less differentiated cells, P2Y-mediated ICW predominate......Intercellular calcium waves (ICW) are calcium transients that spread from cell to cell in response to different stimuli. We previously demonstrated that human osteoblast-like cells in culture propagate ICW in response to mechanical stimulation by two mechanisms. One mechanism involves autocrine...... activation of P2Y receptors, and the other requires gap junctional communication. In the current work we ask whether long-term culture of osteoblast-like cells affects the propagation of ICW by these two mechanisms. Human osteoblast-like cells were isolated from bone marrow. Mechanically induced ICW were...

  2. Catalytic combustion of propane in a membrane reactor with separate feed of reactants—II. Operation in presence of trans-membrane pressure gradients

    NARCIS (Netherlands)

    Saracco, Guido; Veldsink, Jan Willem; Versteeg, Geert F.; Swaaij, Wim P.M. van

    1995-01-01

    This is the second communication of a series dealing with an experimental and modelling study on propane catalytic combustion in a membrane reactor with separate feed of reactants. In paper I the behaviour of the reactor in the absence of trans-membrane pressure gradients was presented and

  3. Regulation of Exocytotic Fusion Pores by SNARE Protein Transmembrane Domains

    Directory of Open Access Journals (Sweden)

    Zhenyong Wu

    2017-10-01

    Full Text Available Calcium-triggered exocytotic release of neurotransmitters and hormones from neurons and neuroendocrine cells underlies neuronal communication, motor activity and endocrine functions. The core of the neuronal exocytotic machinery is composed of soluble N-ethyl maleimide sensitive factor attachment protein receptors (SNAREs. Formation of complexes between vesicle-attached v- and plasma-membrane anchored t-SNAREs in a highly regulated fashion brings the membranes into close apposition. Small, soluble proteins called Complexins (Cpx and calcium-sensing Synaptotagmins cooperate to block fusion at low resting calcium concentrations, but trigger release upon calcium increase. A growing body of evidence suggests that the transmembrane domains (TMDs of SNARE proteins play important roles in regulating the processes of fusion and release, but the mechanisms involved are only starting to be uncovered. Here we review recent evidence that SNARE TMDs exert influence by regulating the dynamics of the fusion pore, the initial aqueous connection between the vesicular lumen and the extracellular space. Even after the fusion pore is established, hormone release by neuroendocrine cells is tightly controlled, and the same may be true of neurotransmitter release by neurons. The dynamics of the fusion pore can regulate the kinetics of cargo release and the net amount released, and can determine the mode of vesicle recycling. Manipulations of SNARE TMDs were found to affect fusion pore properties profoundly, both during exocytosis and in biochemical reconstitutions. To explain these effects, TMD flexibility, and interactions among TMDs or between TMDs and lipids have been invoked. Exocytosis has provided the best setting in which to unravel the underlying mechanisms, being unique among membrane fusion reactions in that single fusion pores can be probed using high-resolution methods. An important role will likely be played by methods that can probe single fusion pores

  4. TMFoldWeb: a web server for predicting transmembrane protein fold class.

    Science.gov (United States)

    Kozma, Dániel; Tusnády, Gábor E

    2015-09-17

    Here we present TMFoldWeb, the web server implementation of TMFoldRec, a transmembrane protein fold recognition algorithm. TMFoldRec uses statistical potentials and utilizes topology filtering and a gapless threading algorithm. It ranks template structures and selects the most likely candidates and estimates the reliability of the obtained lowest energy model. The statistical potential was developed in a maximum likelihood framework on a representative set of the PDBTM database. According to the benchmark test the performance of TMFoldRec is about 77 % in correctly predicting fold class for a given transmembrane protein sequence. An intuitive web interface has been developed for the recently published TMFoldRec algorithm. The query sequence goes through a pipeline of topology prediction and a systematic sequence to structure alignment (threading). Resulting templates are ordered by energy and reliability values and are colored according to their significance level. Besides the graphical interface, a programmatic access is available as well, via a direct interface for developers or for submitting genome-wide data sets. The TMFoldWeb web server is unique and currently the only web server that is able to predict the fold class of transmembrane proteins while assigning reliability scores for the prediction. This method is prepared for genome-wide analysis with its easy-to-use interface, informative result page and programmatic access. Considering the info-communication evolution in the last few years, the developed web server, as well as the molecule viewer, is responsive and fully compatible with the prevalent tablets and mobile devices.

  5. Obif, a Transmembrane Protein, Is Required for Bone Mineralization and Spermatogenesis in Mice.

    Directory of Open Access Journals (Sweden)

    Koji Mizuhashi

    Full Text Available Various kinds of transmembrane and secreted proteins play pivotal roles in development through cell-cell communication. We previously reported that Obif (Osteoblast induction factor, Tmem119, encoding a single transmembrane protein, is expressed in differentiating osteoblasts, and that Obif-/- mice exhibit significantly reduced bone volume in the femur. In the current study, we characterized the Obif protein and further investigated the biological phenotypes of a variety of tissues in Obif-/- mice.First, we found that O-glycosylation of the Obif protein occurs at serine residue 36 in the Obif extracellular domain. Next, we observed that Obif-/- mice exhibit bone dysplasia in association with significantly increased osteoid volume per osteoid surface (OV/OS and osteoid maturation time (Omt, and significantly decreased mineral apposition rate (MAR and bone formation rate per bone surface (BFR/BS. In addition, we observed that Obif-/- mice show a significant decrease in testis weight as well as in sperm number. By histological analysis, we found that Obif is expressed in spermatocytes and spermatids in the developing testis and that spermatogenesis is halted at the round spermatid stage in the Obif-/- testis that lacks sperm. However, the number of litters fathered by male mice was slightly reduced in Obif-/- mice compared with wild-type mice, although this was not statistically significant.Our results, taken together with previous observations, indicate that Obif is a type Ia transmembrane protein whose N-terminal region is O-glycosylated. In addition, we found that Obif is required for normal bone mineralization and late testicular differentiation in vivo. These findings suggest that Obif plays essential roles in the development of multiple tissues.

  6. Obif, a Transmembrane Protein, Is Required for Bone Mineralization and Spermatogenesis in Mice.

    Science.gov (United States)

    Mizuhashi, Koji; Chaya, Taro; Kanamoto, Takashi; Omori, Yoshihiro; Furukawa, Takahisa

    2015-01-01

    Various kinds of transmembrane and secreted proteins play pivotal roles in development through cell-cell communication. We previously reported that Obif (Osteoblast induction factor, Tmem119), encoding a single transmembrane protein, is expressed in differentiating osteoblasts, and that Obif-/- mice exhibit significantly reduced bone volume in the femur. In the current study, we characterized the Obif protein and further investigated the biological phenotypes of a variety of tissues in Obif-/- mice. First, we found that O-glycosylation of the Obif protein occurs at serine residue 36 in the Obif extracellular domain. Next, we observed that Obif-/- mice exhibit bone dysplasia in association with significantly increased osteoid volume per osteoid surface (OV/OS) and osteoid maturation time (Omt), and significantly decreased mineral apposition rate (MAR) and bone formation rate per bone surface (BFR/BS). In addition, we observed that Obif-/- mice show a significant decrease in testis weight as well as in sperm number. By histological analysis, we found that Obif is expressed in spermatocytes and spermatids in the developing testis and that spermatogenesis is halted at the round spermatid stage in the Obif-/- testis that lacks sperm. However, the number of litters fathered by male mice was slightly reduced in Obif-/- mice compared with wild-type mice, although this was not statistically significant. Our results, taken together with previous observations, indicate that Obif is a type Ia transmembrane protein whose N-terminal region is O-glycosylated. In addition, we found that Obif is required for normal bone mineralization and late testicular differentiation in vivo. These findings suggest that Obif plays essential roles in the development of multiple tissues.

  7. Modelling of a transmembrane evaporation module for desalination of seawater

    NARCIS (Netherlands)

    Guijt, C.M.; Racz, I.G.; van Heuven, Jan Willem; Reith, T.; de Haan, A.B.

    1999-01-01

    Transmembrane evaporation (often called membrane distillation) carried out in a countercurrent flow module, in which incoming cold seawater is heated by the condensing product water flow, is a promising technology for low-cost seawater desalination. This paper presents a model for preliminary design

  8. Regulation of germ cell development by intercellular signaling in the mammalian ovarian follicle.

    Science.gov (United States)

    Clarke, Hugh J

    2018-01-01

    Prior to ovulation, the mammalian oocyte undergoes a process of differentiation within the ovarian follicle that confers on it the ability to give rise to an embryo. Differentiation comprises two phases-growth, during which the oocyte increases more than 100-fold in volume as it accumulates macromolecules and organelles that will sustain early embryogenesis; and meiotic maturation, during which the oocyte executes the first meiotic division and prepares for the second division. Entry of an oocyte into the growth phase appears to be triggered when the adjacent granulosa cells produce specific growth factors. As the oocyte grows, it elaborates a thick extracellular coat termed the zona pellucida. Nonetheless, cytoplasmic extensions of the adjacent granulosa cells, termed transzonal projections (TZPs), enable them to maintain contact-dependent communication with the oocyte. Through gap junctions located where the TZP tips meet the oocyte membrane, they provide the oocyte with products that sustain its metabolic activity and signals that regulate its differentiation. Conversely, the oocyte secretes diffusible growth factors that regulate proliferation and differentiation of the granulosa cells. Gap junction-permeable products of the granulosa cells prevent precocious initiation of meiotic maturation, and the gap junctions also enable oocyte maturation to begin in response to hormonal signals received by the granulosa cells. Development of the oocyte or the somatic compartment may also be regulated by extracellular vesicles newly identified in follicular fluid and at TZP tips, which could mediate intercellular transfer of macromolecules. Oocyte differentiation thus depends on continuous signaling interactions with the somatic cells of the follicle. WIREs Dev Biol 2018, 7:e294. doi: 10.1002/wdev.294 This article is categorized under: Gene Expression and Transcriptional Hierarchies > Cellular Differentiation Signaling Pathways > Cell Fate Signaling Early Embryonic

  9. Assessing the utility of photoswitchable fluorescent proteins for tracking intercellular protein movement in the Arabidopsis root.

    Directory of Open Access Journals (Sweden)

    Shuang Wu

    Full Text Available One way in which cells communicate is through the direct transfer of proteins. In plants, many of these proteins are transcription factors, which are made by one cell type and traffic into another. In order to understand how this movement occurs and its role in development, we would like to track this movement in live, intact plants in real-time. Here we examine the utility of the photoconvertible proteins, Dendra2 and (to a lesser extent EosFP as tags for studying intracellular and intercellular protein movement in the Arabidopsis root. To this end, we made fusions between Dendra2 and six mobile transcription factors. Our results show that Dendra2 is an effective tool for studying protein movement between plant cells. Interestingly, we found that Dendra2 could not simply be swapped into existing constructs that had originally contained GFP. Most of the fusions made in this way failed to produce a fluorescent fusion. In addition we found that the optimal settings for photoconversion of Dendra2 in stably transformed roots were different from what has been published for photoconversion in transient assays in plants or in animal cells. By modifying the confocal setting, we were able to photoconvert Dendra2 in all cell layers in the root. However the efficiency of photoconversion was affected by the position of the cell layer within the root, with more internal tissues requiring more energy. By examining the Dendra2 fusions, we confirmed the mobility of the SHORT-ROOT (SHR and CAPRICE (CPC transcription factors between cells and we further discovered that SHR movement in stele and CPC movement in the epidermis are non-directional.

  10. Low dose gamma irradiation enhances defined signaling components of intercellular reactive oxygen-mediated apoptosis induction

    International Nuclear Information System (INIS)

    Bauer, G

    2011-01-01

    Transformed cells are selectively removed by intercellular ROS-mediated induction of apoptosis. Signaling is based on the HOCl and the NO/peroxynitrite pathway (major pathways) and the nitryl chloride and the metal-catalyzed Haber-Weiss pathway (minor pathways). During tumor progression, resistance against intercellular induction of apoptosis is acquired through expression of membrane-associated catalase. Low dose radiation of nontransformed cells has been shown to enhance intercellular induction of apoptosis. The present study was performed to define the signaling components which are modulated by low dose gamma irradiation. Low dose radiation induced the release of peroxidase from nontransformed, transformed and tumor cells. Extracellular superoxide anion generation was strongly enhanced in the case of transformed cells and tumor cells, but not in nontransformed cells. Enhancement of peroxidase release and superoxide anion generation either increased intercellular induction of apoptosis of transformed cells, or caused a partial protection under specific signaling conditions. In tumor cells, low dose radiation enhanced the production of major signaling components, but this had no effect on apoptosis induction, due to the strong resistance mechanism of tumor cells. Our data specify the nature of low dose radiation-induced effects on specific signaling components of intercellular induction of apoptosis at defined stages of multistep carcinogenesis.

  11. Intercellular Communication of Tumor Cells and Immune Cells after Exposure to Different Ionizing Radiation Qualities

    OpenAIRE

    Diegeler, Sebastian; Hellweg, Christine E.

    2017-01-01

    Ionizing radiation can affect the immune system in many ways. Depending on the situation, the whole body or parts of the body can be acutely or chronically exposed to different radiation qualities. In tumor radiotherapy, a fractionated exposure of the tumor (and surrounding tissues) is applied to kill the tumor cells. Currently, mostly photons, and also electrons, neutrons, protons, and heavier particles such as carbon ions, are used in radiotherapy. Tumor elimination can be supported by an e...

  12. Intercellular Communication of Tumor Cells and Immune Cells after Exposure to Different Ionizing Radiation Qualities

    Directory of Open Access Journals (Sweden)

    Sebastian Diegeler

    2017-06-01

    Full Text Available Ionizing radiation can affect the immune system in many ways. Depending on the situation, the whole body or parts of the body can be acutely or chronically exposed to different radiation qualities. In tumor radiotherapy, a fractionated exposure of the tumor (and surrounding tissues is applied to kill the tumor cells. Currently, mostly photons, and also electrons, neutrons, protons, and heavier particles such as carbon ions, are used in radiotherapy. Tumor elimination can be supported by an effective immune response. In recent years, much progress has been achieved in the understanding of basic interactions between the irradiated tumor and the immune system. Here, direct and indirect effects of radiation on immune cells have to be considered. Lymphocytes for example are known to be highly radiosensitive. One important factor in indirect interactions is the radiation-induced bystander effect which can be initiated in unexposed cells by expression of cytokines of the irradiated cells and by direct exchange of molecules via gap junctions. In this review, we summarize the current knowledge about the indirect effects observed after exposure to different radiation qualities. The different immune cell populations important for the tumor immune response are natural killer cells, dendritic cells, and CD8+ cytotoxic T-cells. In vitro and in vivo studies have revealed the modulation of their functions due to ionizing radiation exposure of tumor cells. After radiation exposure, cytokines are produced by exposed tumor and immune cells and a modulated expression profile has also been observed in bystander immune cells. Release of damage-associated molecular patterns by irradiated tumor cells is another factor in immune activation. In conclusion, both immune-activating and -suppressing effects can occur. Enhancing or inhibiting these effects, respectively, could contribute to modified tumor cell killing after radiotherapy.

  13. Intercellular Communication of Tumor Cells and Immune Cells after Exposure to Different Ionizing Radiation Qualities.

    Science.gov (United States)

    Diegeler, Sebastian; Hellweg, Christine E

    2017-01-01

    Ionizing radiation can affect the immune system in many ways. Depending on the situation, the whole body or parts of the body can be acutely or chronically exposed to different radiation qualities. In tumor radiotherapy, a fractionated exposure of the tumor (and surrounding tissues) is applied to kill the tumor cells. Currently, mostly photons, and also electrons, neutrons, protons, and heavier particles such as carbon ions, are used in radiotherapy. Tumor elimination can be supported by an effective immune response. In recent years, much progress has been achieved in the understanding of basic interactions between the irradiated tumor and the immune system. Here, direct and indirect effects of radiation on immune cells have to be considered. Lymphocytes for example are known to be highly radiosensitive. One important factor in indirect interactions is the radiation-induced bystander effect which can be initiated in unexposed cells by expression of cytokines of the irradiated cells and by direct exchange of molecules via gap junctions. In this review, we summarize the current knowledge about the indirect effects observed after exposure to different radiation qualities. The different immune cell populations important for the tumor immune response are natural killer cells, dendritic cells, and CD8+ cytotoxic T-cells. In vitro and in vivo studies have revealed the modulation of their functions due to ionizing radiation exposure of tumor cells. After radiation exposure, cytokines are produced by exposed tumor and immune cells and a modulated expression profile has also been observed in bystander immune cells. Release of damage-associated molecular patterns by irradiated tumor cells is another factor in immune activation. In conclusion, both immune-activating and -suppressing effects can occur. Enhancing or inhibiting these effects, respectively, could contribute to modified tumor cell killing after radiotherapy.

  14. Xenobiotic Modulation of Human Mammary Epithelial Cell Gap junctional Intercellular Communication and Growth

    National Research Council Canada - National Science Library

    Ruch, Randall

    1998-01-01

    ...), phthalate esters, and dioxin have been implicated in this increase. Many xenobiotics such as DDT and PCBs have weak estrogenic activity and may enhance breast cancer formation by an estrogenic effect on breast epithelial cell growth...

  15. Xenobiotic Modulation of Human Mammary Epithelial Cell Gap Junctional Intercellular Communication and Growth

    National Research Council Canada - National Science Library

    Ruch, Randall

    1997-01-01

    ...), phthalate esters, and dioxin have been implicated in this increase. Many xenobiotics such as DDT and PCBs have weak estrogenic activity and may enhance breast cancer formation by an estrogenic effect on breast epithelial cell growth...

  16. HIV-1 intersection with CD4 T cell vesicle exocytosis: intercellular communication goes viral

    Directory of Open Access Journals (Sweden)

    Helena eSoares

    2014-09-01

    Full Text Available In cells of the immune system the secretion of extracellular vesicles is modulated through cellular activation. In particular, T cell activation is achieved through cell-cell contacts with antigen presenting cells and the consequent formation of a specialized signaling junction called the immunological synapse. Recent works on CD4 T cells have elucidated that cognate antigen recognition by the T cell receptor (TCR engages two distinct exocytic events. The first, involves the exocytic targeting of signaling molecules at the synaptic membrane and drives the functional architecture of the immunological synapse. The second, enlists the extracellular secretion of the TCR itself, once the functional architecture of the immunological synapse is accomplished. HIV-1, a human lymphotropic virus, has evolved sophisticated mechanisms to co-opt CD4 T cell physiology. Notably, it has become apparent that HIV-1 intersects the regulated secretory system of CD4 T cells in order to bud from the plasma membrane of the infected cell and to promote bystander cell death. Here, I review the relevance of CD4 vesicle exocytosis to immune regulation and to HIV-1 pathogenesis and discuss their potential therapeutic applications.

  17. On the origin of microparticles: From “platelet dust” to mediators of intercellular communication

    Science.gov (United States)

    Hargett, Leslie A.; Bauer, Natalie N.

    2013-01-01

    Microparticles are submicron vesicles shed from a variety of cells. Peter Wolf first identified microparticles in the midst of ongoing blood coagulation research in 1967 as a product of platelets. He termed them platelet dust. Although initially thought to be useless cellular trash, decades of research focused on the tiny vesicles have defined their roles as participators in coagulation, cellular signaling, vascular injury, and homeostasis. The purpose of this review is to highlight the science leading up to the discovery of microparticles, feature discoveries made by key contributors to the field of microparticle research, and discuss their positive and negative impact on the pulmonary circulation. PMID:24015332

  18. Glycine Perturbs Local and Global Conformational Flexibility of a Transmembrane Helix

    DEFF Research Database (Denmark)

    Högel, Philipp; Götz, Alexander; Kuhne, Felix

    2018-01-01

    Flexible transmembrane helices frequently support the conformational transitions between different functional states of membrane proteins. While proline is well known to distort and destabilize transmembrane helices, the role of glycine is still debated. Here, we systematically investigated the e...

  19. Cavitation of intercellular spaces is critical to establishment of hydraulic properties of compression wood of Chamaecyparis obtusa seedlings.

    Science.gov (United States)

    Nakaba, Satoshi; Hirai, Asami; Kudo, Kayo; Yamagishi, Yusuke; Yamane, Kenichi; Kuroda, Katsushi; Nugroho, Widyanto Dwi; Kitin, Peter; Funada, Ryo

    2016-03-01

    When the orientation of the stems of conifers departs from the vertical as a result of environmental influences, conifers form compression wood that results in restoration of verticality. It is well known that intercellular spaces are formed between tracheids in compression wood, but the function of these spaces remains to be clarified. In the present study, we evaluated the impact of these spaces in artificially induced compression wood in Chamaecyparis obtusa seedlings. We monitored the presence or absence of liquid in the intercellular spaces of differentiating xylem by cryo-scanning electron microscopy. In addition, we analysed the relationship between intercellular spaces and the hydraulic properties of the compression wood. Initially, we detected small intercellular spaces with liquid in regions in which the profiles of tracheids were not rounded in transverse surfaces, indicating that the intercellular spaces had originally contained no gases. In the regions where tracheids had formed secondary walls, we found that some intercellular spaces had lost their liquid. Cavitation of intercellular spaces would affect hydraulic conductivity as a consequence of the induction of cavitation in neighbouring tracheids. Our observations suggest that cavitation of intercellular spaces is the critical event that affects not only the functions of intercellular spaces but also the hydraulic properties of compression wood. © The Author 2016. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  20. Effect of sound on gap-junction-based intercellular signaling: Calcium waves under acoustic irradiation.

    Science.gov (United States)

    Deymier, P A; Swinteck, N; Runge, K; Deymier-Black, A; Hoying, J B

    2015-01-01

    We present a previously unrecognized effect of sound waves on gap-junction-based intercellular signaling such as in biological tissues composed of endothelial cells. We suggest that sound irradiation may, through temporal and spatial modulation of cell-to-cell conductance, create intercellular calcium waves with unidirectional signal propagation associated with nonconventional topologies. Nonreciprocity in calcium wave propagation induced by sound wave irradiation is demonstrated in the case of a linear and a nonlinear reaction-diffusion model. This demonstration should be applicable to other types of gap-junction-based intercellular signals, and it is thought that it should be of help in interpreting a broad range of biological phenomena associated with the beneficial therapeutic effects of sound irradiation and possibly the harmful effects of sound waves on health.

  1. In vivo relevance of intercellular calcium signaling in Drosophila wing development

    OpenAIRE

    Brodskiy, Pavel; Brito-Robinson, Teresa; Levis, Megan; Narciso, Cody; Jangula, Jamison; Huizar, Francisco; Wu, Qinfeng; Zartman, Jeremiah

    2017-01-01

    Recently, organ-scale intercellular Ca2+ transients (ICTs) were reported in the Drosophila wing disc. However, the functional in vivo significance of ICTs remains largely unknown. Here we demonstrate the in vivo relevance of intercellular Ca2+ signaling and its impact on wing development. We report that Ca2+ signaling in vivo decreases as wing discs mature. Ca2+ signaling ex vivo responds to fly extract in a dose-dependent manner. This suggests ICTs occur in vivo due to chemical stimulus that...

  2. A percolation-like model for simulating inter-cellular diffusion in the context of bystander signalling in tumour

    International Nuclear Information System (INIS)

    Moulton, C.R.; Fleming, A.J.; Ebert, M.A.

    2011-01-01

    Full text: Despite ongoing active research, the role of the radiation bystander effect in modifying local tissue response to an ionising radiation dose remains unclear. The present study aims to provide new insight by simulating the diffusion-mediated inter-cellular communication processes in 2D and 3D cell-like structures to calculate likely signal ranges in the diffusion limited case. Random walks of individual signalling molecules were tracked between cells with inclusion of molecule-receptor interactions. The resulting diffusion anomaly is a function of cell density, signal uptake probability and the spatial arrangement of cells local to the signal origin. Uptake probability effects dominate percolation effects in disordered media. Diffu sion through 2D structures is more conducive to anomalous diffusion than diffusion through 3D structures. Values for time-dependent diffusion constants and permeability are derived for typical simulation parameters. Even at low signal uptake probabilities the communication range is restricted to a mean value of less than 100 foun owing to complete signal uptake by 600 s. This should be considered in light of the potential influence of signal relaying, flo dynamics or vasculature-mediated signalling.

  3. Cell-to-cell communication in plants, animals, and fungi: a comparative review.

    Science.gov (United States)

    Bloemendal, Sandra; Kück, Ulrich

    2013-01-01

    Cell-to-cell communication is a prerequisite for differentiation and development in multicellular organisms. This communication has to be tightly regulated to ensure that cellular components such as organelles, macromolecules, hormones, or viruses leave the cell in a precisely organized way. During evolution, plants, animals, and fungi have developed similar ways of responding to this biological challenge. For example, in higher plants, plasmodesmata connect adjacent cells and allow communication to regulate differentiation and development. In animals, two main general structures that enable short- and long-range intercellular communication are known, namely gap junctions and tunneling nanotubes, respectively. Finally, filamentous fungi have also developed specialized structures called septal pores that allow intercellular communication via cytoplasmic flow. This review summarizes the underlying mechanisms for intercellular communication in these three eukaryotic groups and discusses its consequences for the regulation of differentiation and developmental processes.

  4. The association between soluble intercellular adhesion molecule-1 levels in drained dialysate and peritoneal injury in peritoneal dialysis.

    Science.gov (United States)

    Igarashi, Yusuke; Morishita, Yoshiyuki; Yoshizawa, Hiromichi; Imai, Reika; Imai, Toshimi; Hirahara, Ichiro; Akimoto, Tetsu; Ookawara, Susumu; Ishibashi, Kenichi; Muto, Shigeaki; Nagata, Daisuke

    2017-11-01

    Chronic inflammation of the peritoneum causes peritoneal injury in patients on peritoneal dialysis. Intercellular adhesion molecule-1 and its circulating form, soluble intercellular adhesion molecule-1, play pivotal roles in inflammation. However, their role in peritoneal injury is unclear. We measured changes in intercellular adhesion molecule-1 expression in the peritoneum of a peritoneal injury model in rats. The associations between soluble intercellular adhesion molecule-1 levels in drained dialysate and the solute transport rate (D/P-Cr and D/D0-glucose) determined by the peritoneal equilibration test, and matrix metalloproteinase-2 levels in drained dialysate were investigated in 94 peritoneal drained dialysate samples. Intercellular adhesion molecule-1 expression was increased in the peritoneum of rats with peritoneal injury. Soluble intercellular adhesion molecule-1 levels in drained dialysate were significantly positively correlated with D/P-Cr (r = .51, p molecule-1expression is increased in the peritoneum of a peritoneal injury model in the rat, and soluble intercellular adhesion molecule-1 levels in drained dialysate are associated with peritoneal injury in patients on peritoneal dialysis. These results suggest that soluble intercellular adhesion molecule-1 could be a novel biomarker of peritoneal injury in patients on peritoneal dialysis.

  5. The effect of water saturation deficit on the volume of intercellular space in laeves

    Directory of Open Access Journals (Sweden)

    J. Czerski

    2015-01-01

    Full Text Available The volume of intercellular spaces in leaves at various stages of water saturation was determined by method of Czerski (1964, 1968. The investigation were performed with the following plant species: Vicia faba L., Nicotiana tabacum L. var. rustica, Solarium tuberosum L. var. Flisak, Helichrysum bracteatum Wild., Bmssica napus L. var. oleifera, Beta vulgaris L. var. saccharifera.

  6. A transmembrane polar interaction is involved in the functional regulation of integrin alpha L beta 2.

    Science.gov (United States)

    Vararattanavech, Ardcharaporn; Chng, Choon-Peng; Parthasarathy, Krupakar; Tang, Xiao-Yan; Torres, Jaume; Tan, Suet-Mien

    2010-05-14

    Integrins are heterodimeric transmembrane (TM) receptors formed by noncovalent associations of alpha and beta subunits. Each subunit contains a single alpha-helical TM domain. Inside-out activation of an integrin involves the separation of its cytoplasmic tails, leading to disruption of alphabeta TM packing. The leukocyte integrin alpha L beta 2 is required for leukocyte adhesion, migration, proliferation, cytotoxic function, and antigen presentation. In this study, we show by mutagenesis experiments that the packing of alpha L beta 2 TMs is consistent with that of the integrin alpha IIb beta 3 TMs. However, molecular dynamics simulations of alpha L beta 2 TMs in lipids predicted a polar interaction involving the side chains of alpha L Ser1071 and beta2 Thr686 in the outer-membrane association clasp (OMC). This is supported by carbonyl vibrational shifts observed in isotope-labeled alpha L beta 2 TM peptides that were incorporated into lipid bilayers. Molecular dynamics studies simulating the separation of alpha L beta 2 tails showed the presence of polar interaction during the initial perturbation of the inner-membrane association clasp. When the TMs underwent further separation, the polar interaction was disrupted. OMC polar interaction is important in regulating the functions of beta2 integrins because mutations that disrupt the OMC polar interaction generated constitutively activated alpha L beta 2, alpha M beta 2, and alpha X beta 2 in 293T transfectants. We also show that the expression of mutant beta2 Thr686Gly in beta2-deficient T cells rescued cell adhesion to intercellular adhesion molecule 1, but the cells showed overt elongated morphologies in response to chemokine stromal-cell-derived factor 1 alpha treatment as compared to wild-type beta2-expressing cells. These two TM polar residues are totally conserved in other members of the beta2 integrins in humans and across different species. Our results provide an example of the stabilizing effect of polar

  7. Intercellular Adhesion Molecule-1 Levels in Experimental Brain Injury and the Effects of Alpha-tocopherol

    Directory of Open Access Journals (Sweden)

    Nilgun Senol

    2014-06-01

    Full Text Available Aim: The mechanisms, responsible for the secondary injuries occuring after acute injury of the brain are; release of nitrous oxide which is an inflammatory mediator, abnormal formation of free oxygen radicals and excessive stimulation of excitatory aminoacids. In this study, it is aimed to investigate changes in intercellular adhesion molecule levels in the brain, that occur subsequent to blunt head trauma, and after administration of an antioxidant agent, vitamin E. Material and Method: In this study, rats were divided into 4 groups. In group A; rats had only skin incision, group B; rats were traumatized after the skin incision, group C; isotonic (30mg/kg was given intraperitoneally after 30 minutes of the trauma, group D; alpha-tocopherol (30mg/kg was given intraperitoneally, after 30 minutes of the trauma. All the rats in these groups were sacrified after 24 hours. Biparietal and bifrontal lobs were taken about 3x5x1mm tickness and intercellular adhesion molecule-1 levels were studied by enzyme-linked immunosorbent assay kit. Results: As the result of the statistical analysis, it is detected that although there is an increase in intercellular adhesion molecule levels in brain parenchyma after trauma, it is statistically unsignificant. However, as the traumatized group and the group given alpha-tocopherol after trauma was compared, a statistically significant decrease in intercellular adhesion molecule-1 levels in the alpha-tocopherol given group was seen. Discussion: Alpha-tocopherol, an antioxidant agent, causes decrease in intercellular adhesion molecule levels, by decreasing inflammation.

  8. Plasmodesmata-mediated intercellular signaling during plant growth and development

    Directory of Open Access Journals (Sweden)

    Shri Ram eYadav

    2014-02-01

    Full Text Available Plasmodesmata (PD are cytoplasmic channels that connect neighboring cells for cell-to-cell communication. PD structure and function vary temporally and spatially to allow formation of symplastic domains during different stages of plant development. Reversible deposition of callose at PD plays an important role in controlling molecular trafficking through PD by regulating their size exclusion limit (SEL. Previously, we reported several semi-dominant mutants for CALLOSE SYNTHASE 3 (CALS3 gene, which overproduce callose at PD in Arabidopsis. By combining two of these mutations in a LexA-VP16-ER (XVE-based estradiol inducible vector system, a tool known as the icals3m system was developed to temporally obstruct the symplastic connections in a specified spatial domain. The system has been successfully tested and used, in combination with other methods, to investigate the route for mobile signals such as the SHR protein, microRNA165/6, and cytokinins in Arabidopsis roots, and also to understand the role of symplastic domain formation during lateral root development. We envision that this tool may also be useful for identifying tissue-specific symplastic regulatory networks and to analyze symplastic movement of metabolites.

  9. Plasmodesmata-mediated intercellular signaling during plant growth and development.

    Science.gov (United States)

    Yadav, Shri R; Yan, Dawei; Sevilem, Iris; Helariutta, Ykä

    2014-01-01

    Plasmodesmata (PD) are cytoplasmic channels that connect neighboring cells for cell-to-cell communication. PD structure and function vary temporally and spatially to allow formation of symplastic domains during different stages of plant development. Reversible deposition of callose at PD plays an important role in controlling molecular trafficking through PD by regulating their size exclusion limit. Previously, we reported several semi-dominant mutants for CALLOSE SYNTHASE 3 (CALS3) gene, which overproduce callose at PD in Arabidopsis. By combining two of these mutations in a LexA-VP16-ER (XVE)-based estradiol inducible vector system, a tool known as the "icals3m system" was developed to temporally obstruct the symplastic connections in a specified spatial domain. The system has been successfully tested and used, in combination with other methods, to investigate the route for mobile signals such as the SHR protein, microRNA165/6, and cytokinins in Arabidopsis roots, and also to understand the role of symplastic domain formation during lateral root development. We envision that this tool may also be useful for identifying tissue-specific symplastic regulatory networks and to analyze symplastic movement of metabolites.

  10. Molecular Insights into the Transmembrane Domain of the Thyrotropin Receptor.

    Directory of Open Access Journals (Sweden)

    Vanessa Chantreau

    Full Text Available The thyrotropin receptor (TSHR is a G protein-coupled receptor (GPCR that is member of the leucine-rich repeat subfamily (LGR. In the absence of crystal structure, the success of rational design of ligands targeting the receptor internal cavity depends on the quality of the TSHR models built. In this subfamily, transmembrane helices (TM 2 and 5 are characterized by the absence of proline compared to most receptors, raising the question of the structural conformation of these helices. To gain insight into the structural properties of these helices, we carried out bioinformatics and experimental studies. Evolutionary analysis of the LGR family revealed a deletion in TM5 but provided no information on TM2. Wild type residues at positions 2.58, 2.59 or 2.60 in TM2 and/or at position 5.50 in TM5 were substituted to proline. Depending on the position of the proline substitution, different effects were observed on membrane expression, glycosylation, constitutive cAMP activity and responses to thyrotropin. Only proline substitution at position 2.59 maintained complex glycosylation and high membrane expression, supporting occurrence of a bulged TM2. The TSHR transmembrane domain was modeled by homology with the orexin 2 receptor, using a protocol that forced the deletion of one residue in the TM5 bulge of the template. The stability of the model was assessed by molecular dynamics simulations. TM5 straightened during the equilibration phase and was stable for the remainder of the simulations. Our data support a structural model of the TSHR transmembrane domain with a bulged TM2 and a straight TM5 that is specific of glycoprotein hormone receptors.

  11. Bystander effects: intercellular transmission of radiation damage signals

    Energy Technology Data Exchange (ETDEWEB)

    Little, J.B.; Azzam, E.I.; Toledo, S.M. de; Nagasawa, H

    2002-07-01

    Biological effects were examined in confluent cultures of fibroblasts and epithelial cells exposed to very low mean doses of alpha radiation, doses by which only 1-2% of the cells were actually traversed by an alpha particle. Enhanced frequencies of sister chromatid exchanges and HPRT mutations occurred in the non-irradiation, 'bystander' cells associated with a similar increase in the frequency of micronuclei, indicating the induction of DNA damage in these cells. In order to gain information concerning molecular pathways, changes in gene expression were examined in bystander cells by western analysis and in situ immunofluorescence staining. The expression levels of p53, p21 and MDM2 were significantly modulated in bystander cells: the damage signals leading to these changes were transmitted from irradiated to bystander cells by gap junction mediated intracellular communication. The bystander response was suppressed by incubation with superoxide dismutase as well as an inhibitor of NADPH oxidase, suggesting the effect may be mediated by oxidative stress. To examine other signalling pathways responsive to oxidative stress, the activation of stress-related kinases and their downstream transcription factors were analysed in bystander cells by western blotting and electrophoretic mobility shift assays: a 2-4 fold increase in the phosphorylation levels of JNK, EPK1/2, p90RSK, Elk-1 and ATF2 was observed. These changes were detected by 15 min after irradiation and persisted for at least 1 h. These findings indicate the activation of multiple signal transduction pathways in bystander cells, involving signals arising from the plasma membrane as well as from DNA damage. (author)

  12. Promiscuous Seven Transmembrane Receptors Sensing L-α-amino Acids

    DEFF Research Database (Denmark)

    Smajilovic, Sanela; Wellendorph, Petrine; Bräuner-Osborne, Hans

    2014-01-01

    A number of nutrient sensing seven trans-membrane (7TM) receptors have been identified and characterized over the past few years. While the sensing mechanisms to carbohydrates and free fatty acids are well understood, the molecular basis of amino acid sensing has recently come to the limelight....... The present review describes the current status of promiscuous L-α-amino acid sensors, the calcium sensing receptor (CaSR), the GPRC6A receptor, the T1R1/T1R3 receptor and also their molecular pharmacology, expression pattern and physiological significance....

  13. Role of α and β Transmembrane Domains in Integrin Clustering

    Directory of Open Access Journals (Sweden)

    Amir Shamloo

    2015-11-01

    Full Text Available Integrins are transmembrane proteins playing a crucial role in the mechanical signal transduction from the outside to the inside of a cell, and vice versa. Nevertheless, this signal transduction could not be implemented by a single protein. Rather, in order for integrins to be able to participate in signal transduction, they need to be activated and produce clusters first. As integrins consist of α- and β-subunits that are separate in the active state, studying both subunits separately is of a great importance, for, in the active state, the distance between α- and β-subunits is long enough that they do not influence one another significantly. Thus, this study aims to investigate the tendency of transmembrane domains of integrins to form homodimers. We used both Steered and MARTINI Coarse-grained molecular dynamics method to perform our simulations, mainly because of a better resolution and computational feasibility that each of these methods could provide to us. Using the Steered molecular dynamics method for α- and β-subunits, we found that the localized lipid packing prevented them from clustering. Nonetheless, the lipid packing phenomenon was found to be an artifact after investigating this process using a coarse grained (CG model. Exploiting the coarse-grained molecular dynamics simulations, we found that α- and β-subunits tend to form a stable homo-dimer.

  14. Structure and function of the cystic fibrosis transmembrane conductance regulator

    Directory of Open Access Journals (Sweden)

    M.M. Morales

    1999-08-01

    Full Text Available Cystic fibrosis (CF is a lethal autosomal recessive genetic disease caused by mutations in the CF transmembrane conductance regulator (CFTR. Mutations in the CFTR gene may result in a defective processing of its protein and alter the function and regulation of this channel. Mutations are associated with different symptoms, including pancreatic insufficiency, bile duct obstruction, infertility in males, high sweat Cl-, intestinal obstruction, nasal polyp formation, chronic sinusitis, mucus dehydration, and chronic Pseudomonas aeruginosa and Staphylococcus aureus lung infection, responsible for 90% of the mortality of CF patients. The gene responsible for the cellular defect in CF was cloned in 1989 and its protein product CFTR is activated by an increase of intracellular cAMP. The CFTR contains two membrane domains, each with six transmembrane domain segments, two nucleotide-binding domains (NBDs, and a cytoplasmic domain. In this review we discuss the studies that have correlated the role of each CFTR domain in the protein function as a chloride channel and as a regulator of the outwardly rectifying Cl- channels (ORCCs.

  15. PI3K/Akt signaling is involved in the disruption of gap junctional communication caused by v-Src and TNF-α.

    Science.gov (United States)

    Ito, Satoko; Hyodo, Toshinori; Hasegawa, Hitoki; Yuan, Hong; Hamaguchi, Michinari; Senga, Takeshi

    2010-09-17

    Gap junctional communication, which is mediated by the connexin protein family, is essential for the maintenance of normal tissue function and homeostasis. Loss of intercellular communication results in a failure to coordinately regulate cellular functions, and it can facilitate tumorigenesis. Expression of oncogenes and stimulation with cytokines has been shown to suppress intercellular communication; however, the exact mechanism by which intercellular communication is disrupted by these factors remains uncertain. In this report, we show that Akt is essential for the disruption of gap junctional communication in v-Src-transformed cells. In addition, inhibition of Akt restores gap junctional communication after it is suppressed by TNF-α signaling. Furthermore, we demonstrate that the expression of a constitutively active form of Akt1, but not of Akt2 or Akt3, is sufficient to suppress gap junctional communication. Our results clearly define Akt1 as one of the critical regulators of gap junctional communication. Copyright © 2010 Elsevier Inc. All rights reserved.

  16. Expression of a defence-related intercellular barley peroxidase in transgenic tobacco

    DEFF Research Database (Denmark)

    Kristensen, B.K.; Brandt, J.; Bojsen, K.

    1997-01-01

    genetically, phenotypically and biochemically. The T-DNA was steadily inherited through three generations. The barley peroxidase is expressed and sorted to the intercellular space in the transgenic tobacco plants. The peroxidase can be extracted from the intercellular space in two molecular forms from both...... barley and transgenic tobacco. The tobacco expressed forms are indistinguishable from the barley expressed forms as determined by analytical isoelectric focusing (pI 8.5) and Western-blotting. Staining for N-glycosylation showed that one form only was glycosylated. The N-terminus of purified Prx8 from...... transgenic tobacco was blocked by pyroglutamate, after the removal of which, N-terminal sequencing verified the transit signal-peptide cleavage site deduced from the cDNA sequence. Phenotype comparisons show that the constitutive expression of Prx8 lead to growth retardation. However, an infection assay...

  17. Proteins Play Important Role in Intercellular Adhesion Affecting on Fruit Textural Quality

    DEFF Research Database (Denmark)

    Bahadur Adhikari, Khem; Shomer, Ilan

    2012-01-01

    Fruit textural quality is becoming a major quality parameter for export, postharvest preservation, handling and processing. The main determinant of textural quality is intercellular adhesion (ICA) as attributed by the cell wall (CW) and its components. The importance of CW protein in ICA strength......Fruit textural quality is becoming a major quality parameter for export, postharvest preservation, handling and processing. The main determinant of textural quality is intercellular adhesion (ICA) as attributed by the cell wall (CW) and its components. The importance of CW protein in ICA...... strengthening was exempli ed in Medjoul date (Phoenix dactylifera L.) fruit, as a model. Fruit mesocarp sensitively responded to culture environment which was assayed in vitro at pH 3.5( pKa) in presence of organic acid molecules. The max penetration force, as a measure of ICA strength, of p...

  18. Detection of cancerous kidney tissue areas by means of infrared spectroscopy of intercellular fluid

    Science.gov (United States)

    Urboniene, V.; Jankevicius, F.; Zelvys, A.; Steiner, G.; Sablinskas, V.

    2014-03-01

    In this work the infrared absorption spectra of intercellular fluid of normal and tumor kidney tissue were recorded and analyzed. The samples were prepared by stamping freshly resected tissue onto a CaF2 substrate. FT-IR spectra obtained from intracellular fluid of tumor tissue exhibit stronger absorption bands in the spectral region from 1000-1200 cm-1 and around 1750 cm-1 than those obtained from normal tissue. It is likely the spectra of extracellular matrix of kidney tumor tissue with large increases in the intensities of these bands represent a higher concentration of fatty acids and glycerol. Amide I and amide II bands are stronger in spectra of normal tissue indicating a higher level of proteins. The results demonstrate that FT-IR spectroscopy of intercellular fluids is a novel approach for a quick diagnosis during surgical resection, which can improve the therapy of kidney tumors.

  19. Tunneling nanotubes spread fibrillar α-synuclein by intercellular trafficking of lysosomes.

    Science.gov (United States)

    Abounit, Saïda; Bousset, Luc; Loria, Frida; Zhu, Seng; de Chaumont, Fabrice; Pieri, Laura; Olivo-Marin, Jean-Christophe; Melki, Ronald; Zurzolo, Chiara

    2016-10-04

    Synucleinopathies such as Parkinson's disease are characterized by the pathological deposition of misfolded α-synuclein aggregates into inclusions throughout the central and peripheral nervous system. Mounting evidence suggests that intercellular propagation of α-synuclein aggregates may contribute to the neuropathology; however, the mechanism by which spread occurs is not fully understood. By using quantitative fluorescence microscopy with co-cultured neurons, here we show that α-synuclein fibrils efficiently transfer from donor to acceptor cells through tunneling nanotubes (TNTs) inside lysosomal vesicles. Following transfer through TNTs, α-synuclein fibrils are able to seed soluble α-synuclein aggregation in the cytosol of acceptor cells. We propose that donor cells overloaded with α-synuclein aggregates in lysosomes dispose of this material by hijacking TNT-mediated intercellular trafficking. Our findings thus reveal a possible novel role of TNTs and lysosomes in the progression of synucleinopathies. © 2016 The Authors.

  20. Markedly diminished epidermal keratinocyte expression of intercellular adhesion molecule-1 (ICAM-1) in Sezary syndrome

    Energy Technology Data Exchange (ETDEWEB)

    Nickoloff, B.J.; Griffiths, E.M.; Baadsgaard, O.; Voorhees, J.J.; Hanson, C.A.; Cooper, K.D. (Univ. of Michigan Medical Center, Ann Arbor (USA))

    1989-04-21

    In mucosis fungoides the malignant T cells express lymphocyte function-associated antigen-1, which allows them to bind to epidermal keratinocytes expressing the gamma interferon-inducible intercellular adhesion molecule-1. In this report, a patient with leukemic-stage mucosis fungoides (Sezary syndrome) had widespread erythematous dermal infiltrates containing malignant T cells, but without any epidermotropism. The authors discovered that the T cells expressed normal amounts of functional lymphocyte function-associated antigen-1, but the keratinocytes did not express significant levels of intercellular adhesion molecule-1, which was probably due to the inability of the malignant T cells to produce gamma interferon. These results support the concept that the inability of malignant T cells to enter the epidermis may contribute to emergence of more clinically aggressive T-cell clones that are no longer confined to the skin, but infiltrate the blood, lymph nodes, and viscera, as is seen in Sezary syndrome.

  1. Intercellular Uptake of Technetium-99m Pertechnetate by Different Types of Cell Lines

    International Nuclear Information System (INIS)

    Safri Zainal Abidin; Raizulnasuha Abdul Rashid; Muhammad Afiq Khairil Anuar; Wan Nordiana A Abd Rahman

    2015-01-01

    The purpose of this study is to determine the technetium-99m pertechnetate ( 99m TcO 4 ) intercellular uptake by different types of cell lines. HeLa, human fetal osteoblast (hFOB), glial and glioma cell lines grown in 6-wells culture plates were incubated with 99m TcO 4 of activity of 200, 400, 600, 800 and 1000 μCi for 30 minutes at 37 degree Celsius and 5 % CO 2 humidified atmosphere. After incubation, the cells were washed 3 times with phosphate buffer saline to remove the extracellular traces of 99m TcO 4 . Measurement of the intercellular 99m TcO 4 into the cells was calculated. The intercellular uptake of 99m TcO 4 was found to be inversely correlate to the radioactivity. HHeLa cell shows the highest uptake followed by hFOB, glial and glioma cell lines. Comparison of uptake between normal and cancer cells present indistinguishable results. The findings of this study suggest that the intercellular uptake of 99m TcO 4 is highly dependent on the type of cells despite no significant different of uptake was found between normal and cancer cell lines. The level of radioactivity is also an important determinant factor that influence the uptake of 99m TcO 4 into the cell. The study will be the first precedent toward understanding the cellular characteristics and pharmacokinetic of non-invasive imaging tracer for future molecular imaging and therapy. (author)

  2. Optimal cellular mobility for synchronization arising from the gradual recovery of intercellular interactions

    International Nuclear Information System (INIS)

    Uriu, Koichiro; Ares, Saúl; Oates, Andrew C; Morelli, Luis G

    2012-01-01

    Cell movement and intercellular signaling occur simultaneously during the development of tissues, but little is known about how movement affects signaling. Previous theoretical studies have shown that faster moving cells favor synchronization across a population of locally coupled genetic oscillators. An important assumption in these studies is that cells can immediately interact with their new neighbors after arriving at a new location. However, intercellular interactions in cellular systems may need some time to become fully established. How movement affects synchronization in this situation has not been examined. Here, we develop a coupled phase oscillator model in which we consider cell movement and the gradual recovery of intercellular coupling experienced by a cell after movement, characterized by a moving rate and a coupling recovery rate, respectively. We find (1) an optimal moving rate for synchronization and (2) a critical moving rate above which achieving synchronization is not possible. These results indicate that the extent to which movement enhances synchrony is limited by a gradual recovery of coupling. These findings suggest that the ratio of time scales of movement and signaling recovery is critical for information transfer between moving cells. (paper)

  3. Intercellular signaling pathways active during intervertebral disc growth, differentiation, and aging.

    Science.gov (United States)

    Dahia, Chitra Lekha; Mahoney, Eric J; Durrani, Atiq A; Wylie, Christopher

    2009-03-01

    Intervertebral discs at different postnatal ages were assessed for active intercellular signaling pathways. To generate a spatial and temporal map of the signaling pathways active in the postnatal intervertebral disc (IVD). The postnatal IVD is a complex structure, consisting of 3 histologically distinct components, the nucleus pulposus, fibrous anulus fibrosus, and endplate. These differentiate and grow during the first 9 weeks of age in the mouse. Identification of the major signaling pathways active during and after the growth and differentiation period will allow functional analysis using mouse genetics and identify targets for therapy for individual components of the disc. Antibodies specific for individual cell signaling pathways were used on cryostat sections of IVD at different postnatal ages to identify which components of the IVD were responding to major classes of intercellular signal, including sonic hedgehog, Wnt, TGFbeta, FGF, and BMPs. We present a spatial/temporal map of these signaling pathways during growth, differentiation, and aging of the disc. During growth and differentiation of the disc, its different components respond at different times to different intercellular signaling ligands. Most of these are dramatically downregulated at the end of disc growth.

  4. The intercellular synchronization of Ca2+ oscillations evaluates Cx36-dependent coupling.

    Directory of Open Access Journals (Sweden)

    Sabine Bavamian

    Full Text Available Connexin36 (Cx36 plays an important role in insulin secretion by controlling the intercellular synchronization of Ca(2+ transients induced during stimulation. The lack of drugs acting on Cx36 channels is a major limitation in further unraveling the molecular mechanism underlying this effect. To screen for such drugs, we have developed an assay allowing for a semi-automatic, fluorimetric quantification of Ca(2+ transients in large populations of MIN6 cells. Here, we show that (1 compared to control cells, MIN6 cells with reduced Cx36 expression or function showed decreased synchrony of glucose-induced Ca(2+ oscillations; (2 glibenclamide, a sulphonylurea which promotes Cx36 junctions and coupling, increased the number of synchronous MIN6 cells, whereas quinine, an antimalarial drug which inhibits Cx36-dependent coupling, decreased this proportion; (3 several drugs were identified that altered the intercellular Ca(2+ synchronization, cell coupling and distribution of Cx36; (4 some of them also affected insulin content. The data indicate that the intercellular synchronization of Ca(2+ oscillations provides a reliable and non-invasive measurement of Cx36-dependent coupling, which is useful to identify novel drugs affecting the function of β-cells, neurons, and neuron-related cells that express Cx36.

  5. The first transmembrane domain (TM1) of β2-subunit binds to the transmembrane domain S1 of α-subunit in BK potassium channels

    Science.gov (United States)

    Morera, Francisco J.; Alioua, Abderrahmane; Kundu, Pallob; Salazar, Marcelo; Gonzalez, Carlos; Martinez, Agustin D.; Stefani, Enrico; Toro, Ligia; Latorre, Ramon

    2012-01-01

    The BK channel is one of the most broadly expressed ion channels in mammals. In many tissues, the BK channel pore-forming α-subunit is associated to an auxiliary β-subunit that modulates the voltage- and Ca2+-dependent activation of the channel. Structural components present in β-subunits that are important for the physical association with the α-subunit are yet unknown. Here, we show through co-immunoprecipitation that the intracellular C-terminus, the second transmembrane domain (TM2) and the extracellular loop of the β2-subunit are dispensable for association with the α-subunit pointing transmembrane domain 1 (TM1) as responsible for the interaction. Indeed, the TOXCAT assay for transmembrane protein–protein interactions demonstrated for the first time that TM1 of the β2-subunit physically binds to the transmembrane S1 domain of the α-subunit. PMID:22710124

  6. MutHTP: Mutations in Human Transmembrane Proteins.

    Science.gov (United States)

    A, Kulandaisamy; S, Binny Priya; R, Sakthivel; Tarnovskaya, Svetlana; Bizin, Ilya; Hönigschmid, Peter; Frishman, Dmitrij; Gromiha, M Michael

    2018-02-01

    We have developed a novel database, MutHTP, which contains information on 183395 disease-associated and 17827 neutral mutations in human transmembrane proteins. For each mutation site MutHTP provides a description of its location with respect to the membrane protein topology, structural environment (if available) and functional features. Comprehensive visualization, search, display and download options are available. The database is publicly available at http://www.iitm.ac.in/bioinfo/MutHTP/. The website is implemented using HTML, PHP and javascript and supports recent versions of all major browsers, such as Firefox, Chrome and Opera. gromiha@iitm.ac.in. Supplementary data are available at Bioinformatics online. © The Author (2018). Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com

  7. Effect of ionizing radiation on transmembrane potential of Streptococcus

    International Nuclear Information System (INIS)

    Fomenko, B.S.; Akoev, I.G.

    1979-01-01

    Treatment of Streptococcus faecalis with ionizing radiation at doses of 5 to 100 krad is shown to reduce the energy-dependent accumulation of dibenzyldimethylammonium (DDA + ) by the cell. Since transmembrane potential is the moving force of DDA + transport across the membrane, the decrease in DDA + accumulation is suggested to be due to potential reduction. This radiation effect was not due to inactivation of the potential-generating mechanism; thus, the ATPase activity and glycolytic activity of the irradiated cells were higher than in the control. At the same time, the membranes exhibited an increased permeability for K + and protons, which is probably due to structural rearrangements in the membranes after irradiation. It is suggested that the potential reduction results from the increase in proton permeability of membranes

  8. Transmembrane helical interactions in the CFTR channel pore.

    Directory of Open Access Journals (Sweden)

    Jhuma Das

    2017-06-01

    Full Text Available Mutations in the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR gene affect CFTR protein biogenesis or its function as a chloride channel, resulting in dysregulation of epithelial fluid transport in the lung, pancreas and other organs in cystic fibrosis (CF. Development of pharmaceutical strategies to treat CF requires understanding of the mechanisms underlying channel function. However, incomplete 3D structural information on the unique ABC ion channel, CFTR, hinders elucidation of its functional mechanism and correction of cystic fibrosis causing mutants. Several CFTR homology models have been developed using bacterial ABC transporters as templates but these have low sequence similarity to CFTR and are not ion channels. Here, we refine an earlier model in an outward (OWF and develop an inward (IWF facing model employing an integrated experimental-molecular dynamics simulation (200 ns approach. Our IWF structure agrees well with a recently solved cryo-EM structure of a CFTR IWF state. We utilize cysteine cross-linking to verify positions and orientations of residues within trans-membrane helices (TMHs of the OWF conformation and to reconstruct a physiologically relevant pore structure. Comparison of pore profiles of the two conformations reveal a radius sufficient to permit passage of hydrated Cl- ions in the OWF but not the IWF model. To identify structural determinants that distinguish the two conformations and possible rearrangements of TMHs within them responsible for channel gating, we perform cross-linking by bifunctional reagents of multiple predicted pairs of cysteines in TMH 6 and 12 and 6 and 9. To determine whether the effects of cross-linking on gating observed are the result of switching of the channel from open to close state, we also treat the same residue pairs with monofunctional reagents in separate experiments. Both types of reagents prevent ion currents indicating that pore blockage is primarily responsible.

  9. Maize plasma membrane aquaporin ZmPIP2;5, but not ZmPIP1;2, facilitates transmembrane diffusion of hydrogen peroxide.

    Science.gov (United States)

    Bienert, Gerd P; Heinen, Robert B; Berny, Marie C; Chaumont, François

    2014-01-01

    Plant aquaporins play important roles in transmembrane water transport processes, but some also facilitate the diffusion of other small uncharged solutes ranging from gases to metalloids. Recent evidence suggests that the transmembrane movement of hydrogen peroxide, an intra- and intercellular multifunctional signaling and defense compound, can be regulated by aquaporins. We addressed the question whether maize aquaporins belonging to the plasma membrane intrinsic protein (PIP) subfamily facilitate hydrogen peroxide diffusion using heterologous expression in the yeast Saccharomyces cerevisiae. We showed that ZmPIP proteins belonging to the PIP1 and PIP2 groups were significantly expressed in yeast cells only after codon optimization of their cDNA. In accordance with previous localization studies in oocytes and plants, ZmPIP1;2 was mainly retained in intracellular membranes, while ZmPIP2;5 was localized to the plasma membrane. However, upon co-expression with ZmPIP2;5, ZmPIP1;2 was re-localized to the plasma membrane. Using a non-functional plasma membrane-localized ZmPIP2;5 mutant to deliver ZmPIP1;2 to the plasma membrane, we demonstrated that, in contrast to wild type ZmPIP2;5, ZmPIP1;2 was not permeable to hydrogen peroxide. Our study further highlighted the fact that, when using the yeast system, which is widely employed to study substrates for plant aquaporins and other transporters, although positive transport assay results allow direct conclusions to be drawn regarding solute permeability, negative results require additional control experiments to show that the protein is expressed and localized correctly before concluding on the lack of transport activity. © 2013.

  10. Involvement of a putative intercellular signal-recognizing G protein-coupled receptor in the engulfment of Salmonella by the protozoan Tetrahymena

    Directory of Open Access Journals (Sweden)

    P.N. Agbedanu

    2013-07-01

    Full Text Available In an effort to investigate the molecular basis of protozoa engulfment-mediated hypervirulence of Salmonella in cattle, we evaluated protozoan G protein-coupled receptors (GPCRs as transducers of Salmonella engulfment by the model protozoan Tetrahymena. Our laboratory previously demonstrated that non-pathogenic protozoa (including Tetrahymena engulf Salmonella and then exacerbate its virulence in cattle, but the mechanistic details of the phenomenon are not fully understood. GPCRs were investigated since these receptors facilitate phagocytosis of particulates by Tetrahymena, and a GPCR apparently modulates bacterial engulfment for the pathogenic protozoan Entamoeba histolytica. A database search identified three putative Tetrahymena GPCRs, based on sequence homologies and predicted transmembrane domains, that were the focus of this study. Salmonella engulfment by Tetrahymena was assessed in the presence of suramin, a non-specific GPCR inhibitor. Salmonella engulfment was also assessed in Tetrahymena in which expression of putative GPCRs was knocked-down using RNAi. A candidate GPCR was then expressed in a heterologous yeast expression system for further characterization. Our results revealed that Tetrahymena were less efficient at engulfing Salmonella in the presence of suramin. Engulfment was reduced concordantly with a reduction in the density of protozoa. RNAi-based studies revealed that knock-down of one the Tetrahymena GPCRs caused diminished engulfment of Salmonella. Tetrahymena lysates activated this receptor in the heterologous expression system. These data demonstrate that the Tetrahymena receptor is a putative GPCR that facilitates bacterial engulfment by Tetrahymena. Activation of the putative GPCR seemed to be related to protozoan cell density, suggesting that its cognate ligand is an intercellular signaling molecule.

  11. Hidden markov model for the prediction of transmembrane proteins using MATLAB.

    Science.gov (United States)

    Chaturvedi, Navaneet; Shanker, Sudhanshu; Singh, Vinay Kumar; Sinha, Dhiraj; Pandey, Paras Nath

    2011-01-01

    Since membranous proteins play a key role in drug targeting therefore transmembrane proteins prediction is active and challenging area of biological sciences. Location based prediction of transmembrane proteins are significant for functional annotation of protein sequences. Hidden markov model based method was widely applied for transmembrane topology prediction. Here we have presented a revised and a better understanding model than an existing one for transmembrane protein prediction. Scripting on MATLAB was built and compiled for parameter estimation of model and applied this model on amino acid sequence to know the transmembrane and its adjacent locations. Estimated model of transmembrane topology was based on TMHMM model architecture. Only 7 super states are defined in the given dataset, which were converted to 96 states on the basis of their length in sequence. Accuracy of the prediction of model was observed about 74 %, is a good enough in the area of transmembrane topology prediction. Therefore we have concluded the hidden markov model plays crucial role in transmembrane helices prediction on MATLAB platform and it could also be useful for drug discovery strategy. The database is available for free at bioinfonavneet@gmail.comvinaysingh@bhu.ac.in.

  12. Structural organization of intercellular channels II. Amino terminal domain of the connexins: sequence, functional roles, and structure.

    Science.gov (United States)

    Beyer, Eric C; Lipkind, Gregory M; Kyle, John W; Berthoud, Viviana M

    2012-08-01

    The amino terminal domain (NT) of the connexins consists of their first 22-23 amino acids. Site-directed mutagenesis studies have demonstrated that NT amino acids are determinants of gap junction channel properties including unitary conductance, permeability/selectivity, and gating in response to transjunctional voltage. The importance of this region has also been emphasized by the identification of multiple disease-associated connexin mutants affecting amino acid residues in the NT region. The first part of the NT is α-helical. The structure of the Cx26 gap junction channel shows that the NT α-helix localizes within the channel, and lines the wall of the pore. Interactions of the amino acid residues in the NT with those in the transmembrane helices may be critical for holding the channel open. The predicted sites of these interactions and the applicability of the Cx26 structure to the NT of other connexins are considered. This article is part of a Special Issue entitled: The Communicating junctions, composition, structure and characteristics. Copyright © 2011. Published by Elsevier B.V.

  13. Regulation Involved in Colonization of Intercellular Spaces of Host Plants in Ralstonia solanacearum

    Directory of Open Access Journals (Sweden)

    Yasufumi Hikichi

    2017-06-01

    Full Text Available A soil-borne bacterium Ralstonia solanacearum invading plant roots first colonizes the intercellular spaces of the root, and eventually enters xylem vessels, where it replicates at high levels leading to wilting symptoms. After invasion into intercellular spaces, R. solanacearum strain OE1-1 attaches to host cells and expression of the hrp genes encoding components of the type III secretion system (T3SS. OE1-1 then constructs T3SS and secrets effectors into host cells, inducing expression of the host gene encoding phosphatidic acid phosphatase. This leads to suppressing plant innate immunity. Then, OE1-1 grows on host cells, inducing quorum sensing (QS. The QS contributes to regulation of OE1-1 colonization of intercellular spaces including mushroom-type biofilm formation on host cells, leading to its virulence. R. solanacearum strains AW1 and K60 produce methyl 3-hydroxypalmitate (3-OH PAME as a QS signal. The methyltransferase PhcB synthesizes 3-OH PAME. When 3-OH PAME reaches a threshold level, it increases the ability of the histidine kinase PhcS to phosphorylate the response regulator PhcR. This results in elevated levels of functional PhcA, the global virulence regulator. On the other hand, strains OE1-1 and GMI1000 produce methyl 3-hydroxymyristate (3-OH MAME as a QS signal. Among R. solanacearum strains, the deduced PhcB and PhcS amino acid sequences are related to the production of QS signals. R. solanacearum produces aryl-furanone secondary metabolites, ralfuranones, which are extracellularly secreted and required for its virulence, dependent on the QS. Interestingly, ralfuranones affect the QS feedback loop. Taken together, integrated signaling via ralfuranones influences the QS, contributing to pathogen virulence.

  14. Gap junction mediated intercellular metabolite transfer in the cochlea is compromised in connexin30 null mice.

    Directory of Open Access Journals (Sweden)

    Qing Chang

    Full Text Available Connexin26 (Cx26 and connexin30 (Cx30 are two major protein subunits that co-assemble to form gap junctions (GJs in the cochlea. Mutations in either one of them are the major cause of non-syndromic prelingual deafness in humans. Because the mechanisms of cochlear pathogenesis caused by Cx mutations are unclear, we investigated effects of Cx30 null mutation on GJ-mediated ionic and metabolic coupling in the cochlea of mice. A novel flattened cochlear preparation was used to directly assess intercellular coupling in the sensory epithelium of the cochlea. Double-electrode patch clamp recordings revealed that the absence of Cx30 did not significantly change GJ conductance among the cochlear supporting cells. The preserved electrical coupling is consistent with immunolabeling data showing extensive Cx26 GJs in the cochlea of the mutant mice. In contrast, dye diffusion assays showed that the rate and extent of intercellular transfer of multiple fluorescent dyes (including a non-metabolizable D-glucose analogue, 2-NBDG among cochlear supporting cells were severely reduced in Cx30 null mice. Since the sensory epithelium in the cochlea is an avascular organ, GJ-facilitated intercellular transfer of nutrient and signaling molecules may play essential roles in cellular homeostasis. To test this possibility, NBDG was used as a tracer to study the contribution of GJs in transporting glucose into the cochlear sensory epithelium when delivered systemically. NBDG uptake in cochlear supporting cells was significantly reduced in Cx30 null mice. The decrease was also observed with GJ blockers or glucose competition, supporting the specificity of our tests. These data indicate that GJs facilitate efficient uptake of glucose in the supporting cells. This study provides the first direct experimental evidence showing that the transfer of metabolically-important molecules in cochlear supporting cells is dependent on the normal function of GJs, thereby suggesting a

  15. Transmembrane Peptides as Sensors of the Membrane Physical State

    Directory of Open Access Journals (Sweden)

    Stefano Piotto

    2018-05-01

    Full Text Available Cell membranes are commonly considered fundamental structures having multiple roles such as confinement, storage of lipids, sustain and control of membrane proteins. In spite of their importance, many aspects remain unclear. The number of lipid types is orders of magnitude larger than the number of amino acids, and this compositional complexity is not clearly embedded in any membrane model. A diffused hypothesis is that the large lipid palette permits to recruit and organize specific proteins controlling the formation of specialized lipid domains and the lateral pressure profile of the bilayer. Unfortunately, a satisfactory knowledge of lipid abundance remains utopian because of the technical difficulties in isolating definite membrane regions. More importantly, a theoretical framework where to fit the lipidomic data is still missing. In this work, we wish to utilize the amino acid sequence and frequency of the membrane proteins as bioinformatics sensors of cell bilayers. The use of an alignment-free method to find a correlation between the sequences of transmembrane portion of membrane proteins with the membrane physical state (MPS suggested a new approach for the discovery of antimicrobial peptides.

  16. Incorporation of transmembrane hydroxide transport into the chemiosmotic theory.

    Science.gov (United States)

    de Grey, A D

    1999-10-01

    A cornerstone of textbook bioenergetics is that oxidative ATP synthesis in mitochondria requires, in normal conditions of internal and external pH, a potential difference (delta psi) of well over 100 mV between the aqueous compartments that the energy-transducing membrane separates. Measurements of delta psi inferred from diffusion of membrane-permeant ions confirm this, but those using microelectrodes consistently find no such delta psi--a result ostensibly irreconcilable with the chemiosmotic theory. Transmembrane hydroxide transport necessarily accompanies mitochondrial ATP synthesis, due to the action of several carrier proteins; this nullifies some of the proton transport by the respiratory chain. Here, it is proposed that these carriers' structure causes the path of this "lost" proton flow to include a component perpendicular to the membrane but within the aqueous phases, so maintaining a steady-state proton-motive force between the water at each membrane surface and in the adjacent bulk medium. The conflicting measurements of delta psi are shown to be consistent with the response of this system to its chemical environment.

  17. Transmembrane topology of the acetylcholine receptor examined in reconstituted vesicles

    International Nuclear Information System (INIS)

    McCrea, P.D.

    1987-01-01

    Each of the five acetylcholine receptor (AChR) subunits, α 2 β-γδ, is believed to have the same number of transmembrane crossing and to share the same general folding pattern. AChR isolated from the electric organ of electric fish is predominantly dimeric. We have used this bridge as a marker for the C-terminus of the δ subunit, and presumably that of the other subunits in addition. The disulfide's accessibility to hydrophilic reductants, principally glutathione (GSH), was tested in a reconstituted vesicle system. The reduction of the δ-δ desulfide, as evidenced by the transition of AChrR dimers to monomers, was quantitatively monitored on velocity sedimentation sucrose gradients. Alternatively, the reduction of δ 2 to δ was followed by employing non-reducing SDS-PAGE. Reductants such as GSH were able to access the bridge in intact right-side-out vesicles. No acceleration of this process was evident when the vesicles were disrupted by freeze-thaw or by detergents. Control experiments which determined the rate of reduction of entrapped diphtheria toxin, or that of 3 H-GSH efflux, demonstrated that intact reconstituted vesicles provide an adequate permeability barrier to GSH access of their intravesicular space

  18. NMR studies of transmembrane electron transport in human erythrocytes

    International Nuclear Information System (INIS)

    Kennett, E.C.; Bubb, W.A.; Kuchel, P.W.

    2002-01-01

    Full text: Electron transport systems exist in the plasma membranes of all cells. These systems appear to play a role in cell growth and proliferation, intracellular signalling, hormone responses, apoptotic events, cell defence and perhaps most importantly they enable the cell to respond to changes in the redox state of both the intra- and extracellular environments. Previously, 13 C NMR has been used to study transmembrane electron transport in human erythrocytes, specifically the reduction of extracellular 13 C-ferricyanide. NMR is a particularly useful tool for studying such systems as changes in the metabolic state of the cell can be observed concomitantly with extracellular reductase activity. We investigated the oxidation of extracellular NADH by human erythrocytes using 1 H and 31 P NMR spectroscopy. Recent results for glucose-starved human erythrocytes indicate that, under these conditions, extracellular NADH can be oxidised at the plasma membrane with the electron transfer across the membrane resulting in reduction of intracellular NAD + . The activity is inhibited by known trans-plasma membrane electron transport inhibitors (capsaicin and atebrin) and is unaffected by inhibition of the erythrocyte Band 3 anion transporter. These results suggest that electron import from extracellular NADH allows the cell to re-establish a reducing environment after the normal redox balance is disturbed

  19. Transmembrane peptides as sensors of the membrane physical state

    Science.gov (United States)

    Piotto, Stefano; Di Biasi, Luigi; Sessa, Lucia; Concilio, Simona

    2018-05-01

    Cell membranes are commonly considered fundamental structures having multiple roles such as confinement, storage of lipids, sustain and control of membrane proteins. In spite of their importance, many aspects remain unclear. The number of lipid types is orders of magnitude larger than the number of amino acids, and this compositional complexity is not clearly embedded in any membrane model. A diffused hypothesis is that the large lipid palette permits to recruit and organize specific proteins controlling the formation of specialized lipid domains and the lateral pressure profile of the bilayer. Unfortunately, a satisfactory knowledge of lipid abundance remains utopian because of the technical difficulties in isolating definite membrane regions. More importantly, a theoretical framework where to fit the lipidomic data is still missing. In this work, we wish to utilize the amino acid sequence and frequency of the membrane proteins as bioinformatics sensors of cell bilayers. The use of an alignment-free method to find a correlation between the sequences of transmembrane portion of membrane proteins with the membrane physical state suggested a new approach for the discovery of antimicrobial peptides.

  20. Crystal structure of the Haemophilus influenzae Hap adhesin reveals an intercellular oligomerization mechanism for bacterial aggregation

    Science.gov (United States)

    Meng, Guoyu; Spahich, Nicole; Kenjale, Roma; Waksman, Gabriel; St Geme, Joseph W

    2011-01-01

    Bacterial biofilms are complex microbial communities that are common in nature and are being recognized increasingly as an important determinant of bacterial virulence. However, the structural determinants of bacterial aggregation and eventual biofilm formation have been poorly defined. In Gram-negative bacteria, a major subgroup of extracellular proteins called self-associating autotransporters (SAATs) can mediate cell–cell adhesion and facilitate biofilm formation. In this study, we used the Haemophilus influenzae Hap autotransporter as a prototype SAAT to understand how bacteria associate with each other. The crystal structure of the H. influenzae HapS passenger domain (harbouring the SAAT domain) was determined to 2.2 Å by X-ray crystallography, revealing an unprecedented intercellular oligomerization mechanism for cell–cell interaction. The C-terminal SAAT domain folds into a triangular-prism-like structure that can mediate Hap–Hap dimerization and higher degrees of multimerization through its F1–F2 edge and F2 face. The intercellular multimerization can give rise to massive buried surfaces that are required for overcoming the repulsive force between cells, leading to bacterial cell–cell interaction and formation of complex microcolonies. PMID:21841773

  1. Intercellular distribution of mutations induced in oopcytes of Drosophila melanogaster by chemical and physical mutagens

    International Nuclear Information System (INIS)

    Traut, H.

    1979-01-01

    When females of Drosophila melanogaster are treated with chemical or physical mutagens, not only in one but also in both of the two homologous X chromosomes of a given oocyte, a recessive sex-linked lethal mutation may be induced. A method is described that discriminates between such single and double mutations. A theory is developed to show how a comparison betweeen the expected and the observer frequency of double mutations yields an indication of the intercellular distribution (random or nonrandom) of recessive lethal mutations induced by mutagenic agents in oocytes and, consequently, of the distribution (homogenous or nonhomogeneous) of those agents. Three agents were tested: FUdR (12.5, 50.0 and 81.0 μg/ml), mitomycin C (130.0 μg/ml) and x rays (2000 R, 150 kV). After FUdR feeding, no increase in the mutation frequency usually observed in D. melanogaster without mutagenic treatment was obtained (u = 0.13%, namely three single mutations among 2332 chromosomes tested). After mitomycin C feeding 104 single and three double mutations were obtained. All of the 50 mutations observed after x irradiation were single mutations. The results obtained in the mitomycin C and radiation experiments favor the assumption of a random intercellular distribution of recessive lethal mutations induced by these two agents in oocytes of D. melanogaster. Reasons are discussed why for other types of mutagenic agents nonrandom distributions may be observed with our technique

  2. CHANGES OF INTERCELLULAR COOPERATION IN PERIPHERAL BLOOD IN TREATED PATIENTS WITH CARDIOLOGIC DISEASES

    Directory of Open Access Journals (Sweden)

    L. N. Korichkina

    2009-01-01

    Full Text Available Aim. To study changes of intercellular cooperation in peripheral blood induced by treatment in patients with arterial hypertension (HT, ischemic heart disease (IHD and chronic heart failure (CHF.Material and methods. 610 patients were involved into the study, including 250 patients with HT of stages I-III (50 untreated patients, 150 patients with IHD and 210 patients with CHF of stages I-III. All patients were treated except 50 hypertensive ones. 80 healthy patients (40 men, 40 women were included into control group. Blood smears of patients were evaluated (Romanovsky's stain. A number of leukocyte, autorosettes and autorosettes with erythrocyte lysis was calculated. The cellular association consisting of a neutrophil, monocyte or eosinocyte with 3 or more erythrocytes skintight to their surface defined as autorosettes. Erythrocytes number and hemoglobin level determined in peripheral blood.Results. Single autorosettes in peripheral blood were observed in patients of control group and in untreated patients with HT. Treated patients with HT, IHD and CHF had increased number of autorossets and autorosettes with erythrocytes lysis. This phenomenon resulted in reduction of erythrocytes number and hemoglobin level in peripheral blood.Conclusion. Treated patients with cardiologic diseases had changes in intercellular cooperation. It should be considered at intensive and long term therapy.

  3. Intercellular signaling pathways active during and after growth and differentiation of the lumbar vertebral growth plate.

    Science.gov (United States)

    Dahia, Chitra Lekha; Mahoney, Eric J; Durrani, Atiq A; Wylie, Christopher

    2011-06-15

    Vertebral growth plates at different postnatal ages were assessed for active intercellular signaling pathways. To generate a spatial and temporal map of the major signaling pathways active in the postnatal mouse lumbar vertebral growth plate. The growth of all long bones is known to occur by cartilaginous growth plates. The growth plate is composed of layers of chondrocyets that actively proliferate, differentiate, die and, are replaced by bone. The role of major cell signaling pathways has been suggested for regulation of the fetal long bones. But not much is known about the molecular or cellular signals that control the postnatal vertebral growth plate and hence postnatal vertebral bone growth. Understanding such molecular mechanisms will help design therapeutic treatments for vertebral growth disorders such as scoliosis. Antibodies against activated downstream intermediates were used to identify cells in the growth plate responding to BMP, TGFβ, and FGF in cryosections of lumbar vertebrae from different postnatal age mice to identify the zones that were responding to these signals. Reporter mice were used to identify the chondrocytes responding to hedgehog (Ihh), and Wnt signaling. We present a spatial/temporal map of these signaling pathways during growth, and differentiation of the mouse lumbar vertebral growth plate. During growth and differentiation of the vertebral growth plate, its different components respond at different times to different intercellular signaling ligands. Response to most of these signals is dramatically downregulated at the end of vertebral growth.

  4. Intercellular adhesion molecule-1 blockade attenuates inflammatory response and improves microvascular perfusion in rat pancreas grafts.

    Science.gov (United States)

    Preissler, Gerhard; Eichhorn, Martin; Waldner, Helmut; Winter, Hauke; Kleespies, Axel; Massberg, Steffen

    2012-10-01

    After pancreas transplantation (PTx), early capillary malperfusion and leukocyte recruitment indicate the manifestation of severe ischemia/reperfusion injury (IRI). Oscillatory blood-flow redistribution (intermittent capillary perfusion, IP), leading to an overall decrease in erythrocyte flux, precedes complete microvascular perfusion failure with persistent blood flow cessation. We addressed the role of intercellular adhesion molecule-1 (ICAM-1) for leukocyte-endothelial interactions (LEIs) after PTx and evaluated the contribution of IP and malperfusion. Pancreas transplantation was performed in rats after 18-hour preservation, receiving either isotype-matched IgG or monoclonal anti-ICAM-1 antibodies (10 mg/kg intravenously) once before reperfusion. Leukocyte-endothelial interaction, IP, erythrocyte flux, and functional capillary density, respectively, were examined in vivo during 2-hour reperfusion. Nontransplanted animals served as controls. Tissue samples were analyzed by histomorphometry. In grafts of IgG-treated animals, IP was encountered already at an early stage after reperfusion and steadily increased over 2 hours, whereas erythrocyte flux declined continuously. In contrast, inhibition of ICAM-1 significantly improved erythrocyte flux and delayed IP appearance by 2 hours. Further, anti-ICAM-1 significantly reduced LEI and leukocyte tissue infiltration when compared to IgG; edema development was less pronounced in response to anti-ICAM-1 monoclonal antibody. Intercellular adhesion molecule-1 blockade significantly attenuates IRI via immediate reduction of LEI and concomitant improvement of capillary perfusion patterns, emphasizing its central role during IRI in PTx.

  5. Molecular pharmacology of promiscuous seven transmembrane receptors sensing organic nutrients.

    Science.gov (United States)

    Wellendorph, Petrine; Johansen, Lars Dan; Bräuner-Osborne, Hans

    2009-09-01

    A number of highly promiscuous seven transmembrane (7TM) receptors have been cloned and characterized within the last few years. It is noteworthy that many of these receptors are activated broadly by amino acids, proteolytic degradation products, carbohydrates, or free fatty acids and are expressed in taste tissue, the gastrointestinal tract, endocrine glands, adipose tissue, and/or kidney. These receptors thus hold the potential to act as sensors of food intake, regulating, for example, release of incretin hormones from the gut, insulin/glucagon from the pancreas, and leptin from adipose tissue. The promiscuous tendency in ligand recognition of these receptors is in contrast to the typical specific interaction with one physiological agonist seen for most receptors, which challenges the classic "lock-and-key" concept. We here review the molecular mechanisms of nutrient sensing of the calcium-sensing receptor, the G protein-coupled receptor family C, group 6, subtype A (GPRC6A), and the taste1 receptor T1R1/T1R3, which are sensing L-alpha-amino acids, the carbohydrate-sensing T1R2/T1R3 receptor, the proteolytic degradation product sensor GPR93 (also termed GPR92), and the free fatty acid (FFA) sensing receptors FFA1, FFA2, FFA3, GPR84, and GPR120. The involvement of the individual receptors in sensing of food intake has been validated to different degrees because of limited availability of specific pharmacological tools and/or receptor knockout mice. However, as a group, the receptors represent potential drug targets, to treat, for example, type II diabetes by mimicking food intake by potent agonists or positive allosteric modulators. The ligand-receptor interactions of the promiscuous receptors of organic nutrients thus remain an interesting subject of emerging functional importance.

  6. Functional characterization of transmembrane adenylyl cyclases from the honeybee brain.

    Science.gov (United States)

    Balfanz, Sabine; Ehling, Petra; Wachten, Sebastian; Jordan, Nadine; Erber, Joachim; Mujagic, Samir; Baumann, Arnd

    2012-06-01

    The second messenger cAMP has a pivotal role in animals' physiology and behavior. Intracellular concentrations of cAMP are balanced by cAMP-synthesizing adenylyl cyclases (ACs) and cAMP-cleaving phosphodiesterases. Knowledge about ACs in the honeybee (Apis mellifera) is rather limited and only an ortholog of the vertebrate AC3 isoform has been functionally characterized, so far. Employing bioinformatics and functional expression we characterized two additional honeybee genes encoding membrane-bound (tm)ACs. The proteins were designated AmAC2t and AmAC8. Unlike the common structure of tmACs, AmAC2t lacks the first transmembrane domain. Despite this unusual topography, AmAC2t-activity could be stimulated by norepinephrine and NKH477 with EC(50s) of 0.07 μM and 3 μM. Both ligands stimulated AmAC8 with EC(50s) of 0.24 μM and 3.1 μM. In brain cryosections, intensive staining of mushroom bodies was observed with specific antibodies against AmAC8, an expression pattern highly reminiscent of the Drosophila rutabaga AC. In a current release of the honeybee genome database we identified three additional tmAC- and one soluble AC-encoding gene. These results suggest that (1) the AC-gene family in honeybees is comparably large as in other species, and (2) based on the restricted expression of AmAC8 in mushroom bodies, this enzyme might serve important functions in honeybee behavior. Copyright © 2012 Elsevier Ltd. All rights reserved.

  7. Biological amine transport in chromaffin ghosts. Coupling to the transmembrane proton and potential gradients.

    Science.gov (United States)

    Johnson, R G; Pfister, D; Carty, S E; Scarpa, A

    1979-11-10

    The effect of the transmembrane proton gradient (delta pH) and potential gradient (delta psi) upon the rate and extent of amine accumulation was investigated in chromaffin ghosts. The chromaffin ghosts were formed by hypo-osmotic lysis of isolated bovine chromaffin granules and extensive dialysis in order to remove intragranular binding components and dissipate the endogenous electrochemical gradients. Upon ATP addition to suspensions of chromaffin ghosts, a transmembrane proton gradient alone, a transmembrane gradient alone, or both, could be established, depending upon the compositions of the media in which the ghosts were formed and resuspended. When chloride was present in the medium, addition of ATP resulted in the generation of a transmembrane proton gradient, acidic inside of 1 pH unit (measured by [14C]methylamine distribution), and no transmembrane potential (measured by [14C]-thiocyanate distribution). When ATP was added to chromaffin ghosts suspended in a medium in which chloride was substituted by isethionate, a transmembrane potential, inside positive, of 45 mV and no transmembrane proton gradient, was measured. In each medium, the addition of agents known to affect proton or potential gradients, respectively, exerted a predictable mechanism of action. Accumulation of [14C]epinephrine or [14C]5-hydroxytryptamine was over 1 order of magnitude greater in the presence of the transmembrane proton gradient or the transmembrane potential than in the absence of any gradient and, moreover, was related to the magnitude of the proton or potential gradient in a dose-dependent manner. When ghosts were added to a medium containing chloride and isethionate, both a delta pH and delta psi could be generated upon addition of ATP. In this preparation, the maximal rate of amine accumulation was observed. The results indicate that amine accumulation into chromaffin ghosts can occur in the presence of either a transmembrane proton gradient, or a transmembrane potential

  8. The status of intercellular junctions in established lens epithelial cell lines.

    Science.gov (United States)

    Dave, Alpana; Craig, Jamie E; Sharma, Shiwani

    2012-01-01

    Cataract is the major cause of vision-related disability worldwide. Mutations in the crystallin genes are the most common known cause of inherited congenital cataract. Mutations in the genes associated with intercellular contacts, such as Nance-Horan Syndrome (NHS) and Ephrin type A receptor-2 (EPHA2), are other recognized causes of congenital cataract. The EPHA2 gene has been also associated with age-related cataract, suggesting that intercellular junctions are important in not only lens development, but also in maintaining lens transparency. The purpose of this study was to analyze the expression and localization of the key cell junction and cytoskeletal proteins, and of NHS and EPHA2, in established lens epithelial cell lines to determine their suitability as model epithelial systems for the functional investigation of genes involved in intercellular contacts and implicated in cataract. The expression and subcellular localization of occludin and zona occludens protein-1 (ZO-1), which are associated with tight junctions; E-cadherin, which is associated with adherence junctions; and the cytoskeletal actin were analyzed in monolayers of a human lens epithelial cell line (SRA 01/04) and a mouse lens epithelial cell line (αTN4). In addition, the expression and subcellular localization of the NHS and EPHA2 proteins were analyzed in these cell lines. Protein or mRNA expression was respectively determined by western blotting or reverse transcription-polymerase chain reaction (RT-PCR), and localization was determined by immunofluorescence labeling. Human SRA 01/04 and mouse αTN4 lens epithelial cells expressed either the proteins of interest or their encoding mRNA. Occludin, ZO-1, and NHS proteins localized to the cellular periphery, whereas E-cadherin, actin, and EPHA2 localized in the cytoplasm in these cell lines. The human SRA 01/04 and mouse αTN4 lens epithelial cells express the key junctional proteins. The localization patterns of these proteins suggest that

  9. Signaling through intercellular adhesion molecule 1 (ICAM-1) in a B cell lymphoma line

    DEFF Research Database (Denmark)

    Holland, J; Owens, T

    1997-01-01

    Intercellular adhesion molecule 1 (ICAM-1) (CD54) is an adhesion molecule of the immunoglobulin superfamily. The interaction between ICAM-1 on B lymphocytes and leukocyte function-associated antigen 1 on T cells plays a major role in several aspects of the immune response, including T-dependent B...... cell activation. While it was originally believed that ICAM-1 played a purely adhesive role, recent evidence suggests that it can itself transduce biochemical signals. We demonstrate that cross-linking of ICAM-1 results in the up-regulation of class II major histocompatibility complex, and we...... investigate the biochemical mechanism for the signaling role of ICAM-1. We show that cross-linking of ICAM-1 on the B lymphoma line A20 induces an increase in tyrosine phosphorylation of several cellular proteins, including the Src family kinase p53/p56(lyn). In vitro kinase assays showed that Lyn kinase...

  10. 3D Architecture of the Trypanosoma brucei Flagella Connector, a Mobile Transmembrane Junction.

    Directory of Open Access Journals (Sweden)

    Johanna L Höög

    2016-01-01

    Full Text Available Cellular junctions are crucial for the formation of multicellular organisms, where they anchor cells to each other and/or supportive tissue and enable cell-to-cell communication. Some unicellular organisms, such as the parasitic protist Trypanosoma brucei, also have complex cellular junctions. The flagella connector (FC is a three-layered transmembrane junction that moves with the growing tip of a new flagellum and attaches it to the side of the old flagellum. The FC moves via an unknown molecular mechanism, independent of new flagellum growth. Here we describe the detailed 3D architecture of the FC suggesting explanations for how it functions and its mechanism of motility.We have used a combination of electron tomography and cryo-electron tomography to reveal the 3D architecture of the FC. Cryo-electron tomography revealed layers of repetitive filamentous electron densities between the two flagella in the interstitial zone. Though the FC does not change in length and width during the growth of the new flagellum, the interstitial zone thickness decreases as the FC matures. This investigation also shows interactions between the FC layers and the axonemes of the new and old flagellum, sufficiently strong to displace the axoneme in the old flagellum. We describe a novel filament, the flagella connector fibre, found between the FC and the axoneme in the old flagellum.The FC is similar to other cellular junctions in that filamentous proteins bridge the extracellular space and are anchored to underlying cytoskeletal structures; however, it is built between different portions of the same cell and is unique because of its intrinsic motility. The detailed description of its structure will be an important tool to use in attributing structure / function relationships as its molecular components are discovered in the future. The FC is involved in the inheritance of cell shape, which is important for the life cycle of this human parasite.

  11. Conformational constraining of inactive and active States of a seven transmembrane receptor by metal ion site engineering in the extracellular end of transmembrane segment V

    DEFF Research Database (Denmark)

    Rosenkilde, Mette M; David, Ralf; Oerlecke, Ilka

    2006-01-01

    The extracellular part of transmembrane segment V (TM-V) is expected to be involved in the activation process of 7TM receptors, but its role is far from clear. Here, we study the highly constitutively active CXC-chemokine receptor encoded by human herpesvirus 8 (ORF74-HHV8), in which a metal ion ...

  12. Detection of the intercellular adhesion gene cluster (ica in clinical Staphylococcus aureus isolates

    Directory of Open Access Journals (Sweden)

    Namvar, Amirmorteza Ebrahimzadeh

    2013-04-01

    Full Text Available [english] is a major hospital and community pathogen having the aptitude to cause a wide variety of infections in men. The ability of microorganisms to produce biofilm facilitates them to withstand the host immune response and is recognized as one factor contributing to chronic or persistent infections. It was demonstrated that the -encoded genes lead to the biosynthesis of polysaccharide intercellular adhesion (PIA molecules, and may be involved in the accumulation phase of biofilm formation. Different studies have shown the decisive role of the gene as virulence factors in staphylococcal infections. This study was carried out to demonstrate the relationship between gene and production of slime layer in strains. Sixty strains were isolated from patients. The isolates were identified morphologically and biochemically following standard laboratory methods. After identification, the staphylococcal isolates were maintained in trypticase soy broth (TSB, to which 15% glycerol was added, and stored at –20°C. Slime formation and biofilm assay was monitored. A PCR assay was developed to identify the presence of (intercellular adhesion gene gene in all isolates. Thirty-nine slime producing colonies with CRA plates (65% formed black colors, the remaining 21 isolates were pink (35%. In the quantitative biofilm assay 35 (58% produced biofilm while 25 (42% isolates did not exhibit this property. All isolates were positive for detection of gene by PCR method. The interaction of and in the investigated isolates may be important in slime layer formation and biofilm phenomena.We propose PCR detection of the gene locus as a rapid and effective method to be used for discrimination between potentially virulent and nonvirulent isolates, with implications for therapeutic and preventive measures pertainin to the management of colonized indwelling catheters.

  13. Intercellular contact: its influence on the Dsub(q) of mammalian cell survival curves

    International Nuclear Information System (INIS)

    Durand, R.E.; Sutherland, R.M.

    1975-01-01

    Cell survival in tissues exposed to a given dose of ionizing radiation is usually greater than that of similar cells grown individually in vitro, despite the fact that the radiosensitivities (D 0 ) are virtually identical under the two conditions. An analogous increase in cell survival is observed when Chinese hamster V79-171 cells are grown in suspension culture and irradiated as multicell spheroids. Unfortunately, the information gained from the survival curves so obtained is limited by the inhomogeneity of the cell population with respect to both degree of contact and cell cycle position. The latter can be studied using synchronized small spheroids. The ratio of Dsub(q) of spheroid cells to Dsub(q) of single cells increased as the cells progressed through the cell cycle, from a minimum of 1.3 for G 1 phase cells to a maximum of 2.2 for late S-phase cells. The enhanced survival, or 'contact effect', developed slowly as the spheroids grew, after an initial latent period of about one generation cycle of the cells. A second effect of intercellular contact on mammalian cell survival has also been observed. When cells are assayed under conditions in which intercellular contact is maintained, the net cellular survival is increased further. This effect is different from the usual repair of potentially lethal damage, in that it occurs much more slowly and results in modification of the survival-curve shoulder. Not all cell types tested have shown enhanced survival when grown as spheroids. Several MNNG-induced mutants of the Chinese hamster V79-171 line have been isolated and sublines which do and do not show the contact effect are now available. These may permit study of the mechanism(s) of contact effects. (author)

  14. In vitro early changes in intercellular junctions by treatment with a chemical carcinogen.

    Science.gov (United States)

    Tachikawa, T; Kohno, Y; Matsui, Y; Yoshiki, S

    1986-06-01

    To examine early intercellular junction changes caused by treatment with 9,10-dimethyl-1,2-benzanthracene (DMBA), rat lingual epithelium was cultivated in isolation and observed by electrophysiological, freeze-fracture and whole-mount electron microscopy. Electrophysiological measurements showed a transient decrease in membrane potential of -10.2 mV 6 h after the treatment. It returned to almost the same level as that of the control group 1 day later. Six hours after treatment, input resistance decreased rapidly to 5.3 M omega but increased to 18.0 M omega 12 h after treatment. Transient reduction of input resistance and membrane potential occurred prior to the decrease in the coupling ratio 6 h after treatment with DMBA. In freeze-fracture replicas, the number of gap junctions decreased by approximately 45% of the control value 6 h after treatment with DMBA. At 12 h and thereafter, the number and area of gap junctions subsequently decreased by 60-80% of the control value. Alterations in the number and area of desmosomes were similar to those of the gap junctions. The formation of epithelial cytoskeletons, partially devoid of the 2-4 and 5-8 nm filaments was also observed. A decrease in the density of filament networks beneath the plasma membranes was especially apparent. Treatment with a carcinogen brought about morphological cellular changes as early as 6 h after treatment, and such early changes might trigger metabolic cellular abnormalities. Affected cells appear to move away from normal cells in a process of repeated destruction and revision of intercellular junctions, and cytoskeletons.

  15. Interleukin-4 and interleukin-13 compromise the sinonasal epithelial barrier and perturb intercellular junction protein expression.

    Science.gov (United States)

    Wise, Sarah K; Laury, Adrienne M; Katz, Elizabeth H; Den Beste, Kyle A; Parkos, Charles A; Nusrat, Asma

    2014-05-01

    Altered expression of epithelial intercellular junction proteins has been observed in sinonasal biopsies from nasal polyps and epithelial layers cultured from nasal polyp patients. These alterations comprise a "leaky" epithelial barrier phenotype. We hypothesize that T helper 2 (Th2) cytokines interleukin (IL)-4 and IL-13 modulate epithelial junction proteins, thereby contributing to the leaky epithelial barrier. Differentiated primary sinonasal epithelial layers cultured at the air-liquid interface were exposed to IL-4, IL-13, and controls for 24 hours at 37°C. Epithelial resistance measurements were taken every 4 hours during cytokine exposure. Western blot and immunofluorescence staining/confocal microscopy were used to assess changes in a panel of tight and adherens junction proteins. Western blot densitometry was quantified with image analysis. IL-4 and IL-13 exposure resulted in a mean decrease in transepithelial resistance at 24 hours to 51.6% (n = 6) and 68.6% (n = 8) of baseline, respectively. Tight junction protein junctional adhesion molecule-A (JAM-A) expression decreased 42.2% with IL-4 exposure (n = 9) and 37.5% with IL-13 exposure (n = 9). Adherens junction protein E-cadherin expression decreased 35.3% with IL-4 exposure (n = 9) and 32.9% with IL-13 exposure (n = 9). Tight junction protein claudin-2 showed more variability but had a trend toward higher expression with Th2 cytokine exposure. There were no appreciable changes in claudin-1, occludin, or zonula occludens-1 (ZO-1) with IL-4 or IL-13 exposure. Sinonasal epithelial exposure to Th2 cytokines IL-4 and IL-13 results in alterations in intercellular junction proteins, reflecting increased epithelial permeability. Such changes may explain some of the phenotypic manifestations of Th2-mediated sinonasal disease, such as edema, nasal discharge, and environmental reactivity. © 2014 ARS-AAOA, LLC.

  16. IL-4 and IL-13 Compromise the Sinonasal Epithelial Barrier and Perturb Intercellular Junction Protein Expression

    Science.gov (United States)

    Wise, Sarah K.; Laury, Adrienne M.; Katz, Elizabeth H.; Den Beste, Kyle A.; Parkos, Charles A.; Nusrat, Asma

    2014-01-01

    Introduction Altered expression of epithelial intercellular junction proteins has been observed in sinonasal biopsies from nasal polyps and epithelial layers cultured from nasal polyp patients. These alterations comprise a “leaky” epithelial barrier phenotype. We hypothesize that Th2 cytokines IL-4 and IL-13 modulate epithelial junction proteins thereby contributing to the leaky epithelial barrier. Methods Differentiated primary sinonasal epithelial layers cultured at the air-liquid interface were exposed to IL-4, IL-13, and controls for 24 hours at 37°C. Epithelial resistance measurements were taken every 4 hours during cytokine exposure. Western blot and immunofluorescence staining/confocal microscopy were used to assess changes in a panel of tight and adherens junction proteins. Western blot densitometry was quantified with image analysis. Results IL-4 and IL-13 exposure resulted in a mean decrease in transepithelial resistance at 24 hours to 51.6% (n=6) and 68.6% (n=8) of baseline, respectively. Tight junction protein JAM-A expression decreased 42.2% with IL-4 exposure (n=9) and 37.5% with IL-13 exposure (n=9). Adherens junction protein E-cadherin expression decreased 35.3% with IL-4 exposure (n=9) and 32.9% with IL-13 exposure (n=9). Tight junction protein claudin-2 showed more variability but had a trend toward higher expression with Th2 cytokine exposure. There were no appreciable changes in claudin-1, occludin, or ZO-1 with IL-4 or IL-13 exposure. Conclusion Sinonasal epithelial exposure to Th2 cytokines IL-4 and IL-13 results in alterations in intercellular junction proteins, reflecting increased epithelial permeability. Such changes may explain some of the phenotypic manifestations of Th2-mediated sinonasal disease, such as edema, nasal discharge, and environmental reactivity. PMID:24510479

  17. Transformed hairy roots of Discaria trinervis: a valuable tool for studying actinorhizal symbiosis in the context of intercellular infection.

    Science.gov (United States)

    Imanishi, Leandro; Vayssières, Alice; Franche, Claudine; Bogusz, Didier; Wall, Luis; Svistoonoff, Sergio

    2011-11-01

    Among infection mechanisms leading to root nodule symbiosis, the intercellular infection pathway is probably the most ancestral but also one of the least characterized. Intercellular infection has been described in Discaria trinervis, an actinorhizal plant belonging to the Rosales order. To decipher the molecular mechanisms underlying intercellular infection with Frankia bacteria, we set up an efficient genetic transformation protocol for D. trinervis based on Agrobacterium rhizogenes. We showed that composite plants with transgenic roots expressing green fluorescent protein can be specifically and efficiently nodulated by Frankia strain BCU110501. Nitrogen fixation rates and feedback inhibition of nodule formation by nitrogen were similar in control and composite plants. In order to challenge the transformation system, the MtEnod11 promoter, a gene from Medicago truncatula widely used as a marker for early infection-related symbiotic events in model legumes, was introduced in D. trinervis. MtEnod11::GUS expression was related to infection zones in root cortex and in the parenchyma of the developing nodule. The ability to study intercellular infection with molecular tools opens new avenues for understanding the evolution of the infection process in nitrogen-fixing root nodule symbioses.

  18. Intercellular deposits of basement membrane material in active human pituitary adenomas detected by immunostaining for laminin and electron microscopy

    DEFF Research Database (Denmark)

    Holck, S; Wewer, U M; Albrechtsen, R

    1986-01-01

    and one patient with Cushing's syndrome). Concurrently, at the ultrastructural level, bunches of basement membrane-like material intermingled between the adenoma cells were demonstrated in seven of these ten active adenomas. Furthermore, secretory granules were entrapped occasionally in this intercellular...

  19. Radial distribution of dilated intercellular spaces of the esophageal squamous epithelium in patients with reflux disease exhibiting discrete endoscopic lesions

    NARCIS (Netherlands)

    Vieth, M; Fiocca, R; Haringsma, J; Delarive, J; Wiesel, PH; Tam, W; Tytgat, GNJ; Dent, J; Edebo, A; Lundell, L; Stolte, M

    2004-01-01

    Introduction: Dilatation of intercellular spaces of the esophageal squamous epithelium has been suggested as a marker of early acid reflux-induced damage. This change is a potentially useful addition to histomorphological changes that represent so called minimal endoscopic lesions. We have assessed

  20. Transmembrane-sequence-dependent overexpression and secretion of glycoproteins in Saccharomyces cerevisiae.

    Science.gov (United States)

    Schuster, M; Wasserbauer, E; Aversa, G; Jungbauer, A

    2001-02-01

    Protein expression using the secretory pathway in Saccharomyces cerevisiae can lead to high amounts of overexpressed and secreted proteins in culture supernatants in a short period of time. These post-translational modified expression products can be purified up to >90% in a single step. The overexpression and secretion of the transmembrane glycoprotein signaling lymphocytic activation molecule (SLAM) was studied. SLAM belongs to the immunoglobulin superfamily and its engagement results in T-cell expansion and INF-gamma production. The molecule is composed of an extracellular, a single-span transmembrane and a cytoplasmatic domain. The extracellular part may be relevant for stimulation studies in vitro since SLAM is a high-affinity self-ligand. Therefore several fragments of this region have been expressed as Flag-fusions in S. cerevisiae: a full-length fragment containing the transmembrane region and the autologous signal sequence, another without the transmembrane region, and two fragments without the autologous signal sequence with and without the transmembrane region. By molecular cloning, the different deletion mutants of the cDNA encoding the full-length construct have been inserted in a yeast episomal plasmid. Upstream of the cDNA, the alpha-leader sequence of a yeast mating pheromone has been cloned to direct the fusion proteins into the secretory protein maturation pathway. All four fragments were expressed but yield, location, and maturation were highly influenced by the transmembrane domain and the autologous signal sequence. Only the fragment without autologous signal sequence and transmembrane domain could be efficiently secreted. High-mannose glycosylation was analyzed by lectin mapping and digestion with specific glycosidases. After enzyme treatment, a single band product with the theoretical size could be detected and identified as SLAM by a specific monoclonal antibody. The fusion protein concentration in the supernatant was 30 microg/ml. The

  1. Acquired Tumor Cell Radiation Resistance at the Treatment Site Is Mediated Through Radiation-Orchestrated Intercellular Communication

    Energy Technology Data Exchange (ETDEWEB)

    Aravindan, Natarajan, E-mail: naravind@ouhsc.edu [Department of Radiation Oncology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma (United States); Aravindan, Sheeja; Pandian, Vijayabaskar; Khan, Faizan H.; Ramraj, Satish Kumar; Natt, Praveen [Department of Radiation Oncology, University of Oklahoma Health Sciences Center, Oklahoma City, Oklahoma (United States); Natarajan, Mohan [Department of Pathology, The University of Texas Health Science Center at San Antonio, San Antonio, Texas (United States)

    2014-03-01

    Purpose: Radiation resistance induced in cancer cells that survive after radiation therapy (RT) could be associated with increased radiation protection, limiting the therapeutic benefit of radiation. Herein we investigated the sequential mechanistic molecular orchestration involved in radiation-induced radiation protection in tumor cells. Results: Radiation, both in the low-dose irradiation (LDIR) range (10, 50, or 100 cGy) or at a higher, challenge dose IR (CDIR), 4 Gy, induced dose-dependent and sustained NFκB-DNA binding activity. However, a robust and consistent increase was seen in CDIR-induced NFκB activity, decreased DNA fragmentation, apoptosis, and cytotoxicity and attenuation of CDIR-inhibited clonal expansion when the cells were primed with LDIR prior to challenge dose. Furthermore, NFκB manipulation studies with small interfering RNA (siRNA) silencing or p50/p65 overexpression unveiled the influence of LDIR-activated NFκB in regulating CDIR-induced DNA fragmentation and apoptosis. LDIR significantly increased the transactivation/translation of the radiation-responsive factors tumor necrosis factor-α (TNF-α), interleukin-1α (IL-1α), cMYC, and SOD2. Coculture experiments exhibit LDIR-influenced radiation protection and increases in cellular expression, secretion, and activation of radiation-responsive molecules in bystander cells. Individual gene-silencing approach with siRNAs coupled with coculture studies showed the influence of LDIR-modulated TNF-α, IL-1α, cMYC, and SOD2 in induced radiation protection in bystander cells. NFκB inhibition/overexpression studies coupled with coculture experiments demonstrated that TNF-α, IL-1α, cMYC, and SOD2 are selectively regulated by LDIR-induced NFκB. Conclusions: Together, these data strongly suggest that scattered LDIR-induced NFκB-dependent TNF-α, IL-1α, cMYC, and SOD2 mediate radiation protection to the subsequent challenge dose in tumor cells.

  2. [Bacterial synthesis, purification, and solubilization of transmembrane segments of ErbB family members].

    Science.gov (United States)

    Goncharuk, M V; Shul'ga, A A; Ermoliuk, Ia S; Tkach, E N; Goncharuk, S A; Pustovalova, Iu E; Mineev, K S; Bocharov, É V; Maslennikov, I V; Arsen'ev, A S; Kirpichnikov, M P

    2011-01-01

    A family of epidermal growth factor receptors, ErbB, represents an important class of receptor tyrosine kinases, playing a leading role in cellular growth, development and differentiation. Transmembrane domains of these receptors transduce biochemical signals across plasma membrane via lateral homo- and heterodimerization. Relatively small size of complexes of ErbB transmembrane domains with detergents or lipids allows one to study their detailed spatial structure using three-dimensional heteronuclear high-resolution NMR spectroscopy. Here, we describe the effective expression system and purification procedure for preparative-scale production of transmembrane peptides from four representatives of ErbB family, ErbB1, ErbB2, ErbB3, ErbB4, for structural studies. The recombinant peptides were produced in Escherichia coli BL21(DE3)pLysS as C-terminal extensions of thioredoxin A. The fusion protein cleavage was accomplished with the light subunit of human enterokinase. Several (10-30) milligrams of purified isotope-labeled transmembrane peptides were isolated with the use of a simple and convenient procedure, which consists of consecutive steps of immobilized metal affinity chromatography and cation-exchange chromatography. The purified peptides were reconstituted in lipid/detergent environment (micelles or bicelles) and characterized using dynamic light scattering, CD and NMR spectroscopy. The data obtained indicate that the purified ErbB transmembrane peptides are suitable for structural and dynamic studies of their homo- and heterodimer complexes using high resolution NMR spectroscopy.

  3. Gap-junction-mediated communication in human periodontal ligament cells.

    Science.gov (United States)

    Kato, R; Ishihara, Y; Kawanabe, N; Sumiyoshi, K; Yoshikawa, Y; Nakamura, M; Imai, Y; Yanagita, T; Fukushima, H; Kamioka, H; Takano-Yamamoto, T; Yamashiro, T

    2013-07-01

    Periodontal tissue homeostasis depends on a complex cellular network that conveys cell-cell communication. Gap junctions (GJs), one of the intercellular communication systems, are found between adjacent human periodontal ligament (hPDL) cells; however, the functional GJ coupling between hPDL cells has not yet been elucidated. In this study, we investigated functional gap-junction-mediated intercellular communication in isolated primary hPDL cells. SEM images indicated that the cells were in contact with each other via dendritic processes, and also showed high anti-connexin43 (Cx43) immunoreactivity on these processes. Gap-junctional intercellular communication (GJIC) among hPDL cells was assessed by fluorescence recovery after a photobleaching (FRAP) analysis, which exhibited dye coupling between hPDL cells, and was remarkably down-regulated when the cells were treated with a GJ blocker. Additionally, we examined GJs under hypoxic stress. The fluorescence recovery and expression levels of Cx43 decreased time-dependently under the hypoxic condition. Exposure to GJ inhibitor or hypoxia increased RANKL expression, and decreased OPG expression. This study shows that GJIC is responsible for hPDL cells and that its activity is reduced under hypoxia. This is consistent with the possible role of hPDL cells in regulating the biochemical reactions in response to changes in the hypoxic environment.

  4. Bcl-2 overexpression: effects on transmembrane calcium movement

    International Nuclear Information System (INIS)

    Rangaswami, Arun A.; Premack, Brett; Walleczek, Jan; Killoran, Pamela; Gardner, Phyllis; Knox, Susan J.

    1996-01-01

    Purpose/Objective: High levels of expression of the proto-oncogene bcl-2 and its 26 kD protein product Bcl-2 have been correlated with the inhibition of apoptosis and the increased resistance of tumor cells to cytotoxic drugs and ionizing radiation. Unfortunately, the specific mechanism of action of Bcl-2 remains poorly understood. In the studies described here, the role of intracellular calcium fluxes and plasma membrane calcium cycling in the induction of apoptosis, and the effect of Bcl-2 expression on the modulation of transmembrane calcium fluxes following treatment of cells with cytotoxic agents were studied. The relationship between intracellular calcium release, capacitive calcium entry, and the plasma membrane potential were also investigated. Materials and Methods: Human B-cell lymphoma (PW) and human promyelocytic leukemia (HL60) cell lines were transfected with Bcl-2 and a control vector. The Bcl-2 transfectants over expressed the Bcl-2 onco-protein and were more resistant to irradiation than the control cells. Cells were loaded with fluorescent indicators indo-1 and fura-2 AM to quantify the cytosolic calcium concentration and subsequent calcium responses to a variety of cytotoxic stimuli, including the microsomal ATPase inhibitor, thapsigargin, using fluorometric measurements. Comparisons of resting and stimulated cytosolic calcium concentrations were made between the parental, neomycin control, and bcl-2 transfected cells. In order to determine the actual calcium influx rate, cells were loaded with either indo-1 or fura-2 and then exposed to 0.1 mM extracellular manganese, which enters the cells through calcium influx channels and quenches the fluorescent signal in proportion to the calcium influx rate. In order to determine the role of the membrane potential in driving calcium influx, cells were treated with either 0.1 μM Valinomycin or isotonic potassium chloride to either hyper polarize or depolarize the resting membrane potential, and the

  5. Peptide microarray analysis of substrate specificity of the transmembrane Ser/Thr kinase KPI-2 reveals reactivity with cystic fibrosis transmembrane conductance regulator and phosphorylase.

    Science.gov (United States)

    Wang, Hong; Brautigan, David L

    2006-11-01

    Human lemur (Lmr) kinases are predicted to be Tyr kinases based on sequences and are related to neurotrophin receptor Trk kinases. This study used homogeneous recombinant KPI-2 (Lmr2, LMTK2, Cprk, brain-enriched protein kinase) kinase domain and a library of 1,154 peptides on a microarray to analyze substrate specificity. We found that KPI-2 is strictly a Ser/Thr kinase that reacts with Ser either preceded by or followed by Pro residues but unlike other Pro-directed kinases does not strictly require an adjacent Pro residue. The most reactive peptide in the library corresponds to Ser-737 of cystic fibrosis transmembrane conductance regulator, and the recombinant R domain of cystic fibrosis transmembrane conductance regulator was a preferred substrate. Furthermore the KPI-2 kinase phosphorylated peptides corresponding to the single site in phosphorylase and purified phosphorylase b, making this only the second known phosphorylase b kinase. Phosphorylase was used as a specific substrate to show that KPI-2 is inhibited in living cells by addition of nerve growth factor or serum. The results demonstrate the utility of the peptide library to probe specificity and discover kinase substrates and offer a specific assay that reveals hormonal regulation of the activity of this unusual transmembrane kinase.

  6. Advantages of combined transmembrane topology and signal peptide prediction--the Phobius web server

    DEFF Research Database (Denmark)

    Käll, Lukas; Krogh, Anders; Sonnhammer, Erik L L

    2007-01-01

    . The method makes an optimal choice between transmembrane segments and signal peptides, and also allows constrained and homology-enriched predictions. We here present a web interface (http://phobius.cgb.ki.se and http://phobius.binf.ku.dk) to access Phobius. Udgivelsesdato: 2007-Jul......When using conventional transmembrane topology and signal peptide predictors, such as TMHMM and SignalP, there is a substantial overlap between these two types of predictions. Applying these methods to five complete proteomes, we found that 30-65% of all predicted signal peptides and 25-35% of all...

  7. A human/mouse chimeric monoclonal antibody against intercellular adhesion molecule-1 for tumor radioimmunoimaging

    International Nuclear Information System (INIS)

    Yamamura, Miyuki; Hinoda, Yuji; Sasaki, Shigeru; Tsujisaki, Masayuki; Imai, Kohzoh; Oriuchi, Noboru; Endo, Keigo.

    1996-01-01

    A mouse-human chimeric antibody for intercellular adhesion molecule-1 (ICAM-1) was established by using heavy chain loss mouse mutant hybridoma and human immunoglobulin expression vector. The HA58 hybridoma secreted anti-ICAM-1 monoclonal antibody (MoAb) (IgG1,κ). The gene of the mouse variable region of heavy chain was amplified and cloned by the polymerase chain reaction technique directly from the HA58 hybridoma RNA. The variable region of heavy chain was joined with an expression vector which contains human γ1 constant gene. The expression vector was transfected into heavy chain loss mutant cells HA58-7, which produced only murine immunoglobulin light chains. The resultant chimeric MoAb HA58, chHA58, retained full-binding reactivity to ICAM-1 compared with murine HA58 parental antibody. The chimeric MoAb chHA58 showed little antibody dependent cell-mediated cytotoxic activity against cultured tumor cells. Biodistribution studies with 99m Tc-labeled chHA58 in nude mice bearing human gastric carcinoma JRST cells, demonstrated that the tumor-blood ratio was 1.55 at 18 h after injection, when the tumors were clearly visible in gamma scintigraphy. These data suggest that chHA58 may be of practical use for radioimmunoimaging of a wide variety of tumors. (author)

  8. Proteomic insights into intra- and intercellular plant-bacteria symbiotic association during root nodule formation

    Directory of Open Access Journals (Sweden)

    Afshin eSalavati

    2013-02-01

    Full Text Available Over the last several decades, there have been a large number of studies done on the all aspects of legumes and bacteria which participate in nitrogen-fixing symbiosis. The analysis of legume-bacteria interaction is not just a matter of numerical complexity in terms of variants of gene products that can arise from a single gene. Bacteria regulate their quorum-sensing genes to enhance their ability to induce conjugation of plasmids and symbiotic islands, and various protein secretion mechanisms; that can stimulate a collection of chain reactions including species-specific combinations of plant-secretion isoflavonoids, complicated calcium signaling pathways and autoregulation of nodulation mechanisms. Quorum-sensing systems are introduced by the intra- and intercellular organization of gene products lead to protein–protein interactions or targeting of proteins to specific cellular structures. In this study, an attempt has been made to review significant contributions related to nodule formation and development and their impacts on cell proteome for better understanding of plant-bacterium interaction mechanism at protein level. This review would not only provide new insights into the plant-bacteria symbiosis response mechanisms but would also highlights the importance of studying changes in protein abundance inside and outside of cells in response to symbiosis. Furthermore, the application to agriculture programe of plant-bacteria interaction will be discussed.

  9. Herpes simplex virus glycoprotein D relocates nectin-1 from intercellular contacts.

    Science.gov (United States)

    Bhargava, Arjun K; Rothlauf, Paul W; Krummenacher, Claude

    2016-12-01

    Herpes simplex virus (HSV) uses the cell adhesion molecule nectin-1 as a receptor to enter neurons and epithelial cells. The viral glycoprotein D (gD) is used as a non-canonical ligand for nectin-1. The gD binding site on nectin-1 overlaps with a functional adhesive site involved in nectin-nectin homophilic trans-interaction. Consequently, when nectin-1 is engaged with a cellular ligand at cell junctions, the gD binding site is occupied. Here we report that HSV gD is able to disrupt intercellular homophilic trans-interaction of nectin-1 and induce a rapid redistribution of nectin-1 from cell junctions. This movement does not require the receptor's interaction with the actin-binding adaptor afadin. Interaction of nectin-1 with afadin is also dispensable for virion surfing along nectin-1-rich filopodia. Cells seeded on gD-coated surfaces also fail to accumulate nectin-1 at cell contact. These data indicate that HSV gD affects nectin-1 locally through direct interaction and more globally through signaling. Copyright © 2016 Elsevier Inc. All rights reserved.

  10. Association of Intercellular Adhesion Molecule 1 (ICAM1 with Diabetes and Diabetic Nephropathy

    Directory of Open Access Journals (Sweden)

    Harvest F Gu

    2013-01-01

    Full Text Available Diabetes and diabetic nephropathy are complex diseases affected by genetic and environmental factors. Identification of the susceptibility genes and investigation of their roles may provide useful information for better understanding of the pathogenesis and for developing novel therapeutic approaches. Intercellular adhesion molecule 1 (ICAM1 is a cell surface glycoprotein expressed on endothelial cells and leukocytes in the immune system. The ICAM1 gene is located on chromosome 19p13 within the linkage region of diabetes. In the recent years, accumulating reports have implicated that genetic polymorphisms in the ICAM1 gene are associated with diabetes and diabetic nephropathy. Serum ICAM1 levels in diabetes patients and the icam1 gene expression in kidney tissues of diabetic animals are increased compared to the controls. Therefore, ICAM1 may play a role in the development of diabetes and diabetic nephropathy. In this review, we present genomic structure, variation and regulation of the ICAM1 gene, summarized genetic and biological studies of this gene in diabetes and diabetic nephropathy and discussed about the potential application using ICAM1 as a biomarker and target for prediction and treatment of diabetes and diabetic nephropathy.

  11. Effect of intercellular adhesion molecule 1 expression in radiation otitis media murine model

    International Nuclear Information System (INIS)

    Wang Shengzi; Cheng Qingfang; Lu Shenbin; Liu Jianping; Wang Shuyi

    2003-01-01

    Objective: To characterize the dose- and time-dependent changes in intercellular adhesion molecule 1 (ICAM-1) expression and the role of this molecule as a mediator of middle ear inflammation induced by radiation. Methods: Radiation-induced otitis media animal models were established by using guinea pigs after 60 Co irradiation with 3 Gy/fraction per day, 5 times per week to a total dose of 15, 30, 45 Gy. The expression of ICAM-1 was studied by SP immunohistochemistry with the relation between radiation dose and infiltration of leukocytes investigated. Results: ICAM-1 was not expressed in the normal epithelium of the middle ear mucosa. Mucosal epithelium strongly expressed ICAM-1 after having been administered with 45 Gy of irradiation showing a significant correlation between the expression of ICAM-1 and the infiltration of leukocytes. Conclusions: Irradiation increases the expression of ICAM-1 in the middle ear mucosa. ICAM-1 may be related to the inflammation in the middle ear after irradiation

  12. Mechanical Entrapment Is Insufficient and Intercellular Adhesion Is Essential for Metastatic Cell Arrest in Distant Organs

    Directory of Open Access Journals (Sweden)

    Olga V. Glinskii

    2005-05-01

    Full Text Available In this report, we challenge a common perception that tumor embolism is a size-limited event of mechanical arrest, occurring in the first capillary bed encountered by blood-borne metastatic cells. We tested the hypothesis that mechanical entrapment alone, in the absence of tumor cell adhesion to blood vessel walls, is not sufficient for metastatic cell arrest in target organ microvasculature. The in vivo metastatic deposit formation assay was used to assess the number and location of fluorescently labeled tumor cells lodged in selected organs and tissues following intravenous inoculation. We report that a significant fraction of breast and prostate cancer cells escapes arrest in a lung capillary bed and lodges successfully in other organs and tissues. Monoclonal antibodies and carbohydrate-based compounds (anti-Thomsen-Friedenreich antigen antibody, anti-galectin-3 antibody, modified citrus pectin, and lactulosyl-L-leucine, targeting specifically β-galactoside-mediated tumor-endothelial cell adhesive interactions, inhibited by >90% the in vivo formation of breast and prostate carcinoma metastatic deposits in mouse lung and bones. Our results indicate that metastatic cell arrest in target organ microvessels is not a consequence of mechanical trapping, but is supported predominantly by intercellular adhesive interactions mediated by cancer-associated Thomsen-Friedenreich glycoantigen and β-galactoside-binding lectin galectin-3. Efficient blocking of β-galactoside-mediated adhesion precludes malignant cell lodging in target organs.

  13. Effects of lead intoxication on intercellular junctions and biochemical alterations of the renal proximal tubule cells.

    Science.gov (United States)

    Navarro-Moreno, L G; Quintanar-Escorza, M A; González, S; Mondragón, R; Cerbón-Solorzáno, J; Valdés, J; Calderón-Salinas, J V

    2009-10-01

    Lead intoxication is a worldwide health problem which frequently affects the kidney. In this work, we studied the effects of chronic lead intoxication (500 ppm of Pb in drinking water during seven months) on the structure, function and biochemical properties of rat proximal tubule cells. Lead-exposed animals showed increased lead concentration in kidney, reduction of calcium and amino acids uptake, oxidative damage and glucosuria, proteinuria, hematuria and reduced urinary pH. These biochemical and physiological alterations were related to striking morphological modifications in the structure of tubule epithelial cells and in the morphology of their mitochondria, nuclei, lysosomes, basal and apical membranes. Interestingly, in addition to the nuclei, inclusion bodies were found in the cytoplasm and in mitochondria. The epithelial cell structure modifications included an early loss of the apical microvillae, followed by a decrement of the luminal space and the respective apposition and proximity of apical membranes, resulting in the formation of atypical intercellular contacts and adhesion structures. Similar but less marked alterations were observed in subacute lead intoxication as well. Our work contributes in the understanding of the physiopathology of lead intoxication on the structure of renal tubular epithelial cell-cell contacts in vivo.

  14. Role of Intercellular Adhesion Molecule-1 in Radiation-Induced Brain Injury

    International Nuclear Information System (INIS)

    Wu, K.-L.; Tu Ba; Li Yuqing; Wong, C. Shun

    2010-01-01

    Purpose: To determine the role of intercellular adhesion molecule-1 (ICAM-1) in the pathogenesis of brain injury after irradiation (IR). Methods and Materials: We assessed the expression of ICAM-1 in mouse brain after cranial IR and determined the histopathologic and behavioral changes in mice that were either wildtype (+/+) or knockout (-/-) of the ICAM-1 gene after IR. Results: There was an early dose-dependent increase in ICAM-1 mRNA and protein expression after IR. Increased ICAM-1 immunoreactivity was observed in endothelia and glia of ICAM-1+/+ mice up to 8 months after IR. ICAM-1-/- mice showed no expression. ICAM-1+/+ and ICAM-1-/- mice showed similar vascular abnormalities at 2 months after 10-17 Gy, and there was evidence for demyelination and inhibition of hippocampal neurogenesis at 8 months after 10 Gy. After 10 Gy, irradiated ICAM-1+/+ and ICAM-1-/- mice showed similar behavioral changes at 2-6 months in open field, light-dark chamber, and T-maze compared with age-matched genotype controls. Conclusion: There is early and late upregulation of ICAM-1 in the vasculature and glia of mouse brain after IR. ICAM-1, however, does not have a causative role in the histopathologic injury and behavioral dysfunction after moderate single doses of cranial IR.

  15. Rapid construction of mechanically- confined multi- cellular structures using dendrimeric intercellular linker.

    Science.gov (United States)

    Mo, Xuejun; Li, Qiushi; Yi Lui, Lena Wai; Zheng, Baixue; Kang, Chiang Huen; Nugraha, Bramasta; Yue, Zhilian; Jia, Rui Rui; Fu, Hong Xia; Choudhury, Deepak; Arooz, Talha; Yan, Jie; Lim, Chwee Teck; Shen, Shali; Hong Tan, Choon; Yu, Hanry

    2010-10-01

    Tissue constructs that mimic the in vivo cell-cell and cell-matrix interactions are especially useful for applications involving the cell- dense and matrix- poor internal organs. Rapid and precise arrangement of cells into functional tissue constructs remains a challenge in tissue engineering. We demonstrate rapid assembly of C3A cells into multi- cell structures using a dendrimeric intercellular linker. The linker is composed of oleyl- polyethylene glycol (PEG) derivatives conjugated to a 16 arms- polypropylenimine hexadecaamine (DAB) dendrimer. The positively charged multivalent dendrimer concentrates the linker onto the negatively charged cell surface to facilitate efficient insertion of the hydrophobic oleyl groups into the cellular membrane. Bringing linker- treated cells into close proximity to each other via mechanical means such as centrifugation and micromanipulation enables their rapid assembly into multi- cellular structures within minutes. The cells exhibit high levels of viability, proliferation, three- dimensional (3D) cell morphology and other functions in the constructs. We constructed defined multi- cellular structures such as rings, sheets or branching rods that can serve as potential tissue building blocks to be further assembled into complex 3D tissue constructs for biomedical applications. 2010 Elsevier Ltd. All rights reserved.

  16. Loss of intercellular adhesion leads to differential accumulation of hypericin in bladder cancer

    Science.gov (United States)

    Lucky, S. Sasidharan; Bhuvaneswari, Ramaswamy; Chin, William W. L.; Lau, Weber K. O.; Olivo, Malini C. D.

    2009-06-01

    Photodynamic diagnosis (PDD) exploits the photoactive nature of certain compounds, namely photosensitizers, in order to enhance the visual demarcation between normal and neoplastic tissue. Hypericin is one such potent photosensitizer that preferentially accumulate in neoplastic tissue, and fluoresce in the visible spectrum when illuminated with light of an appropriate wavelength. In our study, we investigated the role of E-cadherin in the selective permeation of hypericin in bladder cancer tissues. Clinical studies were done on a series of 43 histologically graded bladder cancer biopsy specimens, obtained from 28 patients who received intravesical instillations with 8μM hypericin solution for at least 2 hours. Immunohistochemical staining was used to assess the expression of E-cadherin, in the cryosectioned tissues. Hypericin uptake was examined by fluorescence microscopy. Immunohistochemical staining showed a clear expression of E-cadherin along the urothelial lining of the normal and pre-malignant tissues. Partial expression of these cell adhesion molecules were still observed in malignant tissues, however there was a loss of expression to variable extends along the urothelium. Thus, loss of intercellular adhesion can be associated with enhanced hypericin permeation through paracellular diffusion.

  17. Intercellular adhesion molecule-1 augments myoblast adhesion and fusion through homophilic trans-interactions.

    Science.gov (United States)

    Pizza, Francis X; Martin, Ryan A; Springer, Evan M; Leffler, Maxwell S; Woelmer, Bryce R; Recker, Isaac J; Leaman, Douglas W

    2017-07-11

    The overall objective of the study was to identify mechanisms through which intercellular adhesion molecule-1 (ICAM-1) augments the adhesive and fusogenic properties of myogenic cells. Hypotheses were tested using cultured myoblasts and fibroblasts, which do not constitutively express ICAM-1, and myoblasts and fibroblasts forced to express full length ICAM-1 or a truncated form lacking the cytoplasmic domain of ICAM-1. ICAM-1 mediated myoblast adhesion and fusion were quantified using novel assays and cell mixing experiments. We report that ICAM-1 augments myoblast adhesion to myoblasts and myotubes through homophilic trans-interactions. Such adhesive interactions enhanced levels of active Rac in adherent and fusing myoblasts, as well as triggered lamellipodia, spreading, and fusion of myoblasts through the signaling function of the cytoplasmic domain of ICAM-1. Rac inhibition negated ICAM-1 mediated lamellipodia, spreading, and fusion of myoblasts. The fusogenic property of ICAM-1-ICAM-1 interactions was restricted to myogenic cells, as forced expression of ICAM-1 by fibroblasts did not augment their fusion to ICAM-1+ myoblasts/myotubes. We conclude that ICAM-1 augments myoblast adhesion and fusion through its ability to self-associate and initiate Rac-mediated remodeling of the actin cytoskeleton.

  18. Intercellular K⁺ accumulation depolarizes Type I vestibular hair cells and their associated afferent nerve calyx.

    Science.gov (United States)

    Contini, D; Zampini, V; Tavazzani, E; Magistretti, J; Russo, G; Prigioni, I; Masetto, S

    2012-12-27

    Mammalian vestibular organs contain two types of sensory receptors, named Type I and Type II hair cells. While Type II hair cells are contacted by several small afferent nerve terminals, the basolateral surface of Type I hair cells is almost entirely enveloped by a single large afferent nerve terminal, called calyx. Moreover Type I, but not Type II hair cells, express a low-voltage-activated outward K(+) current, I(K,L), which is responsible for their much lower input resistance (Rm) at rest as compared to Type II hair cells. The functional meaning of I(K,L) and associated calyx is still enigmatic. By combining the patch-clamp whole-cell technique with the mouse whole crista preparation, we have recorded the current- and voltage responses of in situ hair cells. Outward K(+) current activation resulted in K(+) accumulation around Type I hair cells, since it induced a rightward shift of the K(+) reversal potential the magnitude of which depended on the amplitude and duration of K(+) current flow. Since this phenomenon was never observed for Type II hair cells, we ascribed it to the presence of a residual calyx limiting K(+) efflux from the synaptic cleft. Intercellular K(+) accumulation added a slow (τ>100ms) depolarizing component to the cell voltage response. In a few cases we were able to record from the calyx and found evidence for intercellular K(+) accumulation as well. The resulting depolarization could trigger a discharge of action potentials in the afferent nerve fiber. Present results support a model where pre- and postsynaptic depolarization produced by intercellular K(+) accumulation cooperates with neurotransmitter exocytosis in sustaining afferent transmission arising from Type I hair cells. While vesicular transmission together with the low Rm of Type I hair cells appears best suited for signaling fast head movements, depolarization produced by intercellular K(+) accumulation could enhance signal transmission during slow head movements. Copyright

  19. Destruction of the hepatocyte junction by intercellular invasion of Leptospira causes jaundice in a hamster model of Weil's disease.

    Science.gov (United States)

    Miyahara, Satoshi; Saito, Mitsumasa; Kanemaru, Takaaki; Villanueva, Sharon Y A M; Gloriani, Nina G; Yoshida, Shin-ichi

    2014-08-01

    Weil's disease, the most severe form of leptospirosis, is characterized by jaundice, haemorrhage and renal failure. The mechanisms of jaundice caused by pathogenic Leptospira remain unclear. We therefore aimed to elucidate the mechanisms by integrating histopathological changes with serum biochemical abnormalities during the development of jaundice in a hamster model of Weil's disease. In this work, we obtained three-dimensional images of infected hamster livers using scanning electron microscope together with freeze-cracking and cross-cutting methods for sample preparation. The images displayed the corkscrew-shaped bacteria, which infiltrated the Disse's space, migrated between hepatocytes, detached the intercellular junctions and disrupted the bile canaliculi. Destruction of bile canaliculi coincided with the elevation of conjugated bilirubin, aspartate transaminase and alkaline phosphatase levels in serum, whereas serum alanine transaminase and γ-glutamyl transpeptidase levels increased slightly, but not significantly. We also found in ex vivo experiments that pathogenic, but not non-pathogenic leptospires, tend to adhere to the perijunctional region of hepatocyte couplets isolated from hamsters and initiate invasion of the intercellular junction within 1 h after co-incubation. Our results suggest that pathogenic leptospires invade the intercellular junctions of host hepatocytes, and this invasion contributes in the disruption of the junction. Subsequently, bile leaks from bile canaliculi and jaundice occurs immediately. Our findings revealed not only a novel pathogenicity of leptospires, but also a novel mechanism of jaundice induced by bacterial infection. © 2014 The Authors. International Journal of Experimental Pathology © 2014 International Journal of Experimental Pathology.

  20. Robust synchronization analysis in nonlinear stochastic cellular networks with time-varying delays, intracellular perturbations and intercellular noise.

    Science.gov (United States)

    Chen, Po-Wei; Chen, Bor-Sen

    2011-08-01

    Naturally, a cellular network consisted of a large amount of interacting cells is complex. These cells have to be synchronized in order to emerge their phenomena for some biological purposes. However, the inherently stochastic intra and intercellular interactions are noisy and delayed from biochemical processes. In this study, a robust synchronization scheme is proposed for a nonlinear stochastic time-delay coupled cellular network (TdCCN) in spite of the time-varying process delay and intracellular parameter perturbations. Furthermore, a nonlinear stochastic noise filtering ability is also investigated for this synchronized TdCCN against stochastic intercellular and environmental disturbances. Since it is very difficult to solve a robust synchronization problem with the Hamilton-Jacobi inequality (HJI) matrix, a linear matrix inequality (LMI) is employed to solve this problem via the help of a global linearization method. Through this robust synchronization analysis, we can gain a more systemic insight into not only the robust synchronizability but also the noise filtering ability of TdCCN under time-varying process delays, intracellular perturbations and intercellular disturbances. The measures of robustness and noise filtering ability of a synchronized TdCCN have potential application to the designs of neuron transmitters, on-time mass production of biochemical molecules, and synthetic biology. Finally, a benchmark of robust synchronization design in Escherichia coli repressilators is given to confirm the effectiveness of the proposed methods. Copyright © 2011 Elsevier Inc. All rights reserved.

  1. Cloning and characterization of SCART1, a novel scavenger receptor cysteine-rich type I transmembrane molecule

    DEFF Research Database (Denmark)

    Holm, Dorte; Fink, Dorte Rosenbek; Grønlund, Jørn

    2009-01-01

    We have cloned and characterized a novel murine transmembrane molecule, mSCART1 belonging to the scavenger receptor cysteine-rich superfamily. The cDNA encodes a polypeptide chain of 989 amino acids, organized as a type I transmembrane protein that contains eight extracellular SRCR domains followed...

  2. Structural Insights into Triglyceride Storage Mediated by Fat Storage-Inducing Transmembrane (FIT) Protein 2

    Science.gov (United States)

    Gross, David A.; Snapp, Erik L.; Silver, David L.

    2010-01-01

    Fat storage-Inducing Transmembrane proteins 1 & 2 (FIT1/FITM1 and FIT2/FITM2) belong to a unique family of evolutionarily conserved proteins localized to the endoplasmic reticulum that are involved in triglyceride lipid droplet formation. FIT proteins have been shown to mediate the partitioning of cellular triglyceride into lipid droplets, but not triglyceride biosynthesis. FIT proteins do not share primary sequence homology with known proteins and no structural information is available to inform on the mechanism by which FIT proteins function. Here, we present the experimentally-solved topological models for FIT1 and FIT2 using N-glycosylation site mapping and indirect immunofluorescence techniques. These methods indicate that both proteins have six-transmembrane-domains with both N- and C-termini localized to the cytosol. Utilizing this model for structure-function analysis, we identified and characterized a gain-of-function mutant of FIT2 (FLL(157-9)AAA) in transmembrane domain 4 that markedly augmented the total number and mean size of lipid droplets. Using limited-trypsin proteolysis we determined that the FLL(157-9)AAA mutant has enhanced trypsin cleavage at K86 relative to wild-type FIT2, indicating a conformational change. Taken together, these studies indicate that FIT2 is a 6 transmembrane domain-containing protein whose conformation likely regulates its activity in mediating lipid droplet formation. PMID:20520733

  3. Activation gating kinetics of GIRK channels are mediated by cytoplasmic residues adjacent to transmembrane domains.

    Science.gov (United States)

    Sadja, Rona; Reuveny, Eitan

    2009-01-01

    G-protein-coupled inwardly rectifying potassium channels (GIRK/Kir3.x) are involved in neurotransmission-mediated reduction of excitability. The gating mechanism following G protein activation of these channels likely proceeds from movement of inner transmembrane helices to allow K(+) ions movement through the pore of the channel. There is limited understanding of how the binding of G-protein betagamma subunits to cytoplasmic regions of the channel transduces the signal to the transmembrane regions. In this study, we examined the molecular basis that governs the activation kinetics of these channels, using a chimeric approach. We identified two regions as being important in determining the kinetics of activation. One region is the bottom of the outer transmembrane helix (TM1) and the cytoplasmic domain immediately adjacent (the slide helix); and the second region is the bottom of the inner transmembrane helix (TM2) and the cytoplasmic domain immediately adjacent. Interestingly, both of these regions are sufficient in mediating the kinetics of fast activation gating. This result suggests that there is a cooperative movement of either one of these domains to allow fast and efficient activation gating of GIRK channels.

  4. Structural insights into triglyceride storage mediated by fat storage-inducing transmembrane (FIT protein 2.

    Directory of Open Access Journals (Sweden)

    David A Gross

    2010-05-01

    Full Text Available Fat storage-Inducing Transmembrane proteins 1 & 2 (FIT1/FITM1 and FIT2/FITM2 belong to a unique family of evolutionarily conserved proteins localized to the endoplasmic reticulum that are involved in triglyceride lipid droplet formation. FIT proteins have been shown to mediate the partitioning of cellular triglyceride into lipid droplets, but not triglyceride biosynthesis. FIT proteins do not share primary sequence homology with known proteins and no structural information is available to inform on the mechanism by which FIT proteins function. Here, we present the experimentally-solved topological models for FIT1 and FIT2 using N-glycosylation site mapping and indirect immunofluorescence techniques. These methods indicate that both proteins have six-transmembrane-domains with both N- and C-termini localized to the cytosol. Utilizing this model for structure-function analysis, we identified and characterized a gain-of-function mutant of FIT2 (FLL(157-9AAA in transmembrane domain 4 that markedly augmented the total number and mean size of lipid droplets. Using limited-trypsin proteolysis we determined that the FLL(157-9AAA mutant has enhanced trypsin cleavage at K86 relative to wild-type FIT2, indicating a conformational change. Taken together, these studies indicate that FIT2 is a 6 transmembrane domain-containing protein whose conformation likely regulates its activity in mediating lipid droplet formation.

  5. Molecular dynamics study of the solvation of an alpha-helical transmembrane peptide by DMSO

    NARCIS (Netherlands)

    Duarte, A.M.; Mierlo, van C.P.M.; Hemminga, M.A.

    2008-01-01

    10-ns molecular dynamics study of the solvation of a hydrophobic transmembrane helical peptide in dimethyl sulfoxide (DMSO) is presented. The objective is to analyze how this aprotic polar solvent is able to solvate three groups of amino acid residues (i.e., polar, apolar, and charged) that are

  6. The transmembrane region is responsible for targeting of adaptor protein LAX into "heavy rafts''

    Czech Academy of Sciences Publication Activity Database

    Hrdinka, Matouš; Otáhal, Pavel; Hořejší, Václav

    2012-01-01

    Roč. 7, č. 5 (2012), e36330 E-ISSN 1932-6203 R&D Projects: GA ČR GEMEM/09/E011; GA MŠk 1M0506 Institutional research plan: CEZ:AV0Z50520514 Keywords : LAX * transmembrane domain * DRM Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.730, year: 2012

  7. Trans-membrane area asymmetry controls the shape of cellular organelles

    NARCIS (Netherlands)

    Beznoussenko, Galina V; Pilyugin, Sergei S; Geerts, Willie J C; Kozlov, Michael M; Burger, Koert N J; Luini, Alberto; Derganc, Jure; Mironov, Alexander A

    2015-01-01

    Membrane organelles often have complicated shapes and differ in their volume, surface area and membrane curvature. The ratio between the surface area of the cytosolic and luminal leaflets (trans-membrane area asymmetry (TAA)) determines the membrane curvature within different sites of the organelle.

  8. Intact transmembrane isoforms of the neural cell adhesion molecule are released from the plasma membrane

    DEFF Research Database (Denmark)

    Olsen, M; Krog, L; Edvardsen, K

    1993-01-01

    . By density-gradient centrifugation it was shown that shed transmembrane NCAM-B was present in fractions of high, as well as low, density, indicating that a fraction of the shed NCAM is associated with minor plasma membrane fragments. Finally, it was shown that isolated soluble NCAM inhibited cell binding...

  9. Transmembrane adaptor molecules: a new category of lymphoid-cell markers

    Czech Academy of Sciences Publication Activity Database

    Tedoldi, S.; Paterson, J.C.; Hansmann, M.-L.; Natkunam, Y.; Rüdiger, T.; Angelisová, Pavla; Du, M.Q.; Roberton, H.; Roncador, G.; Sanchez, L.; Pozzobon, M.; Masir, N.; Barry, R.; Pileri, S.; Mason, D.Y.; Marafioti, T.; Hořejší, Václav

    2006-01-01

    Roč. 107, č. 1 (2006), s. 213-221 ISSN 0006-4971 R&D Projects: GA MŠk(CZ) 1M0506 Institutional research plan: CEZ:AV0Z50520514 Keywords : transmembrane adaptors * PAG * LIME Subject RIV: EC - Immunology Impact factor: 10.370, year: 2006

  10. Transmembrane Domain Single-Nucleotide Polymorphisms Impair Expression and Transport Activity of ABC Transporter ABCG2

    NARCIS (Netherlands)

    Sjostedt, N.; Heuvel, J.J.M.W. van den; Koenderink, J.B.; Kidron, H.

    2017-01-01

    PURPOSE: To study the function and expression of nine naturally occurring single-nucleotide polymorphisms (G406R, F431L, S441N, P480L, F489L, M515R, L525R, A528T and T542A) that are predicted to reside in the transmembrane regions of the ABC transporter ABCG2. METHODS: The transport activity of the

  11. NTAL (non-T cell activation linker):a transmembrane adaptor protein involved in immunoreceptor signaling

    Czech Academy of Sciences Publication Activity Database

    Brdička, Tomáš; Imrich, Martin; Angelisová, Pavla; Brdičková, Naděžda; Horváth, Ondřej; Špička, Jiří; Hilgert, Ivan; Lusková, Petra; Dráber, Petr; Novák, P.; Engels, N.; Wienands, J.; Simeoni, L.; Osterreicher, J.; Aguado, E.; Malissen, M.; Schraven, B.; Hořejší, Václav

    2002-01-01

    Roč. 196, č. 12 (2002), s. 16180-16185 ISSN 0022-1007 R&D Projects: GA MŠk LN00A026 Keywords : NTAL * transmembrane adaptor * immunoreceptor signaling Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 15.838, year: 2002

  12. Ligand-mediated negative regulation of a chimeric transmembrane receptor tyrosine phosphatase

    DEFF Research Database (Denmark)

    Desai, D M; Sap, J; Schlessinger, J

    1993-01-01

    CD45, a transmembrane protein tyrosine phosphatase (PTPase), is required for TCR signaling. Multiple CD45 isoforms, differing in the extracellular domain, are expressed in a tissue- and activation-specific manner, suggesting an important function for this domain. We report that a chimeric protein...... that ligand-mediated regulation of receptor-PTPases may have mechanistic similarities with receptor tyrosine kinases....

  13. SCIMP, a transmembrane adaptor protein involved in major histocompatibility complex class II signaling

    Czech Academy of Sciences Publication Activity Database

    Dráber, Peter; Vonková, Ivana; Štěpánek, Ondřej; Hrdinka, Matouš; Kucová, Markéta; Skopcová, Tereza; Otáhal, Pavel; Angelisová, Pavla; Hořejší, Václav; Yeung, M.; Weiss, A.; Brdička, Tomáš

    2011-01-01

    Roč. 31, č. 22 (2011), s. 4550-4562 ISSN 0270-7306 R&D Projects: GA MŠk 1M0506; GA ČR GEMEM/09/E011 Institutional research plan: CEZ:AV0Z50520514 Keywords : SCIMP * transmembrane adaptor protein * MHC II Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 5.527, year: 2011

  14. Molecular pharmacological phenotyping of EBI2. An orphan seven-transmembrane receptor with constitutive activity

    DEFF Research Database (Denmark)

    Rosenkilde, Mette M; Benned-Jensen, Tau; Holst, Peter J

    2006-01-01

    Epstein-Barr virus (EBV)-induced receptor 2 (EBI2) is an orphan seven-transmembrane (7TM) receptor originally identified as the most up-regulated gene (>200-fold) in EBV-infected cells. Here we show that EBI2 signals with constitutive activity through Galpha(i) as determined by a receptor...

  15. Modeling the Structure of SARS 3a Transmembrane Protein Using a ...

    Indian Academy of Sciences (India)

    Modeling the structure of SARS 3a Transmembrane protein using a ... for the implicit membrane molecular dynamics (MD) simulations. ... The coordinates during the simulation were saved every 500 steps, and were used for analysis. ... the pair list for calculation of nonbonded interactions being updated after every 10 steps.

  16. Simulations of Skin Barrier Function: Free Energies of Hydrophobic and Hydrophilic Transmembrane Pores in Ceramide Bilayers

    NARCIS (Netherlands)

    Notman, Rebecca; Anwar, Jamshed; Briels, Willem J.; Noro, Massimo G.; den Otter, Wouter K.

    2008-01-01

    Transmembrane pore formation is central to many biological processes such as ion transport, cell fusion, and viral infection. Furthermore, pore formation in the ceramide bilayers of the stratum corneum may be an important mechanism by which penetration enhancers such as dimethylsulfoxide (DMSO)

  17. Large-scale identification of membrane proteins based on analysis of trypsin-protected transmembrane segments

    Czech Academy of Sciences Publication Activity Database

    Vít, O.; Man, Petr; Kádek, Alan; Hausner, Jiří; Sklenář, A.; Harant, K.; Novák, Petr; Scigelová, M.; Wofferndin, G.; Petrák, J.

    2016-01-01

    Roč. 146, SI (2016), s. 15-22 ISSN 1874-3919 R&D Projects: GA MŠk(CZ) ED1.1.00/02.0109 Institutional support: RVO:61388971 Keywords : Integral membrane proteins * CNBr * Transmembrane Subject RIV: CE - Biochemistry Impact factor: 3.914, year: 2016

  18. Communication without communication

    Directory of Open Access Journals (Sweden)

    Bratina Boris R.

    2017-01-01

    Full Text Available This paper analyses the necessary conditions for successful communication. It is well known that post-modernity, described as an era of control, produces only decentralized, imploded subjectivities, who are neither able to question their own being nor to relate one with another in authentic bonds of communication. Today, virtual communication has become an ultimate model of every possible communication whatsoever. The authors, therefore, pose the question of conditions for possibility of subjectivities who would be able and apt for authentic communication, wherein faith, fidelity, truth, and capability of keeping one's word occupy the central place.

  19. Insight of Transmembrane Processes of Self-Assembling Nanotubes Based on a Cyclic Peptide Using Coarse Grained Molecular Dynamics Simulation.

    Science.gov (United States)

    Fu, Yankai; Yan, Tingxuan; Xu, Xia

    2017-09-28

    Transmembrane self-assembling cyclic peptide (SCP) nanotubes are promising candidates for delivering specific molecules through cell membranes. The detailed mechanisms behind the transmembrane processes, as well as stabilization factors of transmembrane structures, are difficult to elucidate through experiments. In this study, the effects of peptide sequence and oligomeric state on the transmembrane capabilities of SCP nanotubes and the perturbation of embedded SCP nanotubes acting on the membrane were investigated based on coarse grained molecular dynamics simulation. The simulation results reveal that hydrophilic SCP oligomers result in the elevation of the energy barrier while the oligomerization of hydrophobic SCPs causes the reduction of the energy barrier, further leading to membrane insertion. Once SCP nanotubes are embedded, membrane properties such as density, thickness, ordering state and lateral mobility are adjusted along the radial direction. This study provides insight into the transmembrane strategy of SCP nanotubes and sheds light on designing novel transport systems.

  20. Meeting report - Intercellular interactions in context: towards a mechanistic understanding of cells in organs.

    Science.gov (United States)

    Bryant, David; Johnson, Aaron

    2017-07-01

    The Company of Biologists held the workshop 'Intercellular interactions in context: towards a mechanistic understanding of cells in organs' at historic Wiston House in West Sussex, UK, 5-8 February 2017. The meeting brought together around 30 scientists from disparate backgrounds - yet with a common interest of how tissue morphogenesis occurs and its dysregulation leads to pathologies - to intensively discuss their latest research, the current state of the field, as well as any challenges for the future. This report summarises the concepts and challenges that arose as key questions for the fields of cell, cancer and developmental biology. By design of the organizers - Andrew Ewald (John Hopkins University, MA), John Wallingford (University of Texas at Austin, TX) and Peter Friedl (Radboud University, Nijmegen, The Netherlands) - the attendee makeup was cross-sectional: both in terms of career stage and scientific background. This intermingling was mirrored in the workshop format; all participants - irrespective of career stage - were given equal speaking and question time, and all early-career researchers also chaired a session, which promoted an atmosphere for discussions that were open, egalitarian and supportive. This was particularly evident in the scheduled 'out-of-the-box' sessions, which provided an avenue for participants to raise ideas and concepts or to discuss specific problems they wanted feedback or clarification on. In the following, rather than act as court reporters and convey chronological accounting of presentations, we present the questions that arose from the workshop and should be posed to the field at large, by discussing the presentations as they relate to these concepts. © 2017. Published by The Company of Biologists Ltd.

  1. Effect of spironolactone on renal and intercellular adhesion molecule-1 expression in Type 2 diabetic rats

    International Nuclear Information System (INIS)

    Zhang Suwan; Li Sumei; Zhai Fei; Zhang Li; Zhang Rong; Ru Yan

    2011-01-01

    Objective: To observe the influence of spironolactone on the serum and urine intercellular adhesion molecule-1 (ICAM-1) level, and the change of renal structure and function of type 2 diabetic rats. Methods: 30 healthy male SD rats were chosen 10 of them were randomly selected as normal controls (group NC) n=10; Then these rats were randomly divided into type 2 diabetes group (group DM) n=10 and type 2 diabetes + spironolactone treated group (group SPI) n=10. After 8 weeks, the levels of blood glucose, serum lipids, urine biochemical, renal pathological changes were examined; while the serum and urine ICAM-1 levels changes were also detected. Results: 1. Compared with group NC, the levels of fBG and HbA1c were significantly increased in group DM and group SPI (P 0.05). 2. After 8 weeks,the levels of ACR, URBP, UICAM-1, SICAM-1 and kidney/body weight ratio in group DM and group SPI were higher than group NC (P<0.05); the five indexes were significantly lower in group SPI compared with group DM (P<0.05). In addition, UICAM-1 excretion rate and SICAM-1 level showed positive correlations with ACR, URBP excretion rate and kidney/body weight ratio (P<0.01). 3. Pathology showed that the extent of glomerular lesions in rats in group SPI apparently reduced, ICAM-1 expression was decreased compared with that in group DM (P<0.01). Conclusion: Spironolactone can definitely protect type 2 diabetic kidney,and this protective effect was independent on the hypoglycemic effect. The mechanisms might be associated with its inhibition effect on ICAM-1 expression and its excretion. (authors)

  2. Intercellular adhesion molecule-1 expression by skeletal muscle cells augments myogenesis

    International Nuclear Information System (INIS)

    Goh, Qingnian; Dearth, Christopher L.; Corbett, Jacob T.; Pierre, Philippe; Chadee, Deborah N.; Pizza, Francis X.

    2015-01-01

    We previously demonstrated that the expression of intercellular adhesion molecule-1 (ICAM-1) by skeletal muscle cells after muscle overload contributes to ensuing regenerative and hypertrophic processes in skeletal muscle. The objective of the present study is to reveal mechanisms through which skeletal muscle cell expression of ICAM-1 augments regenerative and hypertrophic processes of myogenesis. This was accomplished by genetically engineering C2C12 myoblasts to stably express ICAM-1, and by inhibiting the adhesive and signaling functions of ICAM-1 through the use of a neutralizing antibody or cell penetrating peptide, respectively. Expression of ICAM-1 by cultured skeletal muscle cells augmented myoblast–myoblast adhesion, myotube formation, myonuclear number, myotube alignment, myotube–myotube fusion, and myotube size without influencing the ability of myoblasts to proliferate or differentiate. ICAM-1 augmented myotube formation, myonuclear accretion, and myotube alignment through a mechanism involving adhesion-induced activation of ICAM-1 signaling, as these dependent measures were reduced via antibody and peptide inhibition of ICAM-1. The adhesive and signaling functions of ICAM-1 also facilitated myotube hypertrophy through a mechanism involving myotube–myotube fusion, protein synthesis, and Akt/p70s6k signaling. Our findings demonstrate that ICAM-1 expression by skeletal muscle cells augments myogenesis, and establish a novel mechanism through which the inflammatory response facilitates growth processes in skeletal muscle. - Highlights: • We examined mechanisms through which skeletal muscle cell expression of ICAM-1 facilitates events of in vitro myogenesis. • Expression of ICAM-1 by cultured myoblasts did not influence their ability to proliferate or differentiate. • Skeletal muscle cell expression of ICAM-1 augmented myoblast fusion, myotube alignment, myotube–myotube fusion, and myotube size. • ICAM-1 augmented myogenic processes through

  3. Intercellular signaling via cyclic GMP diffusion through gap junctions restarts meiosis in mouse ovarian follicles.

    Science.gov (United States)

    Shuhaibar, Leia C; Egbert, Jeremy R; Norris, Rachael P; Lampe, Paul D; Nikolaev, Viacheslav O; Thunemann, Martin; Wen, Lai; Feil, Robert; Jaffe, Laurinda A

    2015-04-28

    Meiosis in mammalian oocytes is paused until luteinizing hormone (LH) activates receptors in the mural granulosa cells of the ovarian follicle. Prior work has established the central role of cyclic GMP (cGMP) from the granulosa cells in maintaining meiotic arrest, but it is not clear how binding of LH to receptors that are located up to 10 cell layers away from the oocyte lowers oocyte cGMP and restarts meiosis. Here, by visualizing intercellular trafficking of cGMP in real-time in live follicles from mice expressing a FRET sensor, we show that diffusion of cGMP through gap junctions is responsible not only for maintaining meiotic arrest, but also for rapid transmission of the signal that reinitiates meiosis from the follicle surface to the oocyte. Before LH exposure, the cGMP concentration throughout the follicle is at a uniformly high level of ∼2-4 μM. Then, within 1 min of LH application, cGMP begins to decrease in the peripheral granulosa cells. As a consequence, cGMP from the oocyte diffuses into the sink provided by the large granulosa cell volume, such that by 20 min the cGMP concentration in the follicle is uniformly low, ∼100 nM. The decrease in cGMP in the oocyte relieves the inhibition of the meiotic cell cycle. This direct demonstration that a physiological signal initiated by a stimulus in one region of an intact tissue can travel across many layers of cells via cyclic nucleotide diffusion through gap junctions could provide a general mechanism for diverse cellular processes.

  4. Endothelial targeting of high-affinity multivalent polymer nanocarriers directed to intercellular adhesion molecule 1.

    Science.gov (United States)

    Muro, Silvia; Dziubla, Thomas; Qiu, Weining; Leferovich, John; Cui, Xiumin; Berk, Erik; Muzykantov, Vladimir R

    2006-06-01

    Targeting of diagnostic and therapeutic agents to endothelial cells (ECs) provides an avenue to improve treatment of many maladies. For example, intercellular adhesion molecule 1 (ICAM-1), a constitutive endothelial cell adhesion molecule up-regulated in many diseases, is a good determinant for endothelial targeting of therapeutic enzymes and polymer nanocarriers (PNCs) conjugated with anti-ICAM (anti-ICAM/PNCs). However, intrinsic and extrinsic factors that control targeting of anti-ICAM/PNCs to ECs (e.g., anti-ICAM affinity and PNC valency and flow) have not been defined. In this study we tested in vitro and in vivo parameters of targeting to ECs of anti-ICAM/PNCs consisting of either prototype polystyrene or biodegradable poly(lactic-coglycolic) acid polymers (approximately 200 nm diameter spheres carrying approximately 200 anti-ICAM molecules). Anti-ICAM/PNCs, but not control IgG/PNCs 1) rapidly (t1/2 approximately 5 min) and specifically bound to tumor necrosis factor-activated ECs in a dose-dependent manner (Bmax approximately 350 PNC/cell) at both static and physiological shear stress conditions and 2) bound to ECs and accumulated in the pulmonary vasculature after i.v. injection in mice. Anti-ICAM/PNCs displayed markedly higher EC affinity versus naked anti-ICAM (Kd approximately 80 pM versus approximately 8 nM) in cell culture and, probably because of this factor, higher value (185.3 +/- 24.2 versus 50.5 +/- 1.5% injected dose/g) and selectivity (lung/blood ratio 81.0 +/- 10.9 versus 2.1 +/- 0.02, in part due to faster blood clearance) of pulmonary targeting. These results 1) show that reformatting monomolecular anti-ICAM into high-affinity multivalent PNCs boosts their vascular immuno-targeting, which withstands physiological hydrodynamics and 2) support potential anti-ICAM/PNCs utility for medical applications.

  5. Refractory heartburn: comparison of intercellular space diameter in documented GERD vs. functional heartburn.

    Science.gov (United States)

    Vela, Marcelo F; Craft, Brandon M; Sharma, Neeraj; Freeman, Janice; Hazen-Martin, Debra

    2011-05-01

    Refractory heartburn despite acid suppression may be explained by ongoing gastroesophageal reflux disease (GERD) or functional heartburn (FH), i.e., symptoms without evidence of GERD. Impedance-pH monitoring (impedance-pH) detects acid and nonacid reflux and is useful for evaluating acid-suppressed, refractory patients. Intercellular space diameter (ISD) of esophageal epithelium measured by transmission electron microscopy (TEM) is a marker of epithelial damage present in both erosive and nonerosive reflux disease. ISD has not been used to study refractory heartburn or FH. Our aim was to compare ISD in healthy controls and refractory heartburn patients with GERD and FH. In refractory heartburn patients (heartburn more than twice/week for at least 2 months despite proton pump inhibitor (PPI) b.i.d.), erosive esophagitis and/or abnormal impedance-pH (increased acid exposure or positive symptom index) defined GERD; normal esophagogastroduodenoscopy (EGD)/impedance-pH defined FH. Asymptomatic, healthy controls had normal EGD and pH-metry. Mean ISD in each subject, determined by blinded TEM of esophageal biopsies, was the average of 100 measurements (10 measurements in each of 10 micrographs). In all, 11 healthy controls, 11 FH, and 15 GERD patients were studied. Mean ISD was significantly higher in GERD compared with controls (0.87 vs. 0.32 μm, P=0.003) and FH (0.87 vs. 0.42 μm, P=0.012). Mean ISD was similar in FH and controls (0.42 vs. 0.32 μm, P=0.1). The proportion of patients with abnormal ISD was significantly higher for GERD compared with FH (60 vs. 9%, P=0.014). ISD is increased in refractory heartburn patients with GERD but not those with FH. Our findings suggest that measurement of ISD by TEM might be a useful tool to distinguish GERD from FH in patients with refractory heartburn.

  6. High plasma levels of soluble intercellular adhesion molecule (ICAM)-1 are associated with cerebral malaria.

    Science.gov (United States)

    Adukpo, Selorme; Kusi, Kwadwo A; Ofori, Michael F; Tetteh, John K A; Amoako-Sakyi, Daniel; Goka, Bamenla Q; Adjei, George O; Edoh, Dominic A; Akanmori, Bartholomew D; Gyan, Ben A; Dodoo, Daniel

    2013-01-01

    Cerebral malaria (CM) is responsible for most of the malaria-related deaths in children in sub-Saharan Africa. Although, not well understood, the pathogenesis of CM involves parasite and host factors which contribute to parasite sequestration through cytoadherence to the vascular endothelium. Cytoadherence to brain microvasculature is believed to involve host endothelial receptor, CD54 or intercellular adhesion molecule (ICAM)-1, while other receptors such as CD36 are generally involved in cytoadherence of parasites in other organs. We therefore investigated the contributions of host ICAM-1 expression and levels of antibodies against ICAM-1 binding variant surface antigen (VSA) on parasites to the development of CM. Paediatric malaria patients, 0.5 to 13 years were recruited and grouped into CM and uncomplicated malaria (UM) patients, based on well defined criteria. Standardized ELISA protocol was used to measure soluble ICAM-1 (sICAM-1) levels from acute plasma samples. Levels of IgG to CD36- or ICAM-1-binding VSA were measured by flow cytometry during acute and convalescent states. Wilcoxon sign rank-test analysis to compare groups revealed association between sICAM-1 levels and CM (p0.05). Median levels of antibodies to CD36-binding VSAs were also comparable between acute and convalescent samples within any patient group. Median levels of antibodies to ICAM-1-binding VSAs were however significantly lower at admission time than during recovery in both groups. High levels of sICAM-1 were associated with CM, and the sICAM-1 levels may reflect expression levels of the membrane bound form. Anti-VSA antibody levels to ICAM-binding parasites was more strongly associated with both UM and CM than antibodies to CD36 binding parasites. Thus, increasing host sICAM-1 levels were associated with CM whilst antibodies to parasite expressing non-ICAM-1-binding VSAs were not.

  7. High plasma levels of soluble intercellular adhesion molecule (ICAM-1 are associated with cerebral malaria.

    Directory of Open Access Journals (Sweden)

    Selorme Adukpo

    Full Text Available BACKGROUND: Cerebral malaria (CM is responsible for most of the malaria-related deaths in children in sub-Saharan Africa. Although, not well understood, the pathogenesis of CM involves parasite and host factors which contribute to parasite sequestration through cytoadherence to the vascular endothelium. Cytoadherence to brain microvasculature is believed to involve host endothelial receptor, CD54 or intercellular adhesion molecule (ICAM-1, while other receptors such as CD36 are generally involved in cytoadherence of parasites in other organs. We therefore investigated the contributions of host ICAM-1 expression and levels of antibodies against ICAM-1 binding variant surface antigen (VSA on parasites to the development of CM. METHODOLOGY/PRINCIPAL FINDINGS: Paediatric malaria patients, 0.5 to 13 years were recruited and grouped into CM and uncomplicated malaria (UM patients, based on well defined criteria. Standardized ELISA protocol was used to measure soluble ICAM-1 (sICAM-1 levels from acute plasma samples. Levels of IgG to CD36- or ICAM-1-binding VSA were measured by flow cytometry during acute and convalescent states. Wilcoxon sign rank-test analysis to compare groups revealed association between sICAM-1 levels and CM (p0.05. Median levels of antibodies to CD36-binding VSAs were also comparable between acute and convalescent samples within any patient group. Median levels of antibodies to ICAM-1-binding VSAs were however significantly lower at admission time than during recovery in both groups. CONCLUSIONS/SIGNIFICANCE: High levels of sICAM-1 were associated with CM, and the sICAM-1 levels may reflect expression levels of the membrane bound form. Anti-VSA antibody levels to ICAM-binding parasites was more strongly associated with both UM and CM than antibodies to CD36 binding parasites. Thus, increasing host sICAM-1 levels were associated with CM whilst antibodies to parasite expressing non-ICAM-1-binding VSAs were not.

  8. Intercellular adhesion molecule-1 expression by skeletal muscle cells augments myogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Goh, Qingnian; Dearth, Christopher L.; Corbett, Jacob T. [Department of Kinesiology, The University of Toledo, Toledo, OH (United States); Pierre, Philippe [Centre d’Immunologie de Marseille-Luminy U2M, Aix-Marseille Université, Marseille (France); INSERM U631, Institut National de la Santé et Recherche Médicale, Marseille (France); CNRS UMR6102, Centre National de la Recherche Scientifique, Marseille (France); Chadee, Deborah N. [Department of Biological Sciences, The University of Toledo, Toledo, OH (United States); Pizza, Francis X., E-mail: Francis.Pizza@utoledo.edu [Department of Kinesiology, The University of Toledo, Toledo, OH (United States)

    2015-02-15

    We previously demonstrated that the expression of intercellular adhesion molecule-1 (ICAM-1) by skeletal muscle cells after muscle overload contributes to ensuing regenerative and hypertrophic processes in skeletal muscle. The objective of the present study is to reveal mechanisms through which skeletal muscle cell expression of ICAM-1 augments regenerative and hypertrophic processes of myogenesis. This was accomplished by genetically engineering C2C12 myoblasts to stably express ICAM-1, and by inhibiting the adhesive and signaling functions of ICAM-1 through the use of a neutralizing antibody or cell penetrating peptide, respectively. Expression of ICAM-1 by cultured skeletal muscle cells augmented myoblast–myoblast adhesion, myotube formation, myonuclear number, myotube alignment, myotube–myotube fusion, and myotube size without influencing the ability of myoblasts to proliferate or differentiate. ICAM-1 augmented myotube formation, myonuclear accretion, and myotube alignment through a mechanism involving adhesion-induced activation of ICAM-1 signaling, as these dependent measures were reduced via antibody and peptide inhibition of ICAM-1. The adhesive and signaling functions of ICAM-1 also facilitated myotube hypertrophy through a mechanism involving myotube–myotube fusion, protein synthesis, and Akt/p70s6k signaling. Our findings demonstrate that ICAM-1 expression by skeletal muscle cells augments myogenesis, and establish a novel mechanism through which the inflammatory response facilitates growth processes in skeletal muscle. - Highlights: • We examined mechanisms through which skeletal muscle cell expression of ICAM-1 facilitates events of in vitro myogenesis. • Expression of ICAM-1 by cultured myoblasts did not influence their ability to proliferate or differentiate. • Skeletal muscle cell expression of ICAM-1 augmented myoblast fusion, myotube alignment, myotube–myotube fusion, and myotube size. • ICAM-1 augmented myogenic processes through

  9. Pannexin 1 channels play essential roles in urothelial mechanotransduction and intercellular signaling.

    Directory of Open Access Journals (Sweden)

    Hiromitsu Negoro

    Full Text Available Urothelial cells respond to bladder distension with ATP release, and ATP signaling within the bladder and from the bladder to the CNS is essential for proper bladder function. In other cell types, pannexin 1 (Panx1 channels provide a pathway for mechanically-induced ATP efflux and for ATP-induced ATP release through interaction with P2X7 receptors (P2X7Rs. We report that Panx1 and P2X7R are functionally expressed in the bladder mucosa and in immortalized human urothelial cells (TRT-HU1, and participate in urothelial ATP release and signaling. ATP release from isolated rat bladders induced by distention was reduced by the Panx1 channel blocker mefloquine (MFQ and was blunted in mice lacking Panx1 or P2X7R expression. Hypoosmotic shock induced YoPro dye uptake was inhibited by MFQ and the P2X7R blocker A438079 in TRT-HU1 cells, and was also blunted in primary urothelial cells derived from mice lacking Panx1 or P2X7R expression. Rinsing-induced mechanical stimulation of TRT-HU1 cells triggered ATP release, which was reduced by MFQ and potentiated in low divalent cation solution (LDPBS, a condition known to enhance P2X7R activation. ATP signaling evaluated as intercellular Ca2+ wave radius was significantly larger in LDPBS, reduced by MFQ and by apyrase (ATP scavenger. These findings indicate that Panx1 participates in urothelial mechanotransduction and signaling by providing a direct pathway for mechanically-induced ATP release and by functionally interacting with P2X7Rs.

  10. Effect of flow rate and temperature on transmembrane blood pressure drop in an extracorporeal artificial lung.

    Science.gov (United States)

    Park, M; Costa, E L V; Maciel, A T; Barbosa, E V S; Hirota, A S; Schettino, G de P; Azevedo, L C P

    2014-11-01

    Transmembrane pressure drop reflects the resistance of an artificial lung system to blood transit. Decreased resistance (low transmembrane pressure drop) enhances blood flow through the oxygenator, thereby, enhancing gas exchange efficiency. This study is part of a previous one where we observed the behaviour and the modulation of blood pressure drop during the passage of blood through artificial lung membranes. Before and after the induction of multi-organ dysfunction, the animals were instrumented and analysed for venous-venous extracorporeal membrane oxygenation, using a pre-defined sequence of blood flows. Blood flow and revolutions per minute (RPM) of the centrifugal pump varied in a linear fashion. At a blood flow of 5.5 L/min, pre- and post-pump blood pressures reached -120 and 450 mmHg, respectively. Transmembrane pressures showed a significant spread, particularly at blood flows above 2 L/min; over the entire range of blood flow rates, there was a positive association of pressure drop with blood flow (0.005 mmHg/mL/minute of blood flow) and a negative association of pressure drop with temperature (-4.828 mmHg/(°Celsius). These associations were similar when blood flows of below and above 2000 mL/minute were examined. During its passage through the extracorporeal system, blood is exposed to pressure variations from -120 to 450 mmHg. At high blood flows (above 2 L/min), the drop in transmembrane pressure becomes unpredictable and highly variable. Over the entire range of blood flows investigated (0-5500 mL/min), the drop in transmembrane pressure was positively associated with blood flow and negatively associated with body temperature. © The Author(s) 2014.

  11. Efect of intercellular extracts from banana inoculated leaves with Mycosphaerella fijiensis Morelet, on chloroplast electronic transport of Grande naine (AAA cv.

    Directory of Open Access Journals (Sweden)

    Michel Leiva-Mora

    2003-01-01

    Full Text Available Some foliar pathogens colonize intercellular spaces of damage tissues during infection process, mediated by toxins production and diffusion to kill adjacent healthy cells. Due to the absence of reliable bioassays, the physiologic effects of several phytotoxins are still ignored on cellular membranous systems of the affected cells. In the present work it was extracted the intercellular content from not inoculated and inoculated banana leaves with different Mycosphaerella fijiensis strains. Their effects on chloroplasts of Grande naine cv were evaluated by the absorbance evolution (595 nm of Hill reactive (DCPIP, mixture with 810 ì l of chloroplasts suspension and 99 ì l of the intercellular contents. The electronic exchange on chloroplasts suspension was inhibited by intercellular contents of inoculated leaves. The intercellular contents from leaves inoculated with I1 (high virulence strain had a major inhibiter effect respect to leaves inoculates with G1 strain (low virulence, showing a correspondence between the inhibiter effect of intercellular contents and the affection levels of affected tissues. The procedures used in this work will let to make studies concerned with Mycosphaerella fijiensis-Musa spp interactions and the future breeding programs. Key words: banana breeding, black Sigatoka, host pathogen interaction, physiological bioassays

  12. Conserved allosteric hot spots in the transmembrane domains of cystic fibrosis transmembrane conductance regulator (CFTR) channels and multidrug resistance protein (MRP) pumps.

    Science.gov (United States)

    Wei, Shipeng; Roessler, Bryan C; Chauvet, Sylvain; Guo, Jingyu; Hartman, John L; Kirk, Kevin L

    2014-07-18

    ATP-binding cassette (ABC) transporters are an ancient family of transmembrane proteins that utilize ATPase activity to move substrates across cell membranes. The ABCC subfamily of the ABC transporters includes active drug exporters (the multidrug resistance proteins (MRPs)) and a unique ATP-gated ion channel (cystic fibrosis transmembrane conductance regulator (CFTR)). The CFTR channel shares gating principles with conventional ligand-gated ion channels, but the allosteric network that couples ATP binding at its nucleotide binding domains (NBDs) with conformational changes in its transmembrane helices (TMs) is poorly defined. It is also unclear whether the mechanisms that govern CFTR gating are conserved with the thermodynamically distinct MRPs. Here we report a new class of gain of function (GOF) mutation of a conserved proline at the base of the pore-lining TM6. Multiple substitutions of this proline promoted ATP-free CFTR activity and activation by the weak agonist, 5'-adenylyl-β,γ-imidodiphosphate (AMP-PNP). TM6 proline mutations exhibited additive GOF effects when combined with a previously reported GOF mutation located in an outer collar of TMs that surrounds the pore-lining TMs. Each TM substitution allosterically rescued the ATP sensitivity of CFTR gating when introduced into an NBD mutant with defective ATP binding. Both classes of GOF mutations also rescued defective drug export by a yeast MRP (Yor1p) with ATP binding defects in its NBDs. We conclude that the conserved TM6 proline helps set the energy barrier to both CFTR channel opening and MRP-mediated drug efflux and that CFTR channels and MRP pumps utilize similar allosteric mechanisms for coupling conformational changes in their translocation pathways to ATP binding at their NBDs. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. Stability of Intercellular Exchange of Biochemical Substances Affected by Variability of Environmental Parameters

    Science.gov (United States)

    Mihailović, Dragutin T.; Budinčević, Mirko; Balaž, Igor; Mihailović, Anja

    Communication between cells is realized by exchange of biochemical substances. Due to internal organization of living systems and variability of external parameters, the exchange is heavily influenced by perturbations of various parameters at almost all stages of the process. Since communication is one of essential processes for functioning of living systems it is of interest to investigate conditions for its stability. Using previously developed simplified model of bacterial communication in a form of coupled difference logistic equations we investigate stability of exchange of signaling molecules under variability of internal and external parameters.

  14. Tunneling Nanotubes: Intimate Communication between Myeloid Cells.

    Science.gov (United States)

    Dupont, Maeva; Souriant, Shanti; Lugo-Villarino, Geanncarlo; Maridonneau-Parini, Isabelle; Vérollet, Christel

    2018-01-01

    Tunneling nanotubes (TNT) are dynamic connections between cells, which represent a novel route for cell-to-cell communication. A growing body of evidence points TNT towards a role for intercellular exchanges of signals, molecules, organelles, and pathogens, involving them in a diverse array of functions. TNT form among several cell types, including neuronal cells, epithelial cells, and almost all immune cells. In myeloid cells (e.g., macrophages, dendritic cells, and osteoclasts), intercellular communication via TNT contributes to their differentiation and immune functions. Importantly, TNT enable myeloid cells to communicate with a targeted neighboring or distant cell, as well as with other cell types, therefore creating a complex variety of cellular exchanges. TNT also contribute to pathogen spread as they serve as "corridors" from a cell to another. Herein, we addressed the complexity of the definition and in vitro characterization of TNT in innate immune cells, the different processes involved in their formation, and their relevance in vivo . We also assess our current understanding of how TNT participate in immune surveillance and the spread of pathogens, with a particular interest for HIV-1. Overall, despite recent progress in this growing research field, we highlight that further investigation is needed to better unveil the role of TNT in both physiological and pathological conditions.

  15. Tunneling Nanotubes: Intimate Communication between Myeloid Cells

    Directory of Open Access Journals (Sweden)

    Maeva Dupont

    2018-01-01

    Full Text Available Tunneling nanotubes (TNT are dynamic connections between cells, which represent a novel route for cell-to-cell communication. A growing body of evidence points TNT towards a role for intercellular exchanges of signals, molecules, organelles, and pathogens, involving them in a diverse array of functions. TNT form among several cell types, including neuronal cells, epithelial cells, and almost all immune cells. In myeloid cells (e.g., macrophages, dendritic cells, and osteoclasts, intercellular communication via TNT contributes to their differentiation and immune functions. Importantly, TNT enable myeloid cells to communicate with a targeted neighboring or distant cell, as well as with other cell types, therefore creating a complex variety of cellular exchanges. TNT also contribute to pathogen spread as they serve as “corridors” from a cell to another. Herein, we addressed the complexity of the definition and in vitro characterization of TNT in innate immune cells, the different processes involved in their formation, and their relevance in vivo. We also assess our current understanding of how TNT participate in immune surveillance and the spread of pathogens, with a particular interest for HIV-1. Overall, despite recent progress in this growing research field, we highlight that further investigation is needed to better unveil the role of TNT in both physiological and pathological conditions.

  16. Expression of genes encoding multi-transmembrane proteins in specific primate taste cell populations.

    Directory of Open Access Journals (Sweden)

    Bryan D Moyer

    Full Text Available BACKGROUND: Using fungiform (FG and circumvallate (CV taste buds isolated by laser capture microdissection and analyzed using gene arrays, we previously constructed a comprehensive database of gene expression in primates, which revealed over 2,300 taste bud-associated genes. Bioinformatics analyses identified hundreds of genes predicted to encode multi-transmembrane domain proteins with no previous association with taste function. A first step in elucidating the roles these gene products play in gustation is to identify the specific taste cell types in which they are expressed. METHODOLOGY/PRINCIPAL FINDINGS: Using double label in situ hybridization analyses, we identified seven new genes expressed in specific taste cell types, including sweet, bitter, and umami cells (TRPM5-positive, sour cells (PKD2L1-positive, as well as other taste cell populations. Transmembrane protein 44 (TMEM44, a protein with seven predicted transmembrane domains with no homology to GPCRs, is expressed in a TRPM5-negative and PKD2L1-negative population that is enriched in the bottom portion of taste buds and may represent developmentally immature taste cells. Calcium homeostasis modulator 1 (CALHM1, a component of a novel calcium channel, along with family members CALHM2 and CALHM3; multiple C2 domains; transmembrane 1 (MCTP1, a calcium-binding transmembrane protein; and anoctamin 7 (ANO7, a member of the recently identified calcium-gated chloride channel family, are all expressed in TRPM5 cells. These proteins may modulate and effect calcium signalling stemming from sweet, bitter, and umami receptor activation. Synaptic vesicle glycoprotein 2B (SV2B, a regulator of synaptic vesicle exocytosis, is expressed in PKD2L1 cells, suggesting that this taste cell population transmits tastant information to gustatory afferent nerve fibers via exocytic neurotransmitter release. CONCLUSIONS/SIGNIFICANCE: Identification of genes encoding multi-transmembrane domain proteins

  17. Exosomes: mediators of communication in eukaryotes

    Directory of Open Access Journals (Sweden)

    María A Lopez-Verrilli

    2013-01-01

    Full Text Available In addition to the established mechanisms of intercellular signaling, a new way of communication has gained much attention in the last decade: communication mediated by exosomes. Exosomes are nanovesicles (with a diameter of 40-120 nm secreted into the extracellular space by the multivesicular endosome after its outer membrane fuses with the plasma membrane. Once released, exosomes modulate the response of the recipient cells that recognize them. This indicates that exosomes operate in a specific manner and participate in the regulation of the target cell. Remarkably, exosomes occur from unicellular organisms to mammals, suggesting an evolutionarily conserved mechanism of communication. In this review we describe the cascade of exosome formation, intracellular traffic, secretion, and internalization by recipient cells, and review their most relevant effects. We also highlight important steps that are still poorly understood.

  18. Intracellular disposition of chitosan nanoparticles in macrophages: intracellular uptake, exocytosis, and intercellular transport

    Directory of Open Access Journals (Sweden)

    Jiang LQ

    2017-08-01

    mechanisms, providing the basis for how CsNPs are recognized by the MPS; such information is crucial to numerous medical applications of CsNPs. Keywords: exocytosis, uptake, intercellular transport, chitosan nanoparticles, macrophages

  19. The soluble loop BC region guides, but not dictates, the assembly of the transmembrane cytochrome b6.

    Directory of Open Access Journals (Sweden)

    Lydia Tome-Stangl

    Full Text Available Studying folding and assembly of naturally occurring α-helical transmembrane proteins can inspire the design of membrane proteins with defined functions. Thus far, most studies have focused on the role of membrane-integrated protein regions. However, to fully understand folding pathways and stabilization of α-helical membrane proteins, it is vital to also include the role of soluble loops. We have analyzed the impact of interhelical loops on folding, assembly and stability of the heme-containing four-helix bundle transmembrane protein cytochrome b6 that is involved in charge transfer across biomembranes. Cytochrome b6 consists of two transmembrane helical hairpins that sandwich two heme molecules. Our analyses strongly suggest that the loop connecting the helical hairpins is not crucial for positioning the two protein "halves" for proper folding and assembly of the holo-protein. Furthermore, proteolytic removal of any of the remaining two loops, which connect the two transmembrane helices of a hairpin structure, appears to also not crucially effect folding and assembly. Overall, the transmembrane four-helix bundle appears to be mainly stabilized via interhelical interactions in the transmembrane regions, while the soluble loop regions guide assembly and stabilize the holo-protein. The results of this study might steer future strategies aiming at designing heme-binding four-helix bundle structures, involved in transmembrane charge transfer reactions.

  20. Quantitative measurements of intercellular adhesion between a macrophage and cancer cells using a cup-attached AFM chip.

    Science.gov (United States)

    Kim, Hyonchol; Yamagishi, Ayana; Imaizumi, Miku; Onomura, Yui; Nagasaki, Akira; Miyagi, Yohei; Okada, Tomoko; Nakamura, Chikashi

    2017-07-01

    Intercellular adhesion between a macrophage and cancer cells was quantitatively measured using atomic force microscopy (AFM). Cup-shaped metal hemispheres were fabricated using polystyrene particles as a template, and a cup was attached to the apex of the AFM cantilever. The cup-attached AFM chip (cup-chip) approached a murine macrophage cell (J774.2), the cell was captured on the inner concave of the cup, and picked up by withdrawing the cup-chip from the substrate. The cell-attached chip was advanced towards a murine breast cancer cell (FP10SC2), and intercellular adhesion between the two cells was quantitatively measured. To compare cell adhesion strength, the work required to separate two adhered cells (separation work) was used as a parameter. Separation work was almost 2-fold larger between a J774.2 cell and FP10SC2 cell than between J774.2 cell and three additional different cancer cells (4T1E, MAT-LyLu, and U-2OS), two FP10SC2 cells, or two J774.2 cells. FP10SC2 was established from 4T1E as a highly metastatic cell line, indicates separation work increased as the malignancy of cancer cells became higher. One possible explanation of the strong adhesion of macrophages to cancer cells observed in this study is that the measurement condition mimicked the microenvironment of tumor-associated macrophages (TAMs) in vivo, and J774.2 cells strongly expressed CD204, which is a marker of TAMs. The results of the present study, which were obtained by measuring cell adhesion strength quantitatively, indicate that the fabricated cup-chip is a useful tool for measuring intercellular adhesion easily and quantitatively. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Role of ATP binding and hydrolysis in the gating of the cystic fibrosis transmembrane conductance regulator

    Directory of Open Access Journals (Sweden)

    Taras Gout

    2012-01-01

    Full Text Available The CFTR gene is unique within the ATP-binding cassette (ABC protein family, predominantly of transporters, by coding a chloride channel. The gating mechanism of ABC proteins has been characterized by the ATP Switch model in terms cycles of dimer formation and dissociation linked to ATP binding and hydrolysis, respectively. It would be of interest to assess the extent that Cystic Fibrosis Transmembrane Conductance Regulator (CFTR, a functional channel, fits the ATP Switch model for ABC transporters. Additional transporter mechanisms, namely those of Pgp and HlyB, are discussed for perspective. Literature search of databases selected key references in comparing and contrasting the gating mechanism. CFTR is a functional chloride channel facilitating transmembrane anion flow down electrochemical gradients. A dysfunctional CFTR protein results in cystic fibrosis, a fatal pleiotropic disease currently managed symptomatically. Understanding the gating mechanism will help target drug development aimed at alleviating and curing the disease.

  2. Recombinant expression in E. coli of human FGFR2 with its transmembrane and extracellular domains

    Directory of Open Access Journals (Sweden)

    Adam Bajinting

    2017-06-01

    Full Text Available Fibroblast growth factor receptors (FGFRs are a family of receptor tyrosine kinases containing three domains: an extracellular receptor domain, a single transmembrane helix, and an intracellular tyrosine kinase domain. FGFRs are activated by fibroblast growth factors (FGFs as part of complex signal transduction cascades regulating angiogenesis, skeletal formation, cell differentiation, proliferation, cell survival, and cancer. We have developed the first recombinant expression system in E. coli to produce a construct of human FGFR2 containing its transmembrane and extracellular receptor domains. We demonstrate that the expressed construct is functional in binding heparin and dimerizing. Size exclusion chromatography demonstrates that the purified FGFR2 does not form a complex with FGF1 or adopts an inactive dimer conformation. Progress towards the successful recombinant production of intact FGFRs will facilitate further biochemical experiments and structure determination that will provide insight into how extracellular FGF binding activates intracellular kinase activity.

  3. Intrinsic potential of cell membranes: opposite effects of lipid transmembrane asymmetry and asymmetric salt ion distribution

    DEFF Research Database (Denmark)

    Gurtovenko, Andrey A; Vattulainen, Ilpo

    2009-01-01

    Using atomic-scale molecular dynamics simulations, we consider the intrinsic cell membrane potential that is found to originate from a subtle interplay between lipid transmembrane asymmetry and the asymmetric distribution of monovalent salt ions on the two sides of the cell membrane. It turns out......Cl saline solution and the PE leaflet is exposed to KCl, the outcome is that the effects of asymmetric lipid and salt ion distributions essentially cancel one another almost completely. Overall, our study highlights the complex nature of the intrinsic potential of cell membranes under physiological...... that both the asymmetric distribution of phosphatidylcholine (PC) and phosphatidylethanolamine (PE) lipids across a membrane and the asymmetric distribution of NaCl and KCl induce nonzero drops in the transmembrane potential. However, these potential drops are opposite in sign. As the PC leaflet faces a Na...

  4. TMDIM: an improved algorithm for the structure prediction of transmembrane domains of bitopic dimers

    Science.gov (United States)

    Cao, Han; Ng, Marcus C. K.; Jusoh, Siti Azma; Tai, Hio Kuan; Siu, Shirley W. I.

    2017-09-01

    α-Helical transmembrane proteins are the most important drug targets in rational drug development. However, solving the experimental structures of these proteins remains difficult, therefore computational methods to accurately and efficiently predict the structures are in great demand. We present an improved structure prediction method TMDIM based on Park et al. (Proteins 57:577-585, 2004) for predicting bitopic transmembrane protein dimers. Three major algorithmic improvements are introduction of the packing type classification, the multiple-condition decoy filtering, and the cluster-based candidate selection. In a test of predicting nine known bitopic dimers, approximately 78% of our predictions achieved a successful fit (RMSD PHP, MySQL and Apache, with all major browsers supported.

  5. Cancer Research Advance in CKLF-like MARVEL Transmembrane Domain Containing Member Family (Review).

    Science.gov (United States)

    Lu, Jia; Wu, Qian-Qian; Zhou, Ya-Bo; Zhang, Kai-Hua; Pang, Bing-Xin; Li, Liang; Sun, Nan; Wang, Heng-Shu; Zhang, Song; Li, Wen-Jian; Zheng, Wei; Liu, Wei

    2016-01-01

    CKLF-like MARVEL transmembrane domain-containing family (CMTM) is a novel family of genes first reported at international level by Peking University Human Disease Gene Research Center. The gene products are between chemokines and the transmembrane-4 superfamily. Loaceted in several human chromosomes, CMTMs, which are unregulated in kinds of tumors, are potential tumor suppressor genes consisting of CKLF and CMTM1 to CMTM8. CMTMs play important roles in immune, male reproductive and hematopoietic systems. Also, it has been approved that CMTM family has strong connection with diseases of autoimmunity, haematopoietic system and haematopoietic system. The in-depth study in recent years found the close relation between CMTMs and umorigenesis, tumor development and metastasis. CMTM family has a significant clinical value in diagnosis and treatment to the diseases linking to tumor and immune system.

  6. [Research advances in CKLF-like MARVEL transmembrane domain containing member 5].

    Science.gov (United States)

    Yuan, Ye-qing; Xiao, Yun-bei; Liu, Zhen-hua; Zhang, Xiao-wei; Xu, Tao; Wang, Xiao-feng

    2012-12-01

    CKLF-like MARVEL transmembrane domain containing member(CMTM)is a novel generic family firstly reported by Peking University Center for Human Disease Genomics. CMTM5 belongs to this family and has exhibited tumor-inhibiting activities. It can encode proteins approaching to the transmembrane 4 superfamily(TM4SF). CMTM5 is broadly expressed in normal adult and fetal human tissues, but is undetectable or down-regulated in most carcinoma cell lines and tissues. Restoration of CMTM5 may inhibit the proliferation, migration, and invasion of carcinoma cells. Although the exact mechanism of its anti-tumor activity remains unclear, CMTM5 may be involved in various signaling pathways governing the occurrence and development of tumors. CMTM5 may be a new target in the gene therapies for tumors, while further studies on CMTM5 and its anti-tumor mechanisms are warranted.

  7. First principles design of a core bioenergetic transmembrane electron-transfer protein

    Energy Technology Data Exchange (ETDEWEB)

    Goparaju, Geetha; Fry, Bryan A.; Chobot, Sarah E.; Wiedman, Gregory; Moser, Christopher C.; Leslie Dutton, P.; Discher, Bohdana M.

    2016-05-01

    Here we describe the design, Escherichia coli expression and characterization of a simplified, adaptable and functionally transparent single chain 4-α-helix transmembrane protein frame that binds multiple heme and light activatable porphyrins. Such man-made cofactor-binding oxidoreductases, designed from first principles with minimal reference to natural protein sequences, are known as maquettes. This design is an adaptable frame aiming to uncover core engineering principles governing bioenergetic transmembrane electron-transfer function and recapitulate protein archetypes proposed to represent the origins of photosynthesis. This article is part of a Special Issue entitled Biodesign for Bioenergetics — the design and engineering of electronic transfer cofactors, proteins and protein networks, edited by Ronald L. Koder and J.L. Ross Anderson.

  8. Combined effect of cortical cytoskeleton and transmembrane proteins on domain formation in biomembranes

    DEFF Research Database (Denmark)

    Sikder, K. U.; Stone, K. A.; Kumar, P. B. S.

    2014-01-01

    We investigate the combined effects of transmembrane proteins and the subjacent cytoskeleton on the dynamics of phase separation in multicomponent lipid bilayers using computer simulations of a particle-based implicit solvent model for lipid membranes with soft-core interactions. We find that mic......We investigate the combined effects of transmembrane proteins and the subjacent cytoskeleton on the dynamics of phase separation in multicomponent lipid bilayers using computer simulations of a particle-based implicit solvent model for lipid membranes with soft-core interactions. We find...... that microphase separation can be achieved by the protein confinement by the cytoskeleton. Our results have relevance to the finite size of lipid rafts in the plasma membrane of mammalian cells. (C) 2014 AIP Publishing LLC....

  9. Increasing the endogenous NO level causes catalase inactivation and reactivation of intercellular apoptosis signaling specifically in tumor cells

    Science.gov (United States)

    Bauer, Georg

    2015-01-01

    Tumor cells generate extracellular superoxide anions and are protected against intercellular apoptosis-inducing HOCl- and NO/peroxynitrite signaling through the expression of membrane-associated catalase. This enzyme decomposes H2O2 and thus prevents HOCl synthesis. It efficiently interferes with NO/peroxynitrite signaling through oxidation of NO and decomposition of peroxynitrite. The regulatory potential of catalase at the crosspoint of ROS and RNS chemical biology, as well as its high local concentration on the outside of the cell membrane of tumor cells, establish tight control of intercellular signaling and thus prevent tumor cell apoptosis. Therefore, inhibition of catalase or its inactivation by singlet oxygen reactivate intercellular apoptosis-inducing signaling. Nitric oxide and peroxynitrite are connected with catalase in multiple and meaningful ways, as (i) NO can be oxidated by compound I of catalase, (ii) NO can reversibly inhibit catalase, (iii) peroxynitrite can be decomposed by catalase and (iv) the interaction between peroxynitrite and H2O2 leads to the generation of singlet oxygen that inactivates catalase. Therefore, modulation of the concentration of free NO through addition of arginine, inhibition of arginase, induction of NOS expression or inhibition of NO dioxygenase triggers an autoamplificatory biochemical cascade that is based on initial formation of singlet oxygen, amplification of superoxide anion/H2O2 and NO generation through singlet oxygen dependent stimulation of the FAS receptor and caspase-8. Finally, singlet oxygen is generated at sufficiently high concentration to inactivate protective catalase and to reactivate intercellular apoptosis-inducing ROS signaling. This regulatory network allows to establish several pathways for synergistic interactions, like the combination of modulators of NO metabolism with enhancers of superoxide anion generation, modulators of NO metabolism that act at different targets and between modulators of

  10. Increasing the endogenous NO level causes catalase inactivation and reactivation of intercellular apoptosis signaling specifically in tumor cells.

    Science.gov (United States)

    Bauer, Georg

    2015-12-01

    Tumor cells generate extracellular superoxide anions and are protected against intercellular apoptosis-inducing HOCl- and NO/peroxynitrite signaling through the expression of membrane-associated catalase. This enzyme decomposes H2O2 and thus prevents HOCl synthesis. It efficiently interferes with NO/peroxynitrite signaling through oxidation of NO and decomposition of peroxynitrite. The regulatory potential of catalase at the crosspoint of ROS and RNS chemical biology, as well as its high local concentration on the outside of the cell membrane of tumor cells, establish tight control of intercellular signaling and thus prevent tumor cell apoptosis. Therefore, inhibition of catalase or its inactivation by singlet oxygen reactivate intercellular apoptosis-inducing signaling. Nitric oxide and peroxynitrite are connected with catalase in multiple and meaningful ways, as (i) NO can be oxidated by compound I of catalase, (ii) NO can reversibly inhibit catalase, (iii) peroxynitrite can be decomposed by catalase and (iv) the interaction between peroxynitrite and H2O2 leads to the generation of singlet oxygen that inactivates catalase. Therefore, modulation of the concentration of free NO through addition of arginine, inhibition of arginase, induction of NOS expression or inhibition of NO dioxygenase triggers an autoamplificatory biochemical cascade that is based on initial formation of singlet oxygen, amplification of superoxide anion/H2O2 and NO generation through singlet oxygen dependent stimulation of the FAS receptor and caspase-8. Finally, singlet oxygen is generated at sufficiently high concentration to inactivate protective catalase and to reactivate intercellular apoptosis-inducing ROS signaling. This regulatory network allows to establish several pathways for synergistic interactions, like the combination of modulators of NO metabolism with enhancers of superoxide anion generation, modulators of NO metabolism that act at different targets and between modulators of

  11. Expression and regulation of transmembrane transporters in healthy intestine and gastrointestinal diseases

    OpenAIRE

    Hruz, Petr

    2006-01-01

    Transmembrane transporters mediate energy dependent or independent translocation of drugs, potentially toxic compounds, and of various endogenous substrates such as bile acids and bilirubin across membranes. In this thesis the focus is on two classes of transporters, the ATPbinding cassette (ABC) transporters, which mediate ATP dependent transport and the solute carriers (SLC) which use electrochemical gradients for their transport. The transporters are expressed on membranes o...

  12. Relative transmembrane segment rearrangements during BK channel activation resolved by structurally assigned fluorophore–quencher pairing

    Science.gov (United States)

    Pantazis, Antonios

    2012-01-01

    Voltage-activated proteins can sense, and respond to, changes in the electric field pervading the cell membrane by virtue of a transmembrane helix bundle, the voltage-sensing domain (VSD). Canonical VSDs consist of four transmembrane helices (S1–S4) of which S4 is considered a principal component because it possesses charged residues immersed in the electric field. Membrane depolarization compels the charges, and by extension S4, to rearrange with respect to the field. The VSD of large-conductance voltage- and Ca-activated K+ (BK) channels exhibits two salient inconsistencies from the canonical VSD model: (1) the BK channel VSD possesses an additional nonconserved transmembrane helix (S0); and (2) it exhibits a “decentralized” distribution of voltage-sensing charges, in helices S2 and S3, in addition to S4. Considering these unique features, the voltage-dependent rearrangements of the BK VSD could differ significantly from the standard model of VSD operation. To understand the mode of operation of this unique VSD, we have optically tracked the relative motions of the BK VSD transmembrane helices during activation, by manipulating the quenching environment of site-directed fluorescent labels with native and introduced Trp residues. Having previously reported that S0 and S4 diverge during activation, in this work we demonstrate that S4 also diverges from S1 and S2, whereas S2, compelled by its voltage-sensing charged residues, moves closer to S1. This information contributes spatial constraints for understanding the BK channel voltage-sensing process, revealing the structural rearrangements in a non-canonical VSD. PMID:22802360

  13. Functional relevance of aromatic residues in the first transmembrane domain of P2X receptors

    Czech Academy of Sciences Publication Activity Database

    Jindřichová, Marie; Vávra, Vojtěch; Obšil, Tomáš; Stojilkovic, S. S.; Zemková, Hana

    2009-01-01

    Roč. 109, č. 3 (2009), s. 923-934 ISSN 0022-3042 R&D Projects: GA MŠk(CZ) LC554; GA AV ČR(CZ) IAA5011408; GA AV ČR(CZ) IAA500110702; GA AV ČR(CZ) IAA500110910 Institutional research plan: CEZ:AV0Z50110509 Keywords : purinergic receptors * gating * transmembrane domain Subject RIV: FH - Neuro logy Impact factor: 3.999, year: 2009

  14. Combined effect of cortical cytoskeleton and transmembrane proteins on domain formation in biomembranes

    Science.gov (United States)

    Sikder, Md. Kabir Uddin; Stone, Kyle A.; Kumar, P. B. Sunil; Laradji, Mohamed

    2014-01-01

    We investigate the combined effects of transmembrane proteins and the subjacent cytoskeleton on the dynamics of phase separation in multicomponent lipid bilayers using computer simulations of a particle-based implicit solvent model for lipid membranes with soft-core interactions. We find that microphase separation can be achieved by the protein confinement by the cytoskeleton. Our results have relevance to the finite size of lipid rafts in the plasma membrane of mammalian cells. PMID:25106608

  15. Simulations of Skin Barrier Function: Free Energies of Hydrophobic and Hydrophilic Transmembrane Pores in Ceramide Bilayers

    OpenAIRE

    Notman, Rebecca; Anwar, Jamshed; Briels, W. J.; Noro, Massimo G.; den Otter, Wouter K.

    2008-01-01

    Transmembrane pore formation is central to many biological processes such as ion transport, cell fusion, and viral infection. Furthermore, pore formation in the ceramide bilayers of the stratum corneum may be an important mechanism by which penetration enhancers such as dimethylsulfoxide (DMSO) weaken the barrier function of the skin. We have used the potential of mean constraint force (PMCF) method to calculate the free energy of pore formation in ceramide bilayers in both the innate gel pha...

  16. Electrochemical platform for the detection of transmembrane proteins reconstituted into liposomes

    Czech Academy of Sciences Publication Activity Database

    Vacek, J.; Zatloukalová, M.; Geletičová, J.; Kubala, M.; Modriansky, M.; Fekete, Ladislav; Mašek, J.; Hubatka, F.; Turánek, J.

    2016-01-01

    Roč. 88, č. 8 (2016), s. 4548-4556 ISSN 0003-2700 R&D Projects: GA MŠk LO1409; GA MŠk LM2015088 Institutional support: RVO:68378271 Keywords : detection * transmembrane proteins * liposomes * electrochemistry Subject RIV: BM - Solid Matter Physics ; Magnetism OBOR OECD: Condensed matter physics (including formerly solid state physics, supercond.) Impact factor: 6.320, year: 2016

  17. 15N and 31P solid-state NMR study of transmembrane domain alignment of M2 protein of influenza A virus in hydrated cylindrical lipid bilayers confined to anodic aluminum oxide nanopores.

    Science.gov (United States)

    Chekmenev, Eduard Y; Hu, Jun; Gor'kov, Peter L; Brey, William W; Cross, Timothy A; Ruuge, Andres; Smirnov, Alex I

    2005-04-01

    This communication reports the first example of a high resolution solid-state 15N 2D PISEMA NMR spectrum of a transmembrane peptide aligned using hydrated cylindrical lipid bilayers formed inside nanoporous anodic aluminum oxide (AAO) substrates. The transmembrane domain SSDPLVVA(A-15N)SIIGILHLILWILDRL of M2 protein from influenza A virus was reconstituted in hydrated 1,2-dimyristoyl-sn-glycero-3-phosphatidylcholine bilayers that were macroscopically aligned by a conventional micro slide glass support or by the AAO nanoporous substrate. 15N and 31P NMR spectra demonstrate that both the phospholipids and the protein transmembrane domain are uniformly aligned in the nanopores. Importantly, nanoporous AAO substrates may offer several advantages for membrane protein alignment in solid-state NMR studies compared to conventional methods. Specifically, higher thermal conductivity of aluminum oxide is expected to suppress thermal gradients associated with inhomogeneous radio frequency heating. Another important advantage of the nanoporous AAO substrate is its excellent accessibility to the bilayer surface for exposure to solute molecules. Such high accessibility achieved through the substrate nanochannel network could facilitate a wide range of structure-function studies of membrane proteins by solid-state NMR.

  18. Cell-cell communication in the kidney microcirculation

    DEFF Research Database (Denmark)

    Sørensen, Charlotte Mehlin; von Holstein-Rathlou, Niels-Henrik

    2012-01-01

    the postglomerular vasculature. Cxs form gap junctions between neighboring cells, and as in other organ systems, the major function of Cxs in the kidney appears to be mediation of intercellular communication. Cxs may also form hemichannels that allow cellular secretion of signaling molecules like ATP, and thereby...... mediate paracrine signaling. Renal Cxs facilitate vascular conduction, juxtaglomerlar apparatus calcium signaling, and enable ECs and VSMCs to communicate. Thus, current research suggests multiple roles for Cxs in important regulatory mechanisms within the kidney, including the renin-angiotensin system...

  19. Generation and Nuclear Translocation of Sumoylated Transmembrane Fragment of Cell Adhesion Molecule L1

    Science.gov (United States)

    Lutz, David; Wolters-Eisfeld, Gerrit; Joshi, Gunjan; Djogo, Nevena; Jakovcevski, Igor; Schachner, Melitta; Kleene, Ralf

    2012-01-01

    The functions of the cell adhesion molecule L1 in the developing and adult nervous system are triggered by homophilic and heterophilic interactions that stimulate signal transductions that activate cellular responses. Here, we show that stimulation of signaling by function-triggering L1 antibodies or L1-Fc leads to serine protease-dependent cleavage of full-length L1 at the plasma membrane and generation of a sumoylated transmembrane 70-kDa fragment comprising the intracellular and transmembrane domains and part of the extracellular domain. The 70-kDa transmembrane fragment is transported from the plasma membrane to a late endosomal compartment, released from endosomal membranes into the cytoplasm, and transferred from there into the nucleus by a pathway that depends on importin and chromatin-modifying protein 1. Mutation of the sumoylation site at Lys1172 or of the nuclear localization signal at Lys1147 abolished L1-stimulated generation or nuclear import of the 70-kDa fragment, respectively. Nuclear import of the 70-kDa fragment may activate cellular responses in parallel or in association with phosphorylation-dependent signaling pathways. Alterations in the levels of the 70-kDa fragment during development and in the adult after spinal cord injury or in a mouse model of Alzheimer disease suggest that this fragment is functionally implicated in development, regeneration, neurodegeneration, tumorigenesis, and possibly synaptic plasticity in the mature nervous system. PMID:22431726

  20. The transmembrane collagen COL-99 guides longitudinally extending axons in C. elegans.

    Science.gov (United States)

    Taylor, Jesse; Unsoeld, Thomas; Hutter, Harald

    2018-06-01

    We have identified the transmembrane collagen, COL-99, in a genetic screen for novel genes involved in axon guidance in the nematode C. elegans. COL-99 is similar to transmembrane collagens type XIII, XXIII and XXV in vertebrates. col-99 mutants exhibit guidance defects in axons extending along the major longitudinal axon tracts, most prominently the left ventral nerve cord (VNC). COL-99 is expressed in the hypodermis during the time of axon outgrowth. We provide evidence that a furin cleavage site in COL-99 is essential for function, suggesting that COL-99 is released from the cells producing it. Vertebrate homologs of COL-99 have been shown to be expressed in mammalian nervous systems and linked to various neurological disease but have not been associated with guidance of extending neurons. col-99 acts genetically with the discoidin domain receptors ddr-1 and ddr-2, which are expressed by neurons affected in col-99 mutants. Discoidin domain receptors are activated by collagens in vertebrates. DDR-1 and DDR-2 may function as receptors for COL-99. Our results establish a novel role for a transmembrane collagen in axonal guidance and asymmetry establishment of the VNC. Copyright © 2018 Elsevier Inc. All rights reserved.

  1. The role of membrane microdomains in transmembrane signaling through the epithelial glycoprotein Gp140/CDCP1

    Science.gov (United States)

    Alvares, Stacy M.; Dunn, Clarence A.; Brown, Tod A.; Wayner, Elizabeth E.; Carter, William G.

    2008-01-01

    Cell adhesion to the extracellular matrix (ECM) via integrin adhesion receptors initiates signaling cascades leading to changes in cell behavior. While integrin clustering is necessary to initiate cell attachment to the matrix, additional membrane components are necessary to mediate the transmembrane signals and the cell adhesion response that alter downstream cell behavior. Many of these signaling components reside in glycosphingolipid-rich and cholesterol-rich membrane domains such as Tetraspanin Enriched Microdomains (TEMs)/Glycosynapse 3 and Detergent-Resistant Microdomains (DRMs), also known as lipid rafts. In the following article, we will review examples of how components in these membrane microdomains modulate integrin adhesion after initial attachment to the ECM. Additionally, we will present data on a novel adhesion-responsive transmembrane glycoprotein Gp140/CUB Domain Containing Protein 1, which clusters in epithelial cell-cell contacts. Gp140 can then be phosphorylated by Src Family Kinases at tyrosine 734 in response to outside-in signals- possibly through interactions involving the extracellular CUB domains. Data presented here suggests that outside-in signals through Gp140 in cell-cell contacts assemble membrane clusters that associate with membrane microdomains to recruit and activate SFKs. Active SFKs then mediate phosphorylation of Gp140, SFK and PKCδ with Gp140 acting as a transmembrane scaffold for these kinases. We propose that the clustering of Gp140 and signaling components in membrane microdomains in cell-cell contacts contributes to changes in cell behavior. PMID:18269919

  2. Structure of FGFR3 transmembrane domain dimer: implications for signaling and human pathologies.

    Science.gov (United States)

    Bocharov, Eduard V; Lesovoy, Dmitry M; Goncharuk, Sergey A; Goncharuk, Marina V; Hristova, Kalina; Arseniev, Alexander S

    2013-11-05

    Fibroblast growth factor receptor 3 (FGFR3) transduces biochemical signals via lateral dimerization in the plasma membrane, and plays an important role in human development and disease. Eight different pathogenic mutations, implicated in cancers and growth disorders, have been identified in the FGFR3 transmembrane segment. Here, we describe the dimerization of the FGFR3 transmembrane domain in membrane-mimicking DPC/SDS (9/1) micelles. In the solved NMR structure, the two transmembrane helices pack into a symmetric left-handed dimer, with intermolecular stacking interactions occurring in the dimer central region. Some pathogenic mutations fall within the helix-helix interface, whereas others are located within a putative alternative interface. This implies that although the observed dimer structure is important for FGFR3 signaling, the mechanism of FGFR3-mediated transduction across the membrane is complex. We propose an FGFR3 signaling mechanism that is based on the solved structure, available structures of isolated soluble FGFR domains, and published biochemical and biophysical data. Copyright © 2013 Elsevier Ltd. All rights reserved.

  3. Multi-level communication of human retinal pigment epithelial cells via tunneling nanotubes.

    Directory of Open Access Journals (Sweden)

    Dierk Wittig

    Full Text Available BACKGROUND: Tunneling nanotubes (TNTs may offer a very specific and effective way of intercellular communication. Here we investigated TNTs in the human retinal pigment epithelial (RPE cell line ARPE-19. Morphology of TNTs was examined by immunostaining and scanning electron microscopy. To determine the function of TNTs between cells, we studied the TNT-dependent intercellular communication at different levels including electrical and calcium signalling, small molecular diffusion as well as mitochondrial re-localization. Further, intercellular organelles transfer was assayed by FACS analysis. METHODOLOGY AND PRINCIPAL FINDINGS: Microscopy showed that cultured ARPE-19 cells are frequently connected by TNTs, which are not attached to the substratum. The TNTs were straight connections between cells, had a typical diameter of 50 to 300 nm and a length of up to 120 µm. We observed de novo formation of TNTs by diverging from migrating cells after a short time of interaction. Scanning electron microscopy confirmed characteristic features of TNTs. Fluorescence microscopy revealed that TNTs between ARPE-19 cells contain F-actin but no microtubules. Depolymerisation of F-actin, induced by addition of latrunculin-B, led to disappearance of TNTs. Importantly, these TNTs could function as channels for the diffusion of small molecules such as Lucifer Yellow, but not for large molecules like Dextran Red. Further, organelle exchange between cells via TNTs was observed by microscopy. Using Ca²⁺ imaging we show the intercellular transmission of calcium signals through TNTs. Mechanical stimulation led to membrane depolarisation, which expand through TNT connections between ARPE-19 cells. We further demonstrate that TNTs can mediate electrical coupling between distant cells. Immunolabelling for Cx43 showed that this gap junction protein is interposed at one end of 44% of TNTs between ARPE-19 cells. CONCLUSIONS AND SIGNIFICANCE: Our observations indicate that

  4. Nonpolar interactions between trans-membrane helical EGF peptide and phosphatidylcholines, sphingomyelins and cholesterol. Molecular dynamics simulation studies

    NARCIS (Netherlands)

    Róg, T.; Murzyn, K.; Karttunen, M.E.J.; Pasenkiewicz-Gierula, M.

    2008-01-01

    A molecular dynamics simulation study of four lipid bilayers with inserted trans-membrane helical fragment of epithelial growth factor (EGF) receptor (EGF peptide) was performed. The lipid bilayers differ in their lipid composition and consist of (i) unsaturated phosphatidylcholine

  5. Correlation of Aquaporins and Transmembrane Solute Transporters Revealed by Genome-Wide Analysis in Developing Maize Leaf

    Directory of Open Access Journals (Sweden)

    Xun Yue

    2012-01-01

    Full Text Available Aquaporins are multifunctional membrane channels that facilitate the transmembrane transport of water and solutes. When transmembrane mineral nutrient transporters exhibit the same expression patterns as aquaporins under diverse temporal and physiological conditions, there is a greater probability that they interact. In this study, genome-wide temporal profiling of transcripts analysis and coexpression network-based approaches are used to examine the significant specificity correlation of aquaporins and transmembrane solute transporters in developing maize leaf. The results indicate that specific maize aquaporins are related to specific transmembrane solute transporters. The analysis demonstrates a systems-level correlation between aquaporins, nutrient transporters, and the homeostasis of mineral nutrients in developing maize leaf. Our results provide a resource for further studies into the physiological function of these aquaporins.

  6. Stomatal responses to flooding of the intercellular air spaces suggest a vapor-phase signal between the mesophyll and the guard cells.

    Science.gov (United States)

    Sibbernsen, Erik; Mott, Keith A

    2010-07-01

    Flooding the intercellular air spaces of leaves with water was shown to cause rapid closure of stomata in Tradescantia pallida, Lactuca serriola, Helianthus annuus, and Oenothera caespitosa. The response occurred when water was injected into the intercellular spaces, vacuum infiltrated into the intercellular spaces, or forced into the intercellular spaces by pressurizing the xylem. Injecting 50 mm KCl or silicone oil into the intercellular spaces also caused stomata to close, but the response was slower than with distilled water. Epidermis-mesophyll grafts for T. pallida were created by placing the epidermis of one leaf onto the exposed mesophyll of another leaf. Stomata in these grafts opened under light but closed rapidly when water was allowed to wick between epidermis and the mesophyll. When epidermis-mesophyll grafts were constructed with a thin hydrophobic filter between the mesophyll and epidermis stomata responded normally to light and CO(2). These data, when taken together, suggest that the effect of water on stomata is caused partly by dilution of K(+) in the guard cell and partly by the existence of a vapor-phase signal that originates in the mesophyll and causes stomata to open in the light.

  7. Stomatal Responses to Flooding of the Intercellular Air Spaces Suggest a Vapor-Phase Signal Between the Mesophyll and the Guard Cells1[OA

    Science.gov (United States)

    Sibbernsen, Erik; Mott, Keith A.

    2010-01-01

    Flooding the intercellular air spaces of leaves with water was shown to cause rapid closure of stomata in Tradescantia pallida, Lactuca serriola, Helianthus annuus, and Oenothera caespitosa. The response occurred when water was injected into the intercellular spaces, vacuum infiltrated into the intercellular spaces, or forced into the intercellular spaces by pressurizing the xylem. Injecting 50 mm KCl or silicone oil into the intercellular spaces also caused stomata to close, but the response was slower than with distilled water. Epidermis-mesophyll grafts for T. pallida were created by placing the epidermis of one leaf onto the exposed mesophyll of another leaf. Stomata in these grafts opened under light but closed rapidly when water was allowed to wick between epidermis and the mesophyll. When epidermis-mesophyll grafts were constructed with a thin hydrophobic filter between the mesophyll and epidermis stomata responded normally to light and CO2. These data, when taken together, suggest that the effect of water on stomata is caused partly by dilution of K+ in the guard cell and partly by the existence of a vapor-phase signal that originates in the mesophyll and causes stomata to open in the light. PMID:20472750

  8. Inhibition of luminescence and virulence in the black tiger prawn (Penaeus monodon) pathogen Vibrio harveyi by intercellular signal antagonists.

    Science.gov (United States)

    Manefield, M; Harris, L; Rice, S A; de Nys, R; Kjelleberg, S

    2000-05-01

    Expression of luminescence in the Penaeus monodon pathogen Vibrio harveyi is regulated by an intercellular quorum sensing mechanism involving the synthesis and detection of two signaling molecules, one of which is N-hydroxy butanoyl-L-homoserine lactone and the other of which is uncharacterized. Indirect evidence has suggested that virulence, associated with a toxic extracellular protein, and luminescence in V. harveyi are coregulated. In this study the effects of an acylated homoserine lactone antagonist produced by the marine alga Delisea pulchra on luminescence and toxin production in a virulent strain of V. harveyi were analyzed. Luminescence and toxin production were both inhibited by the signal antagonist at concentrations that had no impact on growth. Toxin production was found to be prematurely induced in V. harveyi cultures incubated in a 10% conditioned medium. Additionally, a significant reduction in the toxicity of concentrated supernatant extracts from V. harveyi cultures incubated in the presence of the signal antagonist, as measured by in vivo toxicity assays in mice and prawns, was observed. These results suggest that intercellular signaling antagonists have potential utility in the control of V. harveyi prawn infections.

  9. Inhibition of Luminescence and Virulence in the Black Tiger Prawn (Penaeus monodon) Pathogen Vibrio harveyi by Intercellular Signal Antagonists

    Science.gov (United States)

    Manefield, Michael; Harris, Lachlan; Rice, Scott A.; de Nys, Rocky; Kjelleberg, Staffan

    2000-01-01

    Expression of luminescence in the Penaeus monodon pathogen Vibrio harveyi is regulated by an intercellular quorum sensing mechanism involving the synthesis and detection of two signaling molecules, one of which is N-hydroxy butanoyl-l-homoserine lactone and the other of which is uncharacterized. Indirect evidence has suggested that virulence, associated with a toxic extracellular protein, and luminescence in V. harveyi are coregulated. In this study the effects of an acylated homoserine lactone antagonist produced by the marine alga Delisea pulchra on luminescence and toxin production in a virulent strain of V. harveyi were analyzed. Luminescence and toxin production were both inhibited by the signal antagonist at concentrations that had no impact on growth. Toxin production was found to be prematurely induced in V. harveyi cultures incubated in a 10% conditioned medium. Additionally, a significant reduction in the toxicity of concentrated supernatant extracts from V. harveyi cultures incubated in the presence of the signal antagonist, as measured by in vivo toxicity assays in mice and prawns, was observed. These results suggest that intercellular signaling antagonists have potential utility in the control of V. harveyi prawn infections. PMID:10788385

  10. Unified communications

    OpenAIRE

    Kravos, Urban

    2011-01-01

    In the modern business world, communication are becoming more and more complex. As a solution to this problem unified communications occurred. Using a single communication approach unified communications are the integration of various communication technologies (eg, telephony, unified messaging, audio, video and web conferencing and collaboration tools). Unified Messaging, which represents only part of the unified communications means the integration of different non real time communication t...

  11. Costimulation of N-methyl-d-aspartate and muscarinic neuronal receptors modulates gap junctional communication in striatal astrocytes

    OpenAIRE

    Rouach, N.; Tencé, M.; Glowinski, J.; Giaume, C.

    2002-01-01

    Cocultures of neurons and astrocytes from the rat striatum were used to determine whether the stimulation of neuronal receptors could affect the level of intercellular communication mediated by gap junctions in astrocytes. The costimulation of N-methyl-D-asparte (NMDA) and muscarinic receptors led to a prominent reduction of astrocyte gap junctional communication (GJC) in coculture. This treatment was not effective in astrocyte cultures, these cells being devoid of NMDA receptors. Both types ...

  12. Cellular communication through light.

    Directory of Open Access Journals (Sweden)

    Daniel Fels

    Full Text Available Information transfer is a fundamental of life. A few studies have reported that cells use photons (from an endogenous source as information carriers. This study finds that cells can have an influence on other cells even when separated with a glass barrier, thereby disabling molecule diffusion through the cell-containing medium. As there is still very little known about the potential of photons for intercellular communication this study is designed to test for non-molecule-based triggering of two fundamental properties of life: cell division and energy uptake. The study was performed with a cellular organism, the ciliate Paramecium caudatum. Mutual exposure of cell populations occurred under conditions of darkness and separation with cuvettes (vials allowing photon but not molecule transfer. The cell populations were separated either with glass allowing photon transmission from 340 nm to longer waves, or quartz being transmittable from 150 nm, i.e. from UV-light to longer waves. Even through glass, the cells affected cell division and energy uptake in neighboring cell populations. Depending on the cuvette material and the number of cells involved, these effects were positive or negative. Also, while paired populations with lower growth rates grew uncorrelated, growth of the better growing populations was correlated. As there were significant differences when separating the populations with glass or quartz, it is suggested that the cell populations use two (or more frequencies for cellular information transfer, which influences at least energy uptake, cell division rate and growth correlation. Altogether the study strongly supports a cellular communication system, which is different from a molecule-receptor-based system and hints that photon-triggering is a fine tuning principle in cell chemistry.

  13. PrP Knockout Cells Expressing Transmembrane PrP Resist Prion Infection.

    Science.gov (United States)

    Marshall, Karen E; Hughson, Andrew; Vascellari, Sarah; Priola, Suzette A; Sakudo, Akikazu; Onodera, Takashi; Baron, Gerald S

    2017-01-15

    Glycosylphosphatidylinositol (GPI) anchoring of the prion protein (PrP C ) influences PrP C misfolding into the disease-associated isoform, PrP res , as well as prion propagation and infectivity. GPI proteins are found in cholesterol- and sphingolipid-rich membrane regions called rafts. Exchanging the GPI anchor for a nonraft transmembrane sequence redirects PrP C away from rafts. Previous studies showed that nonraft transmembrane PrP C variants resist conversion to PrP res when transfected into scrapie-infected N2a neuroblastoma cells, likely due to segregation of transmembrane PrP C and GPI-anchored PrP res in distinct membrane environments. Thus, it remained unclear whether transmembrane PrP C might convert to PrP res if seeded by an exogenous source of PrP res not associated with host cell rafts and without the potential influence of endogenous expression of GPI-anchored PrP C To further explore these questions, constructs containing either a C-terminal wild-type GPI anchor signal sequence or a nonraft transmembrane sequence containing a flexible linker were expressed in a cell line derived from PrP knockout hippocampal neurons, NpL2. NpL2 cells have physiological similarities to primary neurons, representing a novel and advantageous model for studying transmissible spongiform encephalopathy (TSE) infection. Cells were infected with inocula from multiple prion strains and in different biochemical states (i.e., membrane bound as in brain microsomes from wild-type mice or purified GPI-anchorless amyloid fibrils). Only GPI-anchored PrP C supported persistent PrP res propagation. Our data provide strong evidence that in cell culture GPI anchor-directed membrane association of PrP C is required for persistent PrP res propagation, implicating raft microdomains as a location for conversion. Mechanisms of prion propagation, and what makes them transmissible, are poorly understood. Glycosylphosphatidylinositol (GPI) membrane anchoring of the prion protein (PrP C

  14. Proteomic and Functional Analyses of the Virion Transmembrane Proteome of Cyprinid Herpesvirus 3.

    Science.gov (United States)

    Vancsok, Catherine; Peñaranda, M Michelle D; Raj, V Stalin; Leroy, Baptiste; Jazowiecka-Rakus, Joanna; Boutier, Maxime; Gao, Yuan; Wilkie, Gavin S; Suárez, Nicolás M; Wattiez, Ruddy; Gillet, Laurent; Davison, Andrew J; Vanderplasschen, Alain F C

    2017-11-01

    Virion transmembrane proteins (VTPs) mediate key functions in the herpesvirus infectious cycle. Cyprinid herpesvirus 3 (CyHV-3) is the archetype of fish alloherpesviruses. The present study was devoted to CyHV-3 VTPs. Using mass spectrometry approaches, we identified 16 VTPs of the CyHV-3 FL strain. Mutagenesis experiments demonstrated that eight of these proteins are essential for viral growth in vitro (open reading frame 32 [ORF32], ORF59, ORF81, ORF83, ORF99, ORF106, ORF115, and ORF131), and eight are nonessential (ORF25, ORF64, ORF65, ORF108, ORF132, ORF136, ORF148, and ORF149). Among the nonessential proteins, deletion of ORF25, ORF132, ORF136, ORF148, or ORF149 affects viral replication in vitro , and deletion of ORF25, ORF64, ORF108, ORF132, or ORF149 impacts plaque size. Lack of ORF148 or ORF25 causes attenuation in vivo to a minor or major extent, respectively. The safety and efficacy of a virus lacking ORF25 were compared to those of a previously described vaccine candidate deleted for ORF56 and ORF57 (Δ56-57). Using quantitative PCR, we demonstrated that the ORF25 deleted virus infects fish through skin infection and then spreads to internal organs as reported previously for the wild-type parental virus and the Δ56-57 virus. However, compared to the parental wild-type virus, the replication of the ORF25-deleted virus was reduced in intensity and duration to levels similar to those observed for the Δ56-57 virus. Vaccination of fish with a virus lacking ORF25 was safe but had low efficacy at the doses tested. This characterization of the virion transmembrane proteome of CyHV-3 provides a firm basis for further research on alloherpesvirus VTPs. IMPORTANCE Virion transmembrane proteins play key roles in the biology of herpesviruses. Cyprinid herpesvirus 3 (CyHV-3) is the archetype of fish alloherpesviruses and the causative agent of major economic losses in common and koi carp worldwide. In this study of the virion transmembrane proteome of CyHV-3, the

  15. Functional Peptidomics: Combining Discovery-Based Mass Spectrometry and Neurophysiology to Explore Communication of Phase-Resetting Cues in the Rat Suprachiasmatic Nucleus

    Science.gov (United States)

    Atkins, Norman, Jr.

    2009-01-01

    Intercellular signaling is vital to communication within neuronal circuits. The suprachiasmatic nucleus (SCN), the master circadian clock of mammals, contains a dense collection of neurons that align their intrinsic rhythmicity with environmental stimulus and physiological state. While peptide physiology has been demonstrated as a contributor to…

  16. Resolving the biophysics of axon transmembrane polarization in a single closed-form description

    Energy Technology Data Exchange (ETDEWEB)

    Melendy, Robert F., E-mail: rfmelendy@liberty.edu [School of Engineering and Computational Sciences, Liberty University, Lynchburg, Virginia 24515 (United States)

    2015-12-28

    When a depolarizing event occurs across a cell membrane there is a remarkable change in its electrical properties. A complete depolarization event produces a considerably rapid increase in voltage that propagates longitudinally along the axon and is accompanied by changes in axial conductance. A dynamically changing magnetic field is associated with the passage of the action potential down the axon. Over 75 years of research has gone into the quantification of this phenomenon. To date, no unified model exist that resolves transmembrane polarization in a closed-form description. Here, a simple but formative description of propagated signaling phenomena in the membrane of an axon is presented in closed-form. The focus is on using both biophysics and mathematical methods for elucidating the fundamental mechanisms governing transmembrane polarization. The results presented demonstrate how to resolve electromagnetic and thermodynamic factors that govern transmembrane potential. Computational results are supported by well-established quantitative descriptions of propagated signaling phenomena in the membrane of an axon. The findings demonstrate how intracellular conductance, the thermodynamics of magnetization, and current modulation function together in generating an action potential in a unified closed-form description. The work presented in this paper provides compelling evidence that three basic factors contribute to the propagated signaling in the membrane of an axon. It is anticipated this work will compel those in biophysics, physical biology, and in the computational neurosciences to probe deeper into the classical and quantum features of membrane magnetization and signaling. It is hoped that subsequent investigations of this sort will be advanced by the computational features of this model without having to resort to numerical methods of analysis.

  17. Functional Architecture of the Cytoplasmic Entrance to the Cystic Fibrosis Transmembrane Conductance Regulator Chloride Channel Pore.

    Science.gov (United States)

    El Hiani, Yassine; Linsdell, Paul

    2015-06-19

    As an ion channel, the cystic fibrosis transmembrane conductance regulator must form a continuous pathway for the movement of Cl(-) and other anions between the cytoplasm and the extracellular solution. Both the structure and the function of the membrane-spanning part of this pathway are well defined. In contrast, the structure of the pathway that connects the cytoplasm to the membrane-spanning regions is unknown, and functional roles for different parts of the protein forming this pathway have not been described. We used patch clamp recording and substituted cysteine accessibility mutagenesis to identify positively charged amino acid side chains that attract cytoplasmic Cl(-) ions to the inner mouth of the pore. Our results indicate that the side chains of Lys-190, Arg-248, Arg-303, Lys-370, Lys-1041, and Arg-1048, located in different intracellular loops of the protein, play important roles in the electrostatic attraction of Cl(-) ions. Mutation and covalent modification of these residues have charge-dependent effects on the rate of Cl(-) permeation, demonstrating their functional role in maximization of Cl(-) flux. Other nearby positively charged side chains were not involved in electrostatic interactions with Cl(-). The location of these Cl(-)-attractive residues suggests that cytoplasmic Cl(-) ions enter the pore via a lateral portal located between the cytoplasmic extensions to the fourth and sixth transmembrane helices; a secondary, functionally less relevant portal might exist between the extensions to the 10th and 12th transmembrane helices. These results define the cytoplasmic mouth of the pore and show how it attracts Cl(-) ions from the cytoplasm. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  18. Communications article

    KAUST Repository

    Fariborzi, Hossein

    2017-01-01

    Seamless, covert communications using a communications system integrated or incorporated within an article of clothing is described. In one embodiment, the communications system is integrated or incorporated into a shoe insole and includes a haptic

  19. Artificial Diels–Alderase based on the transmembrane protein FhuA

    Directory of Open Access Journals (Sweden)

    Hassan Osseili

    2016-06-01

    Full Text Available Copper(I and copper(II complexes were covalently linked to an engineered variant of the transmembrane protein Ferric hydroxamate uptake protein component A (FhuA ΔCVFtev. Copper(I was incorporated using an N-heterocyclic carbene (NHC ligand equipped with a maleimide group on the side arm at the imidazole nitrogen. Copper(II was attached by coordination to a terpyridyl ligand. The spacer length was varied in the back of the ligand framework. These biohybrid catalysts were shown to be active in the Diels–Alder reaction of a chalcone derivative with cyclopentadiene to preferentially give the endo product.

  20. Ligand Modulation of the Epstein-Barr Virus-induced Seven-transmembrane Receptor EBI2

    DEFF Research Database (Denmark)

    Benned-Jensen, Tau; Smethurst, Christopher; Holst, Peter Johannes

    2011-01-01

    The Epstein-Barr virus-induced receptor 2 (EBI2) is a constitutively active seven-transmembrane receptor, which was recently shown to orchestrate the positioning of B cells in the follicle. To date, no ligands, endogenously or synthetic, have been identified that modulate EBI2 activity. Here we...... with similar potency. Overexpression of EBI2 profoundly potentiated antibody-stimulated ex vivo proliferation of murine B cells compared with WT cells, whereas this was equivalently reduced for EBI2-deficient B cells. Inhibition of EBI2 constitutive activity suppressed the proliferation in all cases...

  1. A portable lipid bilayer system for environmental sensing with a transmembrane protein.

    Directory of Open Access Journals (Sweden)

    Ryuji Kawano

    Full Text Available This paper describes a portable measurement system for current signals of an ion channel that is composed of a planar lipid bilayer. A stable and reproducible lipid bilayer is formed in outdoor environments by using a droplet contact method with a micropipette. Using this system, we demonstrated that the single-channel recording of a transmembrane protein (alpha-hemolysin was achieved in the field at a high-altitude (∼3623 m. This system would be broadly applicable for obtaining environmental measurements using membrane proteins as a highly sensitive sensor.

  2. Trafficking and function of the cystic fibrosis transmembrane conductance regulator: a complex network of posttranslational modifications

    Science.gov (United States)

    McClure, Michelle L.; Barnes, Stephen; Brodsky, Jeffrey L.

    2016-01-01

    Posttranslational modifications add diversity to protein function. Throughout its life cycle, the cystic fibrosis transmembrane conductance regulator (CFTR) undergoes numerous covalent posttranslational modifications (PTMs), including glycosylation, ubiquitination, sumoylation, phosphorylation, and palmitoylation. These modifications regulate key steps during protein biogenesis, such as protein folding, trafficking, stability, function, and association with protein partners and therefore may serve as targets for therapeutic manipulation. More generally, an improved understanding of molecular mechanisms that underlie CFTR PTMs may suggest novel treatment strategies for CF and perhaps other protein conformational diseases. This review provides a comprehensive summary of co- and posttranslational CFTR modifications and their significance with regard to protein biogenesis. PMID:27474090

  3. Early vertebrate origin and diversification of small transmembrane regulators of cellular ion transport.

    Science.gov (United States)

    Pirkmajer, Sergej; Kirchner, Henriette; Lundell, Leonidas S; Zelenin, Pavel V; Zierath, Juleen R; Makarova, Kira S; Wolf, Yuri I; Chibalin, Alexander V

    2017-07-15

    Small transmembrane proteins such as FXYDs, which interact with Na + ,K + -ATPase, and the micropeptides that interact with sarco/endoplasmic reticulum Ca 2+ -ATPase play fundamental roles in regulation of ion transport in vertebrates. Uncertain evolutionary origins and phylogenetic relationships among these regulators of ion transport have led to inconsistencies in their classification across vertebrate species, thus hampering comparative studies of their functions. We discovered the first FXYD homologue in sea lamprey, a basal jawless vertebrate, which suggests small transmembrane regulators of ion transport emerged early in the vertebrate lineage. We also identified 13 gene subfamilies of FXYDs and propose a revised, phylogeny-based FXYD classification that is consistent across vertebrate species. These findings provide an improved framework for investigating physiological and pathophysiological functions of small transmembrane regulators of ion transport. Small transmembrane proteins are important for regulation of cellular ion transport. The most prominent among these are members of the FXYD family (FXYD1-12), which regulate Na + ,K + -ATPase, and phospholamban, sarcolipin, myoregulin and DWORF, which regulate the sarco/endoplasmic reticulum Ca 2+ -ATPase (SERCA). FXYDs and regulators of SERCA are present in fishes, as well as terrestrial vertebrates; however, their evolutionary origins and phylogenetic relationships are obscure, thus hampering comparative physiological studies. Here we discovered that sea lamprey (Petromyzon marinus), a representative of extant jawless vertebrates (Cyclostomata), expresses an FXYD homologue, which strongly suggests that FXYDs predate the emergence of fishes and other jawed vertebrates (Gnathostomata). Using a combination of sequence-based phylogenetic analysis and conservation of local chromosome context, we determined that FXYDs markedly diversified in the lineages leading to cartilaginous fishes (Chondrichthyes) and bony

  4. Structure, function and physiological consequences of virally encoded chemokine seven transmembrane receptors

    DEFF Research Database (Denmark)

    Rosenkilde, M M; Smit, M J; Waldhoer, M

    2008-01-01

    A number of human and animal herpes viruses encode G-protein coupled receptors with seven transmembrane (7TM) segments-most of which are clearly related to human chemokine receptors. It appears, that these receptors are used by the virus for immune evasion, cellular transformation, tissue targeting...... pathogenesis is still poorly understood. Here we focus on the current knowledge of structure, function and trafficking patterns of virally encoded chemokine receptors and further address the putative roles of these receptors in virus survival and host -cell and/or -immune system modulation. Finally, we...

  5. Exploiting hydrophobicity for efficient production of transmembrane helices for structure determination by NMR spectroscopy

    DEFF Research Database (Denmark)

    Bugge, Katrine Østergaard; Steinocher, Helena; Brooks, Andrew J.

    2015-01-01

    -labeled protein. In this work, we have exploited the hydrophobic nature of membrane proteins to develop a simple and efficient production scheme for isotope-labeled single-pass transmembrane domains (TMDs) with or without intrinsically disordered regions. We have evaluated the applicability and limitations...... of the strategy using seven membrane protein variants that differ in their overall hydrophobicity and length and show a recovery for suitable variants of >70%. The developed production scheme is cost-efficient and easy to implement and has the potential to facilitate an increase in the number of structures...

  6. Transmembrane adaptor proteins in the high-affinity IgE receptor signaling

    Czech Academy of Sciences Publication Activity Database

    Dráber, Petr; Hálová, Ivana; Levi-Schaffer, F.; Dráberová, Lubica

    2012-01-01

    Roč. 2, 11.1. (2012), s. 95 ISSN 1664-3224 R&D Projects: GA MŠk 1M0506; GA ČR GA301/09/1826; GA ČR GAP302/10/1759; GA AV ČR KAN200520701 Grant - others:AV ČR(CZ) M200520901 Institutional research plan: CEZ:AV0Z50520514 Institutional support: RVO:68378050 Keywords : IgE receptor * LAT/LAT1 * LAX * NTAL/Lab/LAT2 * PAG/Cbp * mast cells * plasma membrane * transmembrane adaptor proteins Subject RIV: EB - Genetics ; Molecular Biology

  7. Emergency Communication

    Science.gov (United States)

    : Environmental Documents, Reports LANL Home Calendar Search Contacts Resources » Emergency Communication Emergency Communication Stay informed of emergencies, weather delays, closures, other alerts. Find links to

  8. Tactile Communications

    Data.gov (United States)

    National Aeronautics and Space Administration — Communication with the crew is vital and must be maintained regardless of environmental conditions and crew activity. Current spacecraft communication systems depend...

  9. In vitro and in situ intercellular adhesion molecule-1 (ICAM-1) expression by endothelial cells lining a polyester fabric.

    Science.gov (United States)

    Rémy, M; Valli, N; Brethes, D; Labrugère, C; Porté-Durrieu, M C; Dobrova, N B; Novikova, S P; Gorodkov, A J; Bordenave, L

    1999-02-01

    In order to improve long-term patency of vascular grafts, the promising concept of endothelial cell seeding is actually under investigation. Our laboratory tested a polyester coated with albumin and chitosan which permits a rapid colonization by human umbilical vein endothelial cells (HUVEC) and it seems relevant to test in vitro the expression of adhesive molecules expressed by cells with regard to the inflammatory process. We studied intercellular adhesion molecule-1 (ICAM-1) expression and focused our work on the determination of ICAM-1 sites expressed per adherent cell lining the biomaterial, thus in situ, in comparison to control HUVEC on plastic wells: the results obtained by binding experiments were correlated to flow cytometry analyses and showed that the polyester does not induce a proinflammatory state and that HUVEC covering the structure are able to respond to a stimulus.

  10. Indomethacin induced gastropathy in CD18, intercellular adhesion molecule 1, or P-selectin deficient mice

    Science.gov (United States)

    Morise, Z; Granger, D; Fuseler, J; Anderson, D; Grisham, M

    1999-01-01

    BACKGROUND—Neutrophil-endothelial cell interactions are thought to play a critical role in the pathophysiology of non-steroidal anti-inflammatory drug (NSAID) induced gastropathy.
AIMS—To optimise a mouse model of NSAID induced gastropathy and to evaluate the importance of adhesion molecules using adhesion molecule deficient mice.
METHODS—Gastropathy was induced in C57BL/6 mice or their adhesion molecule deficient counterparts via oral administration of indomethacin (20 mg/kg). Lesion scores, mucosal permeability, and histopathology were used to assess gastric mucosal injury.
RESULTS—Intragastric administration of indomethacin induced linear haemorrhagic mucosal lesions, primarily in the corpus of the stomach that were first observed at six hours. These lesions continued to develop over the next six hours with maximal lesion scores and mucosal permeabilities at 12 hours. When indomethacin was administered to mice deficient in CD18, intercellular adhesion molecule 1 (ICAM-1), or P-selectin, there were significant decreases in lesion scores compared with their C57BL/6 controls. In addition, mucosal permeabilities were found to be significantly lower in CD18 or ICAM-1 deficient mice observed at 12 hours.
CONCLUSION—Certain leucocyte and endothelial cell adhesion molecules are important determinants for full expression of indomethacin induced gastropathy. It is proposed that this modification of the mouse model may be useful for the investigation of other pathophysiological mechanisms of NSAID induced gastropathy.


Keywords: indomethacin; gastropathy; cyclooxygenase; intercellular adhesion molecule; VCAM; vascular cell adhesion molecule; P-selectin PMID:10486359

  11. Communications article

    KAUST Repository

    Fariborzi, Hossein

    2017-07-20

    Seamless, covert communications using a communications system integrated or incorporated within an article of clothing is described. In one embodiment, the communications system is integrated or incorporated into a shoe insole and includes a haptic feedback mechanism, a communications module, a flexible pressure sensor, and a battery. The communications module includes a wireless communications module for wireless communications, a wired interface for wired communications, a microcontroller, and a battery charge controller. The flexible pressure sensor can be actuated by an individual\\'s toe, for example, and communication between two communications nodes can be achieved using coded signals sent by individuals using a combination of long and short presses on the pressure sensor. In response to the presses, wireless communications modules can transmit and receive coded signals based on the presses.

  12. Investigation of intercellular salicylic acid accumulation during compatible and incompatible Arabidopsis-pseudomonas syringae interactions using a fast neutron-generated mutant allele of EDS5 identified by genetic mapping and whole-genome sequencing.

    Directory of Open Access Journals (Sweden)

    Jessie L Carviel

    Full Text Available A whole-genome sequencing technique developed to identify fast neutron-induced deletion mutations revealed that iap1-1 is a new allele of EDS5 (eds5-5. RPS2-AvrRpt2-initiated effector-triggered immunity (ETI was compromised in iap1-1/eds5-5 with respect to in planta bacterial levels and the hypersensitive response, while intra- and intercellular free salicylic acid (SA accumulation was greatly reduced, suggesting that SA contributes as both an intracellular signaling molecule and an antimicrobial agent in the intercellular space during ETI. During the compatible interaction between wild-type Col-0 and virulent Pseudomonas syringae pv. tomato (Pst, little intercellular free SA accumulated, which led to the hypothesis that Pst suppresses intercellular SA accumulation. When Col-0 was inoculated with a coronatine-deficient strain of Pst, high levels of intercellular SA accumulation were observed, suggesting that Pst suppresses intercellular SA accumulation using its phytotoxin coronatine. This work suggests that accumulation of SA in the intercellular space is an important component of basal/PAMP-triggered immunity as well as ETI to pathogens that colonize the intercellular space.

  13. Cysteine-rich peptides (CRPs) mediate diverse aspects of cell-cell communication in plant reproduction and development.

    Science.gov (United States)

    Marshall, Eleanor; Costa, Liliana M; Gutierrez-Marcos, Jose

    2011-03-01

    Cell-cell communication in plants is essential for the correct co-ordination of reproduction, growth, and development. Studies to dissect this mode of communication have previously focussed primarily on the action of plant hormones as mediators of intercellular signalling. In animals, peptide signalling is a well-documented intercellular communication system, however, relatively little is known about this system in plants. In recent years, numerous reports have emerged about small, secreted peptides controlling different aspects of plant reproduction. Interestingly, most of these peptides are cysteine-rich, and there is convincing evidence suggesting multiple roles for related cysteine-rich peptides (CRPs) as signalling factors in developmental patterning as well as during plant pathogen responses and symbiosis. In this review, we discuss how CRPs are emerging as key signalling factors in regulating multiple aspects of vegetative growth and reproductive development in plants.

  14. Transmembrane amyloid-related proteins in CSF as potential biomarkers for Alzheimer’s disease

    Directory of Open Access Journals (Sweden)

    Inmaculada eLopez-Font

    2015-06-01

    Full Text Available In the continuing search for new cerebrospinal fluid (CSF biomarkers for Alzheimer’s disease (AD, reasonable candidates are the secretase enzymes involved in the processing of the amyloid precursor protein (APP, as well as the large proteolytic cleavage fragments sAPPα and sAPPβ. The enzymatic activities of some of these secretases, such as BACE1 and TACE, have been investigated as potential AD biomarkers, and it has been assumed that these activities present in human CSF result from the soluble truncated forms of the membrane-bound enzymes. However, we and others recently identified soluble forms of BACE1 and APP in CSF containing the intracellular domains, as well as the multi-pass transmembrane presenilin-1 (PS1 and other subunits of γ-secretase. We also review recent findings that suggest that most of these soluble transmembrane proteins could display self-association properties based on hydrophobic and/or ionic interactions leading to the formation of heteromeric complexes. The oligomerization state of these potential new biomarkers needs to be taken into consideration for assessing their real potential as CSF biomarkers for AD by adequate molecular tools.

  15. Transmembrane Inhibitor of RICTOR/mTORC2 in Hematopoietic Progenitors

    Directory of Open Access Journals (Sweden)

    Dongjun Lee

    2014-11-01

    Full Text Available Central to cellular proliferative, survival, and metabolic responses is the serine/threonine kinase mTOR, which is activated in many human cancers. mTOR is present in distinct complexes that are either modulated by AKT (mTORC1 or are upstream and regulatory of it (mTORC2. Governance of mTORC2 activity is poorly understood. Here, we report a transmembrane molecule in hematopoietic progenitor cells that physically interacts with and inhibits RICTOR, an essential component of mTORC2. Upstream of mTORC2 (UT2 negatively regulates mTORC2 enzymatic activity, reducing AKTS473, PKCα, and NDRG1 phosphorylation and increasing FOXO transcriptional activity in an mTORC2-dependent manner. Modulating UT2 levels altered animal survival in a T cell acute lymphoid leukemia (T-ALL model that is known to be mTORC2 sensitive. These studies identify an inhibitory component upstream of mTORC2 in hematopoietic cells that can reduce mortality from NOTCH-induced T-ALL. A transmembrane inhibitor of mTORC2 may provide an attractive target to affect this critical cell regulatory pathway.

  16. Analysis of Light-Induced Transmembrane Ion Gradients and Membrane Potential in Photosystem I Proteoliposomes

    International Nuclear Information System (INIS)

    Pennisi, Cristian P.; Greenbaum, Elias; Yoshida, Ken

    2010-01-01

    Photosystem I (PSI) complexes can support a light-driven electrochemical gradient for protons, which is the driving force for energy-conserving reactions across biological membranes. In this work, a computational model that enables a quantitative description of the light-induced proton gradients across the membrane of PSI proteoliposomes is presented. Using a set of electrodiffusion equations, a compartmental model of a vesicle suspended in aqueous medium was studied. The light-mediated proton movement was modeled as a single proton pumping step with backpressure of the electric potential. The model fits determinations of pH obtained from PSI proteoliposomes illuminated in the presence of mediators of cyclic electron transport. The model also allows analysis of the proton gradients in relation to the transmembrane ion fluxes and electric potential. Sensitivity analysis enabled a determination of the parameters that have greater influence on steady-state levels and onset/decay rates of transmembrane pH and electric potential. This model could be used as a tool for optimizing PSI proteoliposomes for photo-electrochemical applications.

  17. Transmembrane protein diffusion in gel-supported dual-leaflet membranes.

    Science.gov (United States)

    Wang, Chih-Ying; Hill, Reghan J

    2014-11-18

    Tools to measure transmembrane-protein diffusion in lipid bilayer membranes have advanced in recent decades, providing a need for predictive theoretical models that account for interleaflet leaflet friction on tracer mobility. Here we address the fully three-dimensional flows driven by a (nonprotruding) transmembrane protein embedded in a dual-leaflet membrane that is supported above and below by soft porous supports (e.g., hydrogel or extracellular matrix), each of which has a prescribed permeability and solvent viscosity. For asymmetric configurations, i.e., supports with contrasting permeability, as realized for cells in contact with hydrogel scaffolds or culture media, the diffusion coefficient can reflect interleaflet friction. Reasonable approximations, for sufficiently large tracers on low-permeability supports, are furnished by a recent phenomenological theory from the literature. Interpreting literature data, albeit for hard-supported membranes, provides a theoretical basis for the phenomenological Stokes drag law as well as strengthening assertions that nonhydrodynamic interactions are important in supported bilayer systems, possibly leading to overestimates of the membrane/leaflet viscosity. Our theory provides a theoretical foundation for future experimental studies of tracer diffusion in gel-supported membranes.

  18. Substrate-modulated unwinding of transmembrane helices in the NSS transporter LeuT.

    Science.gov (United States)

    Merkle, Patrick S; Gotfryd, Kamil; Cuendet, Michel A; Leth-Espensen, Katrine Z; Gether, Ulrik; Loland, Claus J; Rand, Kasper D

    2018-05-01

    LeuT, a prokaryotic member of the neurotransmitter:sodium symporter (NSS) family, is an established structural model for mammalian NSS counterparts. We investigate the substrate translocation mechanism of LeuT by measuring the solution-phase structural dynamics of the transporter in distinct functional states by hydrogen/deuterium exchange mass spectrometry (HDX-MS). Our HDX-MS data pinpoint LeuT segments involved in substrate transport and reveal for the first time a comprehensive and detailed view of the dynamics associated with transition of the transporter between outward- and inward-facing configurations in a Na + - and K + -dependent manner. The results suggest that partial unwinding of transmembrane helices 1/5/6/7 drives LeuT from a substrate-bound, outward-facing occluded conformation toward an inward-facing open state. These hitherto unknown, large-scale conformational changes in functionally important transmembrane segments, observed for LeuT in detergent-solubilized form and when embedded in a native-like phospholipid bilayer, could be of physiological relevance for the translocation process.

  19. Monolayer freeze-fracture autoradiography: quantitative analysis of the transmembrane distribution of radioiodinated concanavalin A

    International Nuclear Information System (INIS)

    Fisher, K.A.

    1982-01-01

    The technique of monolayer freeze-fracture autoradiography (MONOFARG) has been developed and the principles, quantitation, and application of the method are described. Cell monolayers attached to polylysine-treated glass were freeze-fractured, shadowed, and coated with dry, Parlodion-supported Ilford L4 photographic emulsion at room temperature. Quantitative aspects of MONOFARG were examined using radioiodinated test systems. Background was routinely -4 grains/μm 2 /day, the highest overall efficiency was between 25% and 45%, and grain density and efficiency were dependent on radiation dose for iodine-125 and D-19 development. Corrected grain densities were linearly proportional to iodine-125 concentration. The method was applied to an examination of the transmembrane distribution of radioiodinated and fluoresceinated concanavalin A ( 125 I-FITC-Con-A). Human erythrocytes were labeled, column-purified, freeze-dried or freeze-fractured, autoradiographed, and examined by electron microscopy. The number of silver grains per square micrometer of unsplit single membrane was essentially identical to that of split extracellular membrane halves. These data demonstrate that 125 I-FITC-Con-A partitions exclusively with the extracellular half of the membrane upon freeze-fracturing and can be used as a quantitative marker for the fraction of extracellular split membrane halves. This method should be able to provide new information about certain transmembrane properties of biological membrane molecules and probes, as well as about the process of freeze-fracture per se

  20. Nanoporous microbead supported bilayers: stability, physical characterization, and incorporation of functional transmembrane proteins.

    Energy Technology Data Exchange (ETDEWEB)

    Davis, Ryan W. (University of New Mexico, Albuquerque, NM); Brozik, James A. (University of New Mexico, Albuquerque, NM); Brozik, Susan Marie; Cox, Jason M. (University of New Mexico, Albuquerque, NM); Lopez, Gabriel P. (University of New Mexico, Albuquerque, NM); Barrick, Todd A. (University of New Mexico, Albuquerque, NM); Flores, Adrean (University of New Mexico, Albuquerque, NM)

    2007-03-01

    The introduction of functional transmembrane proteins into supported bilayer-based biomimetic systems presents a significant challenge for biophysics. Among the various methods for producing supported bilayers, liposomal fusion offers a versatile method for the introduction of membrane proteins into supported bilayers on a variety of substrates. In this study, the properties of protein containing unilamellar phosphocholine lipid bilayers on nanoporous silica microspheres are investigated. The effects of the silica substrate, pore structure, and the substrate curvature on the stability of the membrane and the functionality of the membrane protein are determined. Supported bilayers on porous silica microspheres show a significant increase in surface area on surfaces with structures in excess of 10 nm as well as an overall decrease in stability resulting from increasing pore size and curvature. Comparison of the liposomal and detergent-mediated introduction of purified bacteriorhodopsin (bR) and the human type 3 serotonin receptor (5HT3R) are investigated focusing on the resulting protein function, diffusion, orientation, and incorporation efficiency. In both cases, functional proteins are observed; however, the reconstitution efficiency and orientation selectivity are significantly enhanced through detergent-mediated protein reconstitution. The results of these experiments provide a basis for bulk ionic and fluorescent dye-based compartmentalization assays as well as single-molecule optical and single-channel electrochemical interrogation of transmembrane proteins in a biomimetic platform.

  1. Impact of axial velocity and transmembrane pressure (TMP) on ARP filter performance

    Energy Technology Data Exchange (ETDEWEB)

    Poirier, M. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL); Burket, P. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

    2016-02-29

    The Savannah River Site (SRS) is currently treating radioactive liquid waste with the Actinide Removal Process (ARP) and the Modular Caustic Side Solvent Extraction Unit (MCU). Recently, the low filter flux through the ARP of approximately 5 gallons per minute has limited the rate at which radioactive liquid waste can be treated. Salt Batch 6 had a lower processing rate and required frequent filter cleaning. Savannah River Remediation (SRR) has a desire to understand the causes of the low filter flux and to increase ARP/MCU throughput. One potential method for increasing filter flux is to adjust the axial velocity and transmembrane pressure (TMP). SRR requested SRNL to conduct bench-scale filter tests to evaluate the effects of axial velocity and transmembrane pressure on crossflow filter flux. The objective of the testing was to determine whether increasing the axial velocity at the ARP could produce a significant increase in filter flux. The authors conducted the tests by preparing slurries containing 6.6 M sodium Salt Batch 6 supernate and 2.5 g MST/L, processing the slurry through a bench-scale crossflow filter unit at varying axial velocity and TMP, and measuring filter flux as a function of time.

  2. Different transport behaviors of NH4 (+) and NH3 in transmembrane cyclic peptide nanotubes.

    Science.gov (United States)

    Zhang, Mingming; Fan, Jianfen; Xu, Jian; Weng, Peipei; Lin, Huifang

    2016-10-01

    Two water-filled transmembrane cyclic peptide nanotubes (CPNTs) of 8×cyclo-(WL)n=4,5/POPE were chosen to investigate the dependences of the transport properties of the positive NH4 (+) and neutral NH3 on the channel radius. Molecular dynamic simulations revealed that molecular charge, size, ability to form H-bonds and channel radius all significantly influence the behaviors of NH4 (+) and NH3 in a CPNT. Higher electrostatic interactions, more H-bonds, and water-bridges were found in the NH4 (+) system, resulting in NH4 (+) meeting higher energy barriers, while NH3 can enter, exit and permeate the channels effortlessly. This work sheds a first light on the differences between the mechanisms of NH4 (+) and NH3 moving in a CPNT at an atomic level. Graphical Abstract Snapshot of the simulation system of NH4 (+)_octa-CPNT with an NH4 (+) initially positioned at one mouth of the tube, PMF profiles for single NH4 (+) ion and NH3 molecule moving through water-filled transmembrane CPNTs of 8×cyclo-(WL)n=4,5/POPE and sketch graphs of the possible H-bond forms of NH3 and NH4 (+) with the neighboring water.

  3. Low rate doses effects of gamma radiation on glycoproteins of transmembrane junctions in fibroblasts

    International Nuclear Information System (INIS)

    Bringas, J.E.; Caceres, J.L.

    1996-01-01

    Glycoproteins of trans-membrane junctions are molecules that help to bind cells with the extracellular matrix. Integrins are the most important trans-membrane molecules among others. The damage of gamma radiation on those proteins could be an important early event that causes membrane abnormalities which may lead to cell malfunction and cancer induced by radiation due to cell dissociation. Randomized blocks with 3 repetitions of mouse embryo fibroblast cultures, were irradiated with Cobalt-60 gamma rays, during 20 days. Biological damage to glycoproteins and integrins was evaluated by cellular growth and fibroblast proliferative capacity. Integrins damage was studied by isolation by column immunoaffinity chromatography migrated on SDS-Page under reducing and non reducing conditions, and inhibition of integrins extracellular matrix adhesion by monoclonal antibodies effect. The dose/rate (0.05 Gy/day-0.2 Gy/day) of gamma given to cells did not show damage evidence on glycoproteins and integrins. If damage happened, it was repaired by cells very soon, was delayed by continuous cellular division or by glycoproteins characteristic of being multiple extracellular ligatures. Bio effects became more evident with an irradiation time greater than 20 days or a high dose/rate. (authors). 6 refs

  4. Requirement of transmembrane domain for CD154 association to lipid rafts and subsequent biological events.

    Directory of Open Access Journals (Sweden)

    Nadir Benslimane

    Full Text Available Interaction of CD40 with CD154 leads to recruitment of both molecules into lipid rafts, resulting in bi-directional cell activation. The precise mechanism by which CD154 is translocated into lipid rafts and its impact on CD154 signaling remain largely unknown. Our aim is to identify the domain of CD154 facilitating its association to lipid rafts and the impact of such association on signaling events and cytokine production. Thus, we generated Jurkat cell lines expressing truncated CD154 lacking the cytoplasmic domain or chimeric CD154 in which the transmembrane domain was replaced by that of transferrin receptor I, known to be excluded from lipid rafts. Our results show that cell stimulation with soluble CD40 leads to the association of CD154 wild-type and CD154-truncated, but not CD154-chimera, with lipid rafts. This is correlated with failure of CD154-chimera to activate Akt and p38 MAP kinases, known effectors of CD154 signaling. We also found that CD154-chimera lost the ability to promote IL-2 production upon T cell stimulation with anti-CD3/CD28 and soluble CD40. These results demonstrate the implication of the transmembrane domain of CD154 in lipid raft association, and that this association is necessary for CD154-mediated Akt and p38 activation with consequent enhancement of IL-2 production.

  5. Transmembrane signal transduction by peptide hormones via family B G protein-coupled receptors

    Directory of Open Access Journals (Sweden)

    Kelly J Culhane

    2015-11-01

    Full Text Available Although family B G protein-coupled receptors (GPCRs contain only 15 members, they play key roles in transmembrane signal transduction of hormones. Family B GPCRs are drug targets for developing therapeutics for diseases ranging from metabolic to neurological disorders. Despite their importance, the molecular mechanism of activation of family B GPCRs remains largely unexplored due to the challenges in expression and purification of functional receptors to the quantity for biophysical characterization. Currently, there is no crystal structure available of a full-length family B GPCR. However, structures of key domains, including the extracellular ligand binding regions and seven-helical transmembrane regions, have been solved by X-ray crystallography and NMR, providing insights into the mechanisms of ligand recognition and selectivity, and helical arrangements within the cell membrane. Moreover, biophysical and biochemical methods have been used to explore functions, key residues for signaling, and the kinetics and dynamics of signaling processes. This review summarizes the current knowledge of the signal transduction mechanism of family B GPCRs at the molecular level and comments on the challenges and outlook for mechanistic studies of family B GPCRs.

  6. Role of the vaccinia virus O3 protein in cell entry can be fulfilled by its Sequence flexible transmembrane domain

    Energy Technology Data Exchange (ETDEWEB)

    Satheshkumar, P.S.; Chavre, James; Moss, Bernard, E-mail: bmoss@nih.gov

    2013-09-15

    The vaccinia virus O3 protein, a component of the entry–fusion complex, is encoded by all chordopoxviruses. We constructed truncation mutants and demonstrated that the transmembrane domain, which comprises two-thirds of this 35 amino acid protein, is necessary and sufficient for interaction with the entry–fusion complex and function in cell entry. Nevertheless, neither single amino acid substitutions nor alanine scanning mutagenesis revealed essential amino acids within the transmembrane domain. Moreover, replication-competent mutant viruses were generated by randomization of 10 amino acids of the transmembrane domain. Of eight unique viruses, two contained only two amino acids in common with wild type and the remainder contained one or none within the randomized sequence. Although these mutant viruses formed normal size plaques, the entry–fusion complex did not co-purify with the mutant O3 proteins suggesting a less stable interaction. Thus, despite low specific sequence requirements, the transmembrane domain is sufficient for function in entry. - Highlights: • The 35 amino acid O3 protein is required for efficient vaccinia virus entry. • The transmembrane domain of O3 is necessary and sufficient for entry. • Mutagenesis demonstrated extreme sequence flexibility compatible with function.

  7. Astrocyte physiopathology: At the crossroads of intercellular networking, inflammation and cell death.

    Science.gov (United States)

    Rossi, Daniela

    2015-07-01

    Recent breakthroughs in neuroscience have led to the awareness that we should revise our traditional mode of thinking and studying the CNS, i.e. by isolating the privileged network of "intelligent" synaptic contacts. We may instead need to contemplate all the variegate communications occurring between the different neural cell types, and centrally involving the astrocytes. Basically, it appears that a single astrocyte should be considered as a core that receives and integrates information from thousands of synapses, other glial cells and the blood vessels. In turn, it generates complex outputs that control the neural circuitry and coordinate it with the local microcirculation. Astrocytes thus emerge as the possible fulcrum of the functional homeostasis of the healthy CNS. Yet, evidence indicates that the bridging properties of the astrocytes can change in parallel with, or as a result of, the morphological, biochemical and functional alterations these cells undergo upon injury or disease. As a consequence, they have the potential to transform from supportive friends and interactive partners for neurons into noxious foes. In this review, we summarize the currently available knowledge on the contribution of astrocytes to the functioning of the CNS and what goes wrong in various pathological conditions, with a particular focus on Amyotrophic Lateral Sclerosis, Alzheimer's Disease and ischemia. The observations described convincingly demonstrate that the development and progression of several neurological disorders involve the de-regulation of a finely tuned interplay between multiple cell populations. Thus, it seems that a better understanding of the mechanisms governing the integrated communication and detrimental responses of the astrocytes as well as their impact towards the homeostasis and performance of the CNS is fundamental to open novel therapeutic perspectives. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. Effects of alpha-tocopherol on superoxide production and plasma intercellular adhesion molecule-1 and antibodies to oxidized LDL in chronic smokers

    NARCIS (Netherlands)

    Tits, van L.J.; Waart, de F.; Hak-Lemmers, H.L.M.; Heijst, P.; Graaf, de J.; Demacker, P.N.; Stalenhoef, A.F.

    2001-01-01

    Antioxidants have been postulated to exert beneficial effects in atherosclerosis. Atherosclerosis is associated with raised plasma levels of soluble intercellular adhesion molecule-1 (sICAM-1) and autoantibodies against oxidized low-density lipoprotein (oxLDL). It is not known whether antioxidants

  9. Soluble intercellular adhesion molecule 1 and flow-mediated dilatation are related to the estimated risk of coronary heart disease independently from each other

    NARCIS (Netherlands)

    Witte, D.R.; Broekmans, W.M.R.; Kardinaal, A.F.M.; Klöpping-Ketelaars, I.A.A.; Poppel, G. van; Bots, M.L.; Kluft, C.; Princen, J.M.G.

    2003-01-01

    Background: Flow mediated dilatation (FMD) of the brachial artery and soluble intercellular adhesion molecule 1 (sICAM-1) are measures of distinct functions of the endothelium, reflecting nitric oxide (NO)-mediated and pro-inflammatory status, respectively. The comparative value of the two measures

  10. Using Information Theory to Assess the Communicative Capacity of Circulating MicroRNA

    OpenAIRE

    Finn, Nnenna A.; Searles, Charles D.

    2013-01-01

    The discovery of extracellular microRNAs (miRNAs) and their transport modalities (i.e. microparticles, exosomes, proteins and lipoproteins) has sparked theories regarding their role in intercellular communication. Here, we assessed the information transfer capacity of different miRNA transport modalities in human serum by utilizing basic principles of information theory. Zipf Statistics were calculated for each of the miRNA transport modalities identified in human serum. Our analyses revealed...

  11. Strategizing Communication

    DEFF Research Database (Denmark)

    Gulbrandsen, Ib Tunby; Just, Sine Nørholm

    beyond, but not past instrumental, rational plans in order to become better able to understand and manage the concrete, incremental practices and contexts in which communication becomes strategic. Thus, we argue that although strategic communicators do (and should) make plans, a plan in itself does...... of the specific communicative disciplines and practices employed by the organization and/or its individual members, be they marketing, public relations, corporate communication, branding, public affairs or social advocacy. In all cases, strategic communicators do well to focus more on the process of communicating...... for understanding and managing strategic communication processes....

  12. Extracellular vesicle communication pathways as regulatory targets of oncogenic transformation.

    Science.gov (United States)

    Choi, Dongsic; Lee, Tae Hoon; Spinelli, Cristiana; Chennakrishnaiah, Shilpa; D'Asti, Esterina; Rak, Janusz

    2017-07-01

    Pathogenesis of human cancers bridges intracellular oncogenic driver events and their impact on intercellular communication. Among multiple mediators of this 'pathological connectivity' the role of extracellular vesicles (EVs) and their subsets (exosomes, ectosomes, oncosomes) is of particular interest for several reasons. The release of EVs from cancer cells represents a unique mechanism of regulated expulsion of bioactive molecules, a process that also mediates cell-to-cell transfer of lipids, proteins, and nucleic acids. Biological effects of these processes have been implicated in several aspects of cancer-related pathology, including tumour growth, invasion, angiogenesis, metastasis, immunity and thrombosis. Notably, the emerging evidence suggests that oncogenic mutations may impact several aspects of EV-mediated cell-cell communication including: (i) EV release rate and protein content; (ii) molecular composition of cancer EVs; (iii) the inclusion of oncogenic and mutant macromolecules in the EV cargo; (iv) EV-mediated release of genomic DNA; (v) deregulation of mechanisms responsible for EV biogenesis (vesiculome) and (vi) mechanisms of EV uptake by cancer cells. Intriguingly, EV-mediated intercellular transfer of mutant and oncogenic molecules between subpopulations of cancer cells, their indolent counterparts and stroma may exert profound biological effects that often resemble (but are not tantamount to) oncogenic transformation, including changes in cell growth, clonogenicity and angiogenic phenotype, or cause cell stress and death. However, several biological barriers likely curtail a permanent horizontal transformation of normal cells through EV-mediated mechanisms. The ongoing analysis and targeting of EV-mediated intercellular communication pathways can be viewed as a new therapeutic paradigm in cancer, while the analysis of oncogenic cargo contained in EVs released from cancer cells into biofluids is being developed for clinical use as a biomarker

  13. Microvesicle and tunneling nanotube mediated intercellular transfer of g-protein coupled receptors in cell cultures

    International Nuclear Information System (INIS)

    Guescini, M.; Leo, G.; Genedani, S.; Carone, C.; Pederzoli, F.; Ciruela, F.; Guidolin, D.; Stocchi, V.; Mantuano, M.; Borroto-Escuela, D.O.; Fuxe, K.; Agnati, L.F.

    2012-01-01

    Recent evidence shows that cells exchange collections of signals via microvesicles (MVs) and tunneling nano-tubes (TNTs). In this paper we have investigated whether in cell cultures GPCRs can be transferred by means of MVs and TNTs from a source cell to target cells. Western blot, transmission electron microscopy and gene expression analyses demonstrate that A 2A and D 2 receptors are present in released MVs. In order to further demonstrate the involvement of MVs in cell-to-cell communication we created two populations of cells (HEK293T and COS-7) transiently transfected with D 2 R-CFP or A 2A R-YFP. These two types of cells were co-cultured, and FRET analysis demonstrated simultaneously positive cells to the D 2 R-CFP and A 2A R-YFP. Fluorescence microscopy analysis also showed that GPCRs can move from one cell to another also by means of TNTs. Finally, recipient cells pre-incubated for 24 h with A 2A R positive MVs were treated with the adenosine A 2A receptor agonist CGS-21680. The significant increase in cAMP accumulation clearly demonstrated that A 2A Rs were functionally competent in target cells. These findings demonstrate that A 2A receptors capable of recognizing and decoding extracellular signals can be safely transferred via MVs from source to target cells.

  14. Microvesicle and tunneling nanotube mediated intercellular transfer of g-protein coupled receptors in cell cultures

    Energy Technology Data Exchange (ETDEWEB)

    Guescini, M. [Department of Biomolecular Sciences, University of Urbino ' Carlo Bo' , 61029 Urbino (Italy); Leo, G.; Genedani, S. [Department Biomedical Sciences, University of Modena and Reggio Emilia (Italy); Carone, C. [Department Biomedical Sciences, University of Modena and Reggio Emilia (Italy); IRCCS San Camillo Lido, Venezia (Italy); Pederzoli, F. [Department Biomedical Sciences, University of Modena and Reggio Emilia (Italy); Ciruela, F. [Departament Patologia i Terapeutica Experimental, Universitat de Barcelona (Spain); Guidolin, D. [Department of Human Anatomy and Physiology, University of Padua (Italy); Stocchi, V.; Mantuano, M. [Department of Biomolecular Sciences, University of Urbino ' Carlo Bo' , 61029 Urbino (Italy); Borroto-Escuela, D.O.; Fuxe, K. [Department of Neuroscience, Karolinska Institutet, Stockholm (Sweden); Agnati, L.F., E-mail: luigiagnati@tin.it [IRCCS San Camillo Lido, Venezia (Italy)

    2012-03-10

    Recent evidence shows that cells exchange collections of signals via microvesicles (MVs) and tunneling nano-tubes (TNTs). In this paper we have investigated whether in cell cultures GPCRs can be transferred by means of MVs and TNTs from a source cell to target cells. Western blot, transmission electron microscopy and gene expression analyses demonstrate that A{sub 2A} and D{sub 2} receptors are present in released MVs. In order to further demonstrate the involvement of MVs in cell-to-cell communication we created two populations of cells (HEK293T and COS-7) transiently transfected with D{sub 2}R-CFP or A{sub 2A}R-YFP. These two types of cells were co-cultured, and FRET analysis demonstrated simultaneously positive cells to the D{sub 2}R-CFP and A{sub 2A}R-YFP. Fluorescence microscopy analysis also showed that GPCRs can move from one cell to another also by means of TNTs. Finally, recipient cells pre-incubated for 24 h with A{sub 2A}R positive MVs were treated with the adenosine A{sub 2A} receptor agonist CGS-21680. The significant increase in cAMP accumulation clearly demonstrated that A{sub 2A}Rs were functionally competent in target cells. These findings demonstrate that A{sub 2A} receptors capable of recognizing and decoding extracellular signals can be safely transferred via MVs from source to target cells.

  15. Comparative characteristic of transmembrane currents and caffeine-induced responses of intact and irradiated small intestine smooth muscle cells

    International Nuclear Information System (INIS)

    Stepanov, Yu.V.; Gordienko, D.V.; Preobrazhenskaya, T.D.; Stepanova, L.I.; Vojtsitskij, V.M.

    1994-01-01

    A comparative investigation of transmembrane ion currents and caffeine-induced responses of single smooth muscle cells isolated from the circular layer of rat small intestine was curried out by the method of 'patch-clamp'. No reliable difference in potential-dependent and amplitude-kinetic characteristics of transmembrane ion currents in cells of intact and irradiated with dose of 3 Gy rats was revealed. In cells of irradiated animals external application of caffeine (4 mM) was not accompanied by strong quick-inactivated transient Ca 2+ -dependent potassium current as in control

  16. Coordinated movement of cytoplasmic and transmembrane domains of RyR1 upon gating.

    Directory of Open Access Journals (Sweden)

    Montserrat Samsó

    2009-04-01

    Full Text Available Ryanodine receptor type 1 (RyR1 produces spatially and temporally defined Ca2+ signals in several cell types. How signals received in the cytoplasmic domain are transmitted to the ion gate and how the channel gates are unknown. We used EGTA or neuroactive PCB 95 to stabilize the full closed or open states of RyR1. Single-channel measurements in the presence of FKBP12 indicate that PCB 95 inverts the thermodynamic stability of RyR1 and locks it in a long-lived open state whose unitary current is indistinguishable from the native open state. We analyzed two datasets of 15,625 and 18,527 frozen-hydrated RyR1-FKBP12 particles in the closed and open conformations, respectively, by cryo-electron microscopy. Their corresponding three-dimensional structures at 10.2 A resolution refine the structure surrounding the ion pathway previously identified in the closed conformation: two right-handed bundles emerging from the putative ion gate (the cytoplasmic "inner branches" and the transmembrane "inner helices". Furthermore, six of the identifiable transmembrane segments of RyR1 have similar organization to those of the mammalian Kv1.2 potassium channel. Upon gating, the distal cytoplasmic domains move towards the transmembrane domain while the central cytoplasmic domains move away from it, and also away from the 4-fold axis. Along the ion pathway, precise relocation of the inner helices and inner branches results in an approximately 4 A diameter increase of the ion gate. Whereas the inner helices of the K+ channels and of the RyR1 channel cross-correlate best with their corresponding open/closed states, the cytoplasmic inner branches, which are not observed in the K+ channels, appear to have at least as important a role as the inner helices for RyR1 gating. We propose a theoretical model whereby the inner helices, the inner branches, and the h1 densities together create an efficient novel gating mechanism for channel opening by relaxing two right

  17. CSR communication

    DEFF Research Database (Denmark)

    Golob, Ursa; Podnar, Klement; Elving, Wim

    2013-01-01

    Purpose – This paper aims to introduce the special issue on CSR communication attached to the First International CSR Communication Conference held in Amsterdam in October 2011. The aim of the introduction is also to review CSR communication papers published in scholarly journals in order to make...... a summary of the state of CSR communication knowledge. Design/methodology/approach – The existing literature on CSR communication was approached via systematic review. with a combination of conventional and summative qualitative content analysis. The final dataset contained 90 papers from two main business...... communications. The most important outlets for CSR communication-related topics are Journal of Business Ethics and Corporate Communications: An International Journal. Originality/value – This paper represents the first attempt to perform a systematic and comprehensive overview of CSR communication papers...

  18. ‘Gap Junctions and Cancer: Communicating for 50 Years’

    Science.gov (United States)

    Aasen, Trond; Mesnil, Marc; Naus, Christian C.; Lampe, Paul D.; Laird, Dale W.

    2017-01-01

    Fifty years ago, tumour cells were found to lack electrical coupling, leading to the hypothesis that loss of direct intercellular communication is commonly associated with cancer onset and progression. Subsequent studies linked this phenomenon to gap junctions composed of connexin proteins. While many studies support the notion that connexins are tumour suppressors, recent evidence suggests that, in some tumour types, they may facilitate specific stages of tumour progression through both junctional and non-junctional signalling pathways. This Timeline article highlights the milestones connecting gap junctions to cancer, and underscores important unanswered questions, controversies and therapeutic opportunities in the field. PMID:27782134

  19. Scientific communication

    Directory of Open Access Journals (Sweden)

    Aleksander Kobylarek

    2017-09-01

    Full Text Available The article tackles the problem of models of communication in science. The formal division of communication processes into oral and written does not resolve the problem of attitude. The author defines successful communication as a win-win game, based on the respect and equality of the partners, regardless of their position in the world of science. The core characteristics of the process of scientific communication are indicated , such as openness, fairness, support, and creation. The task of creating the right atmosphere for science communication belongs to moderators, who should not allow privilege and differentiation of position to affect scientific communication processes.

  20. Quantitative phosphoproteomics dissection of seven-transmembrane receptor signaling using full and biased agonists

    DEFF Research Database (Denmark)

    Christensen, Gitte L; Kelstrup, Christian D; Lyngsø, Christina

    2010-01-01

    (q)-dependent and -independent AT(1)R signaling. This study provides substantial novel insight into angiotensin II signal transduction and is the first study dissecting the differences between a full agonist and a biased agonist from a 7TMR on a systems-wide scale. Importantly, it reveals a previously unappreciated diversity......Seven-transmembrane receptors (7TMRs) signal through the well described heterotrimeric G proteins but can also activate G protein-independent signaling pathways of which the impact and complexity are less understood. The angiotensin II type 1 receptor (AT(1)R) is a prototypical 7TMR...... and quantity of Galpha(q) protein-independent signaling and uncovers novel signaling pathways. We foresee that the amount and diversity of G protein-independent signaling may be more pronounced than previously recognized for other 7TMRs as well. Quantitative mass spectrometry is a promising tool for evaluation...

  1. Fluorophores, environments, and quantification techniques in the analysis of transmembrane helix interaction using FRET.

    Science.gov (United States)

    Khadria, Ambalika S; Senes, Alessandro

    2015-07-01

    Förster resonance energy transfer (FRET) has been widely used as a spectroscopic tool in vitro to study the interactions between transmembrane (TM) helices in detergent and lipid environments. This technique has been instrumental to many studies that have greatly contributed to quantitative understanding of the physical principles that govern helix-helix interactions in the membrane. These studies have also improved our understanding of the biological role of oligomerization in membrane proteins. In this review, we focus on the combinations of fluorophores used, the membrane mimetic environments, and measurement techniques that have been applied to study model systems as well as biological oligomeric complexes in vitro. We highlight the different formalisms used to calculate FRET efficiency and the challenges associated with accurate quantification. The goal is to provide the reader with a comparative summary of the relevant literature for planning and designing FRET experiments aimed at measuring TM helix-helix associations. © 2015 Wiley Periodicals, Inc.

  2. Emerging role of cystic fibrosis transmembrane conductance regulator- an epithelial chloride channel in gastrointestinal cancers

    Institute of Scientific and Technical Information of China (English)

    Yuning Hou; Xiaoqing Guan; Zhe Yang; Chunying Li

    2016-01-01

    Cystic fibrosis transmembrane conductance regulator(CFTR), a glycoprotein with 1480 amino acids, has been well established as a chloride channel mainly expressed in the epithelial cells of various tissues and organs such as lungs, sweat glands, gastrointestinal system, and reproductive organs. Although defective CFTR leads to cystic fibrosis, a common genetic disorder in the Caucasian population, there is accumulating evidence that suggests a novel role of CFTR in various cancers, especially in gastroenterological cancers, such as pancreatic cancer and colon cancer. In this review, we summarize the emerging findings that link CFTR with various cancers, with focus on the association between CFTR defects and gastrointestinal cancers as well as the underlying mechanisms. Further study of CFTR in cancer biology may help pave a new way for the diagnosis and treatment of gastrointestinal cancers.

  3. Purification and crystallization of the cystic fibrosis transmembrane conductance regulator (CFTR).

    Science.gov (United States)

    Rosenberg, Mark F; Kamis, Alhaji Bukar; Aleksandrov, Luba A; Ford, Robert C; Riordan, John R

    2004-09-10

    The cystic fibrosis transmembrane conductance regulator (CFTR) is a membrane protein that is mutated in patients suffering from cystic fibrosis. Here we report the purification and first crystallization of wild-type human CFTR. Functional characterization of the material showed it to be highly active. Electron crystallography of negatively stained two-dimensional crystals of CFTR has revealed the overall architecture of this channel for two different conformational states. These show a strong structural homology to two conformational states of another eukaryotic ATP-binding cassette transporter, P-glycoprotein. In contrast to P-glycoprotein, however, both conformational states can be observed in the presence of a nucleotide, which may be related to the role of CFTR as an ion channel rather than a transporter. The hypothesis that the two conformations could represent the "open" and "closed" states of the channel is considered.

  4. Coarse Grained Molecular Dynamics Simulations of Transmembrane Protein-Lipid Systems

    Directory of Open Access Journals (Sweden)

    Peter Spijker

    2010-06-01

    Full Text Available Many biological cellular processes occur at the micro- or millisecond time scale. With traditional all-atom molecular modeling techniques it is difficult to investigate the dynamics of long time scales or large systems, such as protein aggregation or activation. Coarse graining (CG can be used to reduce the number of degrees of freedom in such a system, and reduce the computational complexity. In this paper the first version of a coarse grained model for transmembrane proteins is presented. This model differs from other coarse grained protein models due to the introduction of a novel angle potential as well as a hydrogen bonding potential. These new potentials are used to stabilize the backbone. The model has been validated by investigating the adaptation of the hydrophobic mismatch induced by the insertion of WALP-peptides into a lipid membrane, showing that the first step in the adaptation is an increase in the membrane thickness, followed by a tilting of the peptide.

  5. Research Advances in CKLFSF-like MARVEL Transmembrane Domain Containing Member 3.

    Science.gov (United States)

    Hu, Feng-zhan; Sheng, Zheng-zuo; Qin, Cai-peng; Xu, Tao

    2016-06-10

    CKLF-like MARVEL transmembrane domain containing member/chemokine-like factor super family member (CKLFSF/CMTM) is a novel tumor suppressor gene. CMTM3 is broadly expressed in normal human tissues and evolutionary conserved,especially in testis,spleen,and some cells of peripheral blood mononuclear cells. However,its expression is undetectable or down-regulated in most carcinoma cell lines and tissues. Restoration of CMTM3 may inhibit the proliferation,migration,and invasion of carcinoma cells. Although the exact mechanism of its anti-tumor activity remains unclear,CKLFSF3/CMTM3 is closely connected with immune system and associated with sex during tumorigenesis. The study advances of CKLFSF3/CMTM3 are elaborated in this review as CMTM3 may be a new target in the gene therapies for tumors,especially genitourinary tumors,while further studies on CMTM3 and its anti-tumor mechanisms are warranted.

  6. Deorphanizing the human transmembrane genome: A landscape of uncharacterized membrane proteins.

    Science.gov (United States)

    Babcock, Joseph J; Li, Min

    2014-01-01

    The sequencing of the human genome has fueled the last decade of work to functionally characterize genome content. An important subset of genes encodes membrane proteins, which are the targets of many drugs. They reside in lipid bilayers, restricting their endogenous activity to a relatively specialized biochemical environment. Without a reference phenotype, the application of systematic screens to profile candidate membrane proteins is not immediately possible. Bioinformatics has begun to show its effectiveness in focusing the functional characterization of orphan proteins of a particular functional class, such as channels or receptors. Here we discuss integration of experimental and bioinformatics approaches for characterizing the orphan membrane proteome. By analyzing the human genome, a landscape reference for the human transmembrane genome is provided.

  7. Transmembrane molecular transport during versus after extremely large, nanosecond electric pulses.

    Science.gov (United States)

    Smith, Kyle C; Weaver, James C

    2011-08-19

    Recently there has been intense and growing interest in the non-thermal biological effects of nanosecond electric pulses, particularly apoptosis induction. These effects have been hypothesized to result from the widespread creation of small, lipidic pores in the plasma and organelle membranes of cells (supra-electroporation) and, more specifically, ionic and molecular transport through these pores. Here we show that transport occurs overwhelmingly after pulsing. First, we show that the electrical drift distance for typical charged solutes during nanosecond pulses (up to 100 ns), even those with very large magnitudes (up to 10 MV/m), ranges from only a fraction of the membrane thickness (5 nm) to several membrane thicknesses. This is much smaller than the diameter of a typical cell (∼16 μm), which implies that molecular drift transport during nanosecond pulses is necessarily minimal. This implication is not dependent on assumptions about pore density or the molecular flux through pores. Second, we show that molecular transport resulting from post-pulse diffusion through minimum-size pores is orders of magnitude larger than electrical drift-driven transport during nanosecond pulses. While field-assisted charge entry and the magnitude of flux favor transport during nanosecond pulses, these effects are too small to overcome the orders of magnitude more time available for post-pulse transport. Therefore, the basic conclusion that essentially all transmembrane molecular transport occurs post-pulse holds across the plausible range of relevant parameters. Our analysis shows that a primary direct consequence of nanosecond electric pulses is the creation (or maintenance) of large populations of small pores in cell membranes that govern post-pulse transmembrane transport of small ions and molecules. Copyright © 2011 Elsevier Inc. All rights reserved.

  8. Can membrane-bound carotenoid pigment zeaxanthin carry out a transmembrane proton transfer?

    Science.gov (United States)

    Kupisz, Kamila; Sujak, Agnieszka; Patyra, Magdalena; Trebacz, Kazimierz; Gruszecki, Wiesław I

    2008-10-01

    Polar carotenoid pigment zeaxanthin (beta,beta-carotene-3,3'-diol) incorporated into planar lipid membranes formed with diphytanoyl phosphatidylcholine increases the specific electric resistance of the membrane from ca. 4 to 13 x 10(7) Omega cm2 (at 5 mol% zeaxanthin with respect to lipid). Such an observation is consistent with the well known effect of polar carotenoids in decreasing fluidity and structural stabilization of lipid bilayers. Zeaxanthin incorporated into the lipid membrane at 1 mol% has very small effect on the overall membrane resistance but facilitates equilibration of the transmembrane proton gradient, as demonstrated with the application of the H+-sensitive antimony electrodes. Relatively low changes in the electrical potential suggest that the equilibration process may be associated with a symport/antiport activity or with a transmembrane transfer of the molecules of acid. UV-Vis linear dichroism analysis of multibilayer formed with the same lipid-carotenoid system shows that the transition dipole moment of the pigment molecules forms a mean angle of 21 degrees with respect to the axis normal to the plane of the membrane. This means that zeaxanthin spans the membrane and tends to have its two hydroxyl groups anchored in the opposite polar zones of the membrane. Detailed FTIR analysis of beta-carotene and zeaxanthin indicates that the polyene chain of carotenoids is able to form weak hydrogen bonds with water molecules. Possible molecular mechanisms responsible for proton transport by polyenes are discussed, including direct involvement of the polyene chain in proton transfer and indirect effect of the pigment on physical properties of the membrane.

  9. Comparing side chain packing in soluble proteins, protein-protein interfaces, and transmembrane proteins.

    Science.gov (United States)

    Gaines, J C; Acebes, S; Virrueta, A; Butler, M; Regan, L; O'Hern, C S

    2018-05-01

    We compare side chain prediction and packing of core and non-core regions of soluble proteins, protein-protein interfaces, and transmembrane proteins. We first identified or created comparable databases of high-resolution crystal structures of these 3 protein classes. We show that the solvent-inaccessible cores of the 3 classes of proteins are equally densely packed. As a result, the side chains of core residues at protein-protein interfaces and in the membrane-exposed regions of transmembrane proteins can be predicted by the hard-sphere plus stereochemical constraint model with the same high prediction accuracies (>90%) as core residues in soluble proteins. We also find that for all 3 classes of proteins, as one moves away from the solvent-inaccessible core, the packing fraction decreases as the solvent accessibility increases. However, the side chain predictability remains high (80% within 30°) up to a relative solvent accessibility, rSASA≲0.3, for all 3 protein classes. Our results show that ≈40% of the interface regions in protein complexes are "core", that is, densely packed with side chain conformations that can be accurately predicted using the hard-sphere model. We propose packing fraction as a metric that can be used to distinguish real protein-protein interactions from designed, non-binding, decoys. Our results also show that cores of membrane proteins are the same as cores of soluble proteins. Thus, the computational methods we are developing for the analysis of the effect of hydrophobic core mutations in soluble proteins will be equally applicable to analyses of mutations in membrane proteins. © 2018 Wiley Periodicals, Inc.

  10. Induction of IL-1 during hemodialysis: Transmembrane passage of intact endotoxins (LPS)

    Energy Technology Data Exchange (ETDEWEB)

    Laude-Sharp, M.; Caroff, M.; Simard, L.; Pusineri, C.; Kazatchkine, M.D.; Haeffner-Cavaillon, N. (INSERM U 28, Hopital Broussais, Paris (France))

    1990-12-01

    Circulating monocytes of patients undergoing chronic hemodialysis are triggered to produce interleukin-1 (IL-1) in vivo. Intradialytic induction of IL-1 is associated with complement activation in patients dialyzed with first-use cellulose membranes. Chronic stimulation of IL-1 production occurs because of an yet unidentified mechanism in patients dialyzed with high permeability membranes. The present study demonstrates that intact bacterial lipopolysaccharide (LPS) molecules may cross cuprophan, AN69 and polysulfone membranes under in vitro conditions simulating in vivo hemodialysis. The experiments used purified LPS from Neisseria meningitidis and LPS from Pseudomonas testosteroni, a bacterial strain grown out from a clinically used dialysate. LPS were purified to homogeneity and radiolabeled. Transmembrane passage of 3H-labeled LPS was observed within the first five minutes of dialysis. A total of 0.1 to 1% of 3H-labeled LPS were recovered in the dialysate compartment after one hour of dialysis. High amounts of LPS, representing 40 to 70% of the amount originally present in the dialysate, were absorbed onto high permeability membranes. Low amounts of LPS were absorbed onto cuprophan membranes. The amount of LPS absorbed decreased with the concentration of LPS in the dialysate. LPS recovered from the blood compartment exhibited the same molecular weight as that used to contaminate the dialysate. Biochemically detectable transmembrane passage of LPS was not associated with that of material detectable using the limulus amebocyte lysate (LAL) assay. An IL-1-inducing activity was, however, detected in the blood compartment upon dialysis with high permeability membranes, as previously found by others with cuprophan membranes.

  11. Overexpression of transmembrane protein 168 in the mouse nucleus accumbens induces anxiety and sensorimotor gating deficit.

    Directory of Open Access Journals (Sweden)

    Kequan Fu

    Full Text Available Transmembrane protein 168 (TMEM168 comprises 697 amino acid residues, including some putative transmembrane domains. It is reported that TMEM168 controls methamphetamine (METH dependence in the nucleus accumbens (NAc of mice. Moreover, a strong link between METH dependence-induced adaptive changes in the brain and mood disorders has been evaluated. In the present study, we investigated the effects of accumbal TMEM168 in a battery of behavioral paradigms. The adeno-associated virus (AAV Tmem168 vector was injected into the NAc of C57BL/6J mice (NAc-TMEM mice. Subsequently, the accumbal TMEM168 mRNA was increased approximately by seven-fold when compared with the NAc-Mock mice (controls. The NAc-TMEM mice reported no change in the locomotor activity, cognitive ability, social interaction, and depression-like behaviors; however, TMEM168 overexpression enhanced anxiety in the elevated-plus maze and light/dark box test. The increased anxiety was reversed by pretreatment with the antianxiety drug diazepam (0.3 mg/kg i.p.. Moreover, the NAc-TMEM mice exhibited decreased prepulse inhibition (PPI in the startle response test, and the induced schizophrenia-like behavior was reversed by pretreatment with the antipsychotic drug risperidone (0.01 mg/kg i.p.. Furthermore, accumbal TMEM168 overexpression decreased the basal levels of extracellular GABA in the NAc and the high K+ (100 mM-stimulated GABA elevation; however, the total contents of GABA in the NAc remained unaffected. These results suggest that the TMEM168-regulated GABAergic neuronal system in the NAc might become a novel target while studying the etiology of anxiety and sensorimotor gating deficits.

  12. Induction of IL-1 during hemodialysis: Transmembrane passage of intact endotoxins (LPS)

    International Nuclear Information System (INIS)

    Laude-Sharp, M.; Caroff, M.; Simard, L.; Pusineri, C.; Kazatchkine, M.D.; Haeffner-Cavaillon, N.

    1990-01-01

    Circulating monocytes of patients undergoing chronic hemodialysis are triggered to produce interleukin-1 (IL-1) in vivo. Intradialytic induction of IL-1 is associated with complement activation in patients dialyzed with first-use cellulose membranes. Chronic stimulation of IL-1 production occurs because of an yet unidentified mechanism in patients dialyzed with high permeability membranes. The present study demonstrates that intact bacterial lipopolysaccharide (LPS) molecules may cross cuprophan, AN69 and polysulfone membranes under in vitro conditions simulating in vivo hemodialysis. The experiments used purified LPS from Neisseria meningitidis and LPS from Pseudomonas testosteroni, a bacterial strain grown out from a clinically used dialysate. LPS were purified to homogeneity and radiolabeled. Transmembrane passage of 3H-labeled LPS was observed within the first five minutes of dialysis. A total of 0.1 to 1% of 3H-labeled LPS were recovered in the dialysate compartment after one hour of dialysis. High amounts of LPS, representing 40 to 70% of the amount originally present in the dialysate, were absorbed onto high permeability membranes. Low amounts of LPS were absorbed onto cuprophan membranes. The amount of LPS absorbed decreased with the concentration of LPS in the dialysate. LPS recovered from the blood compartment exhibited the same molecular weight as that used to contaminate the dialysate. Biochemically detectable transmembrane passage of LPS was not associated with that of material detectable using the limulus amebocyte lysate (LAL) assay. An IL-1-inducing activity was, however, detected in the blood compartment upon dialysis with high permeability membranes, as previously found by others with cuprophan membranes

  13. Data communications

    International Nuclear Information System (INIS)

    Preckshot, G.G.

    1993-08-01

    The purpose of this paper is to recommend regulatory guidance for reviewers examining computer communication systems used in nuclear power plants. The recommendations cover three areas important to these communications systems: system design, communication protocols, and communication media. The first area, system design, considers three aspects of system design--questions about architecture, specific risky design elements or omissions to look for in designs being reviewed, and recommendations for multiplexed data communication systems used in safety systems. The second area reviews pertinent aspects of communication protocol design and makes recommendations for newly designed protocols or the selection of existing protocols for safety system, information display, and non-safety control system use. The third area covers communication media selection, which differs significantly from traditional wire and cable. The recommendations for communication media extend or enhance the concerns of published IEEE standards about three subjects: data rate, imported hazards and maintainability

  14. Communication, Communication, Communication! Growth through Laboratory Instructing

    Science.gov (United States)

    Peterson, Jamie J.; DeAngelo, Samantha; Mack, Nancy; Thompson, Claudia; Cooper, Jennifer; Sesma, Arturo, Jr.

    2014-01-01

    This study examined gains undergraduate students made in their communication and collaboration skills when they served as peer teachers, i.e., laboratory instructors (LIs), for a General Psychology laboratory. Self-ratings of communication and collaboration skills were completed before and after teaching the laboratory. When compared to before the…

  15. Participatory Communication

    DEFF Research Database (Denmark)

    Tufte, Thomas

    This user guide on participatory communication aims to answer the following questions: What do we mean when we say participatory communication? What are the practical implications of working with participatory communication strategies in development and social change processes? What practical exp......, tools, and experiences on how to implement participatory communications strategies. It is targeted toward government officials, World Bank staff, develompent workers in the field, and civil society....

  16. Cultural Communications.

    Science.gov (United States)

    Armas, Jose

    It is too often taken for granted that the communication process with culturally different children takes place as readily as it might with children from Anglo cultures. Most teachers receive training in verbal and formal communication skills; children come to school with nonverbal and informal communication skills. This initially can create…

  17. Stereotypes Communication

    Science.gov (United States)

    Zhang, Shuli; Deng, Dongyuan

    2009-01-01

    We live in a world, which is becoming a Global Village in which information and communication attract people's attention more than ever before. Our desire to communicate with strangers and our relationships with them depend on the degree to which we are effective in communicating with them. There are so many factors restricting or improving…

  18. Communication Speaks

    Science.gov (United States)

    Kinman, Robin Lynn

    2010-01-01

    When the author recently turned her attention to the National Council of Teachers of Mathematics (NCTM) "Principles and Standards," she was startled to see communication as key. She adjusted her teaching to meet the NCTM Communication Standard and promote communication in her classroom by providing a safe environment, developing discourse and…

  19. Ripple Communication.

    Science.gov (United States)

    Wilcox, R. Stimson

    1980-01-01

    Discusses how surface-dwelling animals use the water surface as a mode of communication by making ripple signals while they swim about. Provides information about surfaces and surface waves, ripple communication in water striders, ripple signal characteristics, sensing and orienting, other modes of communication, and evolution of ripple…

  20. What Can We Learn about Cholesterol's Transmembrane Distribution Based on Cholesterol-Induced Changes in Membrane Dipole Potential?

    DEFF Research Database (Denmark)

    Falkovich, Stanislav G.; Martinez-Seara, Hector; Nesterenko, Alexey M.

    2016-01-01

    Cholesterol is abundant in the plasma membranes of animal cells and is known to regulate a variety of membrane properties. Despite decades of research, the transmembrane distribution of cholesterol is still a matter of debate. Here we consider this outstanding issue through atomistic simulations ...

  1. A monocyte chemotaxis inhibiting factor in serum of HIV infected men shares epitopes with the HIV transmembrane protein gp41

    NARCIS (Netherlands)

    Tas, M.; Drexhage, H. A.; Goudsmit, J.

    1988-01-01

    This report describes that gp41, the transmembranous envelope protein of HIV, is able to inhibit monocyte chemotaxis (measured as FMLP-induced polarization). To study the presence of such immunosuppressive HIV env proteins in the circulation of HIV-infected men, fractions were prepared from serum

  2. Impact of biofilm accumulation on transmembrane and feed channel pressure drop: Effects of crossflow velocity, feed spacer and biodegradable nutrient

    KAUST Repository

    Dreszer, C.; Flemming, H. C.; Zwijnenburg, A.; Kruithof, J. C.; Vrouwenvelder, Johannes S.

    2014-01-01

    . As biodegradable nutrient, acetate was dosed to the feed water (1.0 and 0.25mgL-1 carbon) to enhance biofilm accumulation in the monitors. The studies showed that biofilm formation caused an increased transmembrane resistance and feed channel pressure drop

  3. Localization of the transmembrane proteoglycan syndecan-4 and its regulatory kinases in costameres of rat cardiomyocytes: a deconvolution microscopic study

    DEFF Research Database (Denmark)

    VanWinkle, W Barry; Snuggs, Mark B; De Hostos, Eugenio L

    2002-01-01

    Syndecan-4 (syn-4), a transmembrane heparan sulfate-containing proteoglycan, is unique among the four members of the syndecan family in its specific cellular localization to complex cytoskeletal adhesion sites, i.e., focal adhesions. During early phenotypic redifferentiation of neonatal cardiomyo...

  4. Characterization of the GXXXG motif in the first transmembrane segment of Japanese encephalitis virus precursor membrane (prM protein

    Directory of Open Access Journals (Sweden)

    Wu Suh-Chin

    2010-05-01

    Full Text Available Abstract The interaction between prM and E proteins in flavivirus-infected cells is a major driving force for the assembly of flavivirus particles. We used site-directed mutagenesis to study the potential role of the transmembrane domains of the prM proteins of Japanese encephalitis virus (JEV in prM-E heterodimerization as well as subviral particle formation. Alanine insertion scanning mutagenesis within the GXXXG motif in the first transmembrane segment of JEV prM protein affected the prM-E heterodimerization; its specificity was confirmed by replacing the two glycines of the GXXXG motif with alanine, leucine and valine. The GXXXG motif was found to be conserved in the JEV serocomplex viruses but not other flavivirus groups. These mutants with alanine inserted in the two prM transmembrane segments all impaired subviral particle formation in cell cultures. The prM transmembrane domains of JEV may play importation roles in prM-E heterodimerization and viral particle assembly.

  5. Selective elimination of high constitutive activity or chemokine binding in the human herpesvirus 8 encoded seven transmembrane oncogene ORF74

    DEFF Research Database (Denmark)

    Rosenkilde, M M; Kledal, T N; Holst, Peter Johannes

    2000-01-01

    Open reading frame 74 (ORF74) encoded by human herpesvirus 8 is a highly constitutively active seven transmembrane (7TM) receptor stimulated by angiogenic chemokines, e.g. growth-related oncogene-alpha, and inhibited by angiostatic chemokines e.g. interferon-gamma-inducible protein. Transgenic mice...

  6. Agonists and inverse agonists for the herpesvirus 8-encoded constitutively active seven-transmembrane oncogene product, ORF-74

    DEFF Research Database (Denmark)

    Rosenkilde, M M; Kledal, T N; Bräuner-Osborne, Hans

    1999-01-01

    A number of CXC chemokines competed with similar, nanomolar affinity against 125I-interleukin-8 (IL-8) binding to ORF-74, a constitutively active seven-transmembrane receptor encoded by human herpesvirus 8. However, in competition against 125I-labeled growth-related oncogene (GRO)-alpha, the ORF-74...

  7. What Can We Learn about Cholesterol's Transmembrane Distribution Based on Cholesterol-Induced Changes in Membrane Dipole Potential?

    Czech Academy of Sciences Publication Activity Database

    Falkovich, S. G.; Martinez-Seara, Hector; Nesterenko, A. M.; Vattulainen, I.; Gurtovenko, A. A.

    2016-01-01

    Roč. 7, č. 22 (2016), s. 4585-4590 ISSN 1948-7185 Institutional support: RVO:61388963 Keywords : membrane * cholesterol * membrane asymmetry * membrane dipole potential * transmembrane distribution Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 9.353, year: 2016

  8. Defining the transmembrane helix of M2 protein from influenza A by molecular dynamics simulations in a lipid bilayer

    NARCIS (Netherlands)

    Forrest, LR; Tieleman, DP; Sansom, MSP

    Integral membrane proteins containing at least one transmembrane (TM) alpha-helix are believed to account for between 20% and 30% of most genomes. There are several algorithms that accurately predict the number and position of TM helices within,a membrane protein sequence. However, these methods

  9. A mutual support mechanism through intercellular movement of CAPRICE and GLABRA3 can pattern the Arabidopsis root epidermis.

    Directory of Open Access Journals (Sweden)

    Natasha Saint Savage

    2008-09-01

    Full Text Available The patterning of the Arabidopsis root epidermis depends on a genetic regulatory network that operates both within and between cells. Genetic studies have identified a number of key components of this network, but a clear picture of the functional logic of the network is lacking. Here, we integrate existing genetic and biochemical data in a mathematical model that allows us to explore both the sufficiency of known network interactions and the extent to which additional assumptions about the model can account for wild-type and mutant data. Our model shows that an existing hypothesis concerning the autoregulation of WEREWOLF does not account fully for the expression patterns of components of the network. We confirm the lack of WEREWOLF autoregulation experimentally in transgenic plants. Rather, our modelling suggests that patterning depends on the movement of the CAPRICE and GLABRA3 transcriptional regulators between epidermal cells. Our combined modelling and experimental studies show that WEREWOLF autoregulation does not contribute to the initial patterning of epidermal cell fates in the Arabidopsis seedling root. In contrast to a patterning mechanism relying on local activation, we propose a mechanism based on lateral inhibition with feedback. The active intercellular movements of proteins that are central to our model underlie a mechanism for pattern formation in planar groups of cells that is centred on the mutual support of two cell fates rather than on local activation and lateral inhibition.

  10. A Wnt5 Activity Asymmetry and Intercellular Signaling via PCP Proteins Polarize Node Cells for Left-Right Symmetry Breaking.

    Science.gov (United States)

    Minegishi, Katsura; Hashimoto, Masakazu; Ajima, Rieko; Takaoka, Katsuyoshi; Shinohara, Kyosuke; Ikawa, Yayoi; Nishimura, Hiromi; McMahon, Andrew P; Willert, Karl; Okada, Yasushi; Sasaki, Hiroshi; Shi, Dongbo; Fujimori, Toshihiko; Ohtsuka, Toshihisa; Igarashi, Yasunobu; Yamaguchi, Terry P; Shimono, Akihiko; Shiratori, Hidetaka; Hamada, Hiroshi

    2017-03-13

    Polarization of node cells along the anterior-posterior axis of mouse embryos is responsible for left-right symmetry breaking. How node cells become polarized has remained unknown, however. Wnt5a and Wnt5b are expressed posteriorly relative to the node, whereas genes for Sfrp inhibitors of Wnt signaling are expressed anteriorly. Here we show that polarization of node cells is impaired in Wnt5a -/- Wnt5b -/- and Sfrp mutant embryos, and also in the presence of a uniform distribution of Wnt5a or Sfrp1, suggesting that Wnt5 and Sfrp proteins act as instructive signals in this process. The absence of planar cell polarity (PCP) core proteins Prickle1 and Prickle2 in individual cells or local forced expression of Wnt5a perturbed polarization of neighboring wild-type cells. Our results suggest that opposing gradients of Wnt5a and Wnt5b and of their Sfrp inhibitors, together with intercellular signaling via PCP proteins, polarize node cells along the anterior-posterior axis for breaking of left-right symmetry. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. cGAS-Mediated Innate Immunity Spreads Intercellularly through HIV-1 Env-Induced Membrane Fusion Sites.

    Science.gov (United States)

    Xu, Shuting; Ducroux, Aurélie; Ponnurangam, Aparna; Vieyres, Gabrielle; Franz, Sergej; Müsken, Mathias; Zillinger, Thomas; Malassa, Angelina; Ewald, Ellen; Hornung, Veit; Barchet, Winfried; Häussler, Susanne; Pietschmann, Thomas; Goffinet, Christine

    2016-10-12

    Upon sensing cytoplasmic retroviral DNA in infected cells, cyclic GMP-AMP (cGAMP) synthase (cGAS) produces the cyclic dinucleotide cGAMP, which activates STING to trigger a type I interferon (IFN) response. We find that membrane fusion-inducing contact between donor cells expressing the HIV envelope (Env) and primary macrophages endogenously expressing the HIV receptor CD4 and coreceptor enable intercellular transfer of cGAMP. This cGAMP exchange results in STING-dependent antiviral IFN responses in target macrophages and protection from HIV infection. Furthermore, under conditions allowing cell-to-cell transmission of HIV-1, infected primary T cells, but not cell-free virions, deliver cGAMP to autologous macrophages through HIV-1 Env and CD4/coreceptor-mediated membrane fusion sites and induce a STING-dependent, but cGAS-independent, IFN response in target cells. Collectively, these findings identify an infection-specific mode of horizontal transfer of cGAMP between primary immune cells that may boost antiviral responses, particularly in infected tissues in which cell-to-cell transmission of virions exceeds cell-free infection. Copyright © 2016 Elsevier Inc. All rights reserved.

  12. Effects of anisodamine on the expressions of vascular endothelial growth factor and intercellular adhesion molecule 1 in experimental infusion phlebitis.

    Science.gov (United States)

    Zhang, Zhen-Xiang; Wang, Peng; Zhang, Qiu-Shi; Pan, Xue; Zhao, Qing-Xia; Wang, Xiao-Kai

    2012-01-01

    Infusion phlebitis is the most common side effect of clinical intravenous drug therapy and several clinical studies have demonstrated that anisodamine can effectively prevent the occurrence of infusion phlebitis. This study was designed to investigate effects of anisodamine on the expressions of vascular endothelial growth factor (VEGF) and intercellular adhesion molecule 1 (ICAM-1) in a rabbit model of infusion phlebitis and to analyze the mechanisms of anisodamine effect on the prevention and treatment of experimental infusion phlebitis. Twenty-four specific pathogen-free male Japanese white rabbits were randomly assigned to the control group, the model group, the magnesium sulfate group and the anisodamine group. The rabbit model of infusion phlebitis, induced by intravenous administration, was established and expressions of VEGF and ICAM-1 were determined and contrasted with the control group treated with normal saline. We evaluated expression by histopathology, immunohistochemistry, reverse transcription-polymerase chain reaction, and Western blotting assay. Pathohistological changes of the model group were observed, such as loss of venous endothelial cells, inflammatory cell infiltration, edema and thrombus. The magnesium sulfate group and the anisodamine group showed significant protective effects on vascular congestion, inflammatory cell infiltration, proliferation, swelling of endothelium and perivascular hemorrhage. The model group showed the highest expressions of VEGF and ICAM-1 of the four groups (P 0.05). Anisodamine alleviates inflammatory damage by significantly reducing the expressions of VEGF and ICAM-1, and shows significant protective effects in an animal model of infusion phlebitis.

  13. Delivery of acid sphingomyelinase in normal and niemann-pick disease mice using intercellular adhesion molecule-1-targeted polymer nanocarriers.

    Science.gov (United States)

    Garnacho, Carmen; Dhami, Rajwinder; Simone, Eric; Dziubla, Thomas; Leferovich, John; Schuchman, Edward H; Muzykantov, Vladimir; Muro, Silvia

    2008-05-01

    Type B Niemann-Pick disease (NPD) is a multiorgan system disorder caused by a genetic deficiency of acid sphingomyelinase (ASM), for which lung is an important and challenging therapeutic target. In this study, we designed and evaluated new delivery vehicles for enzyme replacement therapy of type B NPD, consisting of polystyrene and poly(lactic-coglycolic) acid polymer nanocarriers targeted to intercellular adhesion molecule (ICAM)-1, an endothelial surface protein up-regulated in many pathologies, including type B NPD. Real-time vascular imaging using intravital microscopy and postmortem imaging of mouse organs showed rapid, uniform, and efficient binding of fluorescently labeled ICAM-1-targeted ASM nanocarriers (anti-ICAM/ASM nanocarriers) to endothelium after i.v. injection in mice. Fluorescence microscopy of lung alveoli actin, tissue histology, and 125I-albumin blood-to-lung transport showed that anti-ICAM nanocarriers cause neither detectable lung injury, nor abnormal vascular permeability in animals. Radioisotope tracing showed rapid disappearance from the circulation and enhanced accumulation of anti-ICAM/125I-ASM nanocarriers over the nontargeted naked enzyme in kidney, heart, liver, spleen, and primarily lung, both in wild-type and ASM knockout mice. These data demonstrate that ICAM-1-targeted nanocarriers may enhance enzyme replacement therapy for type B NPD and perhaps other lysosomal storage disorders.

  14. Mobility of the native Bacillus subtilis conjugative plasmid pLS20 is regulated by intercellular signaling.

    Science.gov (United States)

    Singh, Praveen K; Ramachandran, Gayetri; Ramos-Ruiz, Ricardo; Peiró-Pastor, Ramón; Abia, David; Wu, Ling J; Meijer, Wilfried J J

    2013-10-01

    Horizontal gene transfer mediated by plasmid conjugation plays a significant role in the evolution of bacterial species, as well as in the dissemination of antibiotic resistance and pathogenicity determinants. Characterization of their regulation is important for gaining insights into these features. Relatively little is known about how conjugation of Gram-positive plasmids is regulated. We have characterized conjugation of the native Bacillus subtilis plasmid pLS20. Contrary to the enterococcal plasmids, conjugation of pLS20 is not activated by recipient-produced pheromones but by pLS20-encoded proteins that regulate expression of the conjugation genes. We show that conjugation is kept in the default "OFF" state and identified the master repressor responsible for this. Activation of the conjugation genes requires relief of repression, which is mediated by an anti-repressor that belongs to the Rap family of proteins. Using both RNA sequencing methodology and genetic approaches, we have determined the regulatory effects of the repressor and anti-repressor on expression of the pLS20 genes. We also show that the activity of the anti-repressor is in turn regulated by an intercellular signaling peptide. Ultimately, this peptide dictates the timing of conjugation. The implications of this regulatory mechanism and comparison with other mobile systems are discussed.

  15. Expression of intercellular adhesion molecule-1 in UVA-irradiated human skin cells in vitro and in vivo

    International Nuclear Information System (INIS)

    Treina, G.; Scaletta, C.; Frenk, E.; Applegate, L.A.; Fourtanier, A.; Seite, S.

    1996-01-01

    Ultraviolet A (UVA) radiation represents an important oxidative stress to human skin and certain forms of oxidative stress have been shown to modulate intercellular adhesion molecule-1 (ICAM-1) expression. ICAM-1 has been shown to play an important part in many immune reactions and the perturbations of this molecule by ultraviolet radiation could have implications in many inflammatory responses. An enhancement immunohistochemical method with avidin/biotin was used for analysing the early effects of UVA radiation on human cell cultures and human skin (340-400 nm). Both in vitro and in vivo data show that ICAM-1 staining in epidermal keratinocytes, which was expressed constitutively, decreased in a UVA dose-dependent manner. The decrease was most noted at 3-6 h following UVA radiation with some ICAM-1 staining returning by 48 h post-UVA. ICAM-1 positive staining in the dermis was specific for vascular structures and was increased 24 h after UVA radiation. Cultured dermal fibroblasts exhibited ICAM-1 staining which increased slightly within 6-48 h post-UVA radiation. As epidermal ICAM-1 expression is depleted following UVA radiation and dermal expression increases due to an increase in the vascular structures, ICAM-1 provides a valuable marker following UVA radiation in human skin that can be readily measured in situ. (author)

  16. Domino-Like Intercellular Delivery of Undecylenic Acid-Conjugated Porous Silicon Nanoparticles for Deep Tumor Penetration.

    Science.gov (United States)

    Yong, Tuying; Hu, Jun; Zhang, Xiaoqiong; Li, Fuying; Yang, Hao; Gan, Lu; Yang, Xiangliang

    2016-10-05

    Improving the intratumoral distribution of anticancer agents remains the critical challenge for developing efficient cancer chemotherapy. Luminescent porous silicon nanoparticles (PSiNPs) have attracted considerable attention in the biomedical field especially in drug delivery. Here, we described the lysosomal exocytosis-mediated domino-like intercellular delivery of undecylenic acid-conjugated PSiNPs (UA-PSiNPs) for deep tumor penetration. UA-PSiNPs with significantly improved stability in physiological conditions were internalized into tumor cells by macropinocytosis-, caveolae-, and clathrin-mediated endocytosis and mainly colocalized with Golgi apparatus and lysosomes. Substantial evidence showed that UA-PSiNPs was excreted from cells via lysosomal exocytosis after cellular uptake. The exocytosed UA-PSiNPs induced a domino-like infection of adjacent cancer cells and allowed encapsulated doxorubicin (DOX) to deeply penetrate into both three-dimensional tumor spheroids and in vivo tumors. In addition, DOX-loaded UA-PSiNPs exhibited strong antitumor activity and few side effects in vivo. This study demonstrated that UA-PSiNPs as a drug carrier might be applied for deep tumor penetration, offering a new insight into the design of more efficient delivery systems of anticancer drugs.

  17. Remodelling of cellular excitation (reaction) and intercellular coupling (diffusion) by chronic atrial fibrillation represented by a reaction-diffusion system

    Science.gov (United States)

    Zhang, Henggui; Garratt, Clifford J.; Kharche, Sanjay; Holden, Arun V.

    2009-06-01

    Human atrial tissue is an excitable system, in which myocytes are excitable elements, and cell-to-cell electrotonic interactions are via diffusive interactions of cell membrane potentials. We developed a family of excitable system models for human atrium at cellular, tissue and anatomical levels for both normal and chronic atrial fibrillation (AF) conditions. The effects of AF-induced remodelling of cell membrane ionic channels (reaction kinetics) and intercellular gap junctional coupling (diffusion) on atrial excitability, conduction of excitation waves and dynamics of re-entrant excitation waves are quantified. Both ionic channel and gap junctional coupling remodelling have rate dependent effects on atrial propagation. Membrane channel conductance remodelling allows the propagation of activity at higher rates than those sustained in normal tissue or in tissue with gap junctional remodelling alone. Membrane channel conductance remodelling is essential for the propagation of activity at rates higher than 300/min as seen in AF. Spatially heterogeneous gap junction coupling remodelling increased the risk of conduction block, an essential factor for the genesis of re-entry. In 2D and 3D anatomical models, the dynamical behaviours of re-entrant excitation waves are also altered by membrane channel modelling. This study provides insights to understand the pro-arrhythmic effects of AF-induced reaction and diffusion remodelling in atrial tissue.

  18. House dust mite induces expression of intercellular adhesion molecule-1 in EoL-1 human eosinophilic leukemic cells.

    Science.gov (United States)

    Kwon, Byoung Chul; Sohn, Myung Hyun; Kim, Kyung Won; Kim, Eun Soo; Kim, Kyu-Earn; Shin, Myeong Heon

    2007-10-01

    The house dust mite (HDM) is considered to be the most common indoor allergen associated with bronchial asthma. In this study, we investigated whether crude extract of the HDM Dermatophagoides farinae could activate human eosinophilic leukemic cells (EoL-1) to induce upregulation of cell-surface adhesion molecules. When EoL-1 cells were incubated with D. farinae extract, expression of intercellular adhesion molecule-1 (ICAM-1) significantly increased on the cell surfaces compared to cells incubated with medium alone. In contrast, surface expression of CD11b and CD49d in EoL-1 cells was not affected by D. farinae extract. In addition, pretreatment of cells with NF-kappaB inhibitor (MG-132) or JNK inhibitor (SP600125) significantly inhibited ICAM-1 expression promoted by HDM extract. However, neither p38 MAP kinase inhibitor nor MEK inhibitor prevented HDM-induced ICAM-1 expression in EoL-1 cells. These results suggest that crude extract of D. farinae induces ICAM-1 expression in EoL-1 cells through signaling pathways involving both NF-kappaB and JNK.

  19. Inflammation-Specific T1 Imaging Using Anti-Intercellular Adhesion Molecule 1 Antibody-Conjugated Gadolinium Diethylenetriaminepentaacetic Acid

    Directory of Open Access Journals (Sweden)

    Kyu-Sil Choi

    2007-03-01

    Full Text Available To examine inflammatory tissue, an initial and common symptom of various types of pathogenesis, we designed inflammation-targeted T1 contrast agents prepared by bioconjugation of gadolinium diethylenetriaminepentaacetic acid (Gd-DTPA with anti-intercellular adhesion molecule 1 (ICAM-1 antibody. The anti-ICAM-1 antibody was coupled with DTPA and was then conjugated with Gd. The specific binding of the Gd-DTPA-anti-ICAM-1 antibody complex to the ICAM-1-expressing cells was examined in the cultured endothelial cells where ICAM-1 expression was stimulated. Inflammation-specific T1 imaging was then assessed using a mouse abscess model with the 1.5-Tesla module. The Gd-DTPA-anti-ICAM-1 antibody displayed increased r1, which was two times higher than that of Gd-DTPA and showed predominant binding to cultured endothelial cells, which expressed a high level of ICAM-1. Moreover, the inflammation-specific T1 enhancement was imaged with the Gd-DTPA-anti-ICAM-1 antibody in the mouse acute inflammation model. The Gd-DTPA-anti-ICAM-1 antibody showed significantly increased vascular circulation time, which thereby offered a greater chance for its binding to the target cells. The Gd-DTPA-anti-ICAM-1 antibody displays a potential targeted T1 contrast agent specific to the inflammatory tissue that expresses ICAM-1.

  20. Staphylococcus epidermidis polysaccharide intercellular adhesin induces IL-8 expression in human astrocytes via a mechanism involving TLR2.

    LENUS (Irish Health Repository)

    Stevens, Niall T

    2009-03-01

    Staphylococcus epidermidis is an opportunistic biofilm-forming pathogen associated with neurosurgical device-related meningitis. Expression of the polysaccharide intercellular adhesin (PIA) on its surface promotes S. epidermidis biofilm formation. Here we investigated the pro-inflammatory properties of PIA against primary and transformed human astrocytes. PIA induced IL-8 expression in a dose- and\\/or time-dependent manner from U373 MG cells and primary normal human astrocytes. This effect was inhibited by depletion of N-acetyl-beta-d-glucosamine polymer from the PIA preparation with Lycopersicon esculentum lectin or sodium meta-periodate. Expression of dominant-negative versions of the TLR2 and TLR4 adaptor proteins MyD88 and Mal in U373 MG cells inhibited PIA-induced IL-8 production. Blocking IL-1 had no effect. PIA failed to induce IL-8 production from HEK293 cells stably expressing TLR4. However, in U373 MG cells which express TLR2, neutralization of TLR2 impaired PIA-induced IL-8 production. In addition to IL-8, PIA also induced expression of other cytokines from U373 MG cells including IL-6 and MCP-1. These data implicate PIA as an important immunogenic component of the S. epidermidis biofilm that can regulate pro-inflammatory cytokine production from human astrocytes, in part, via TLR2.

  1. The Prognostic Value of Soluble Intercellular Adhesion Molecule 1 Plasma Level in Children With Acute Lung Injury.

    Science.gov (United States)

    Al-Biltagi, Mohammed A; Abo-Elezz, Ahmed Ahmed Abd ElBasset; Abu-Ela, Khaled Talaat; Suliman, Ghada Abudelmomen; Sultan, Tamer Gomaa Hassan

    2017-06-01

    The objective of this study was to evaluate the prognostic significance of soluble intercellular adhesion molecule 1 (sICAM-1) measurement in plasma for the prediction of outcome of acute lung injury (ALI) in children that may allow early recognition of critical cases. The study was performed as a prospective, controlled cohort study involving 40 children with ALI and 30 healthy children. The plasma level of sICAM-1 was measured at days 1 and 3 of development of ALI for the patient group and measured only once for the control group. C-Reactive protein was measured in both groups on day 1 only. There was significant increase in sICAM-1 in the patient group than in the control group ( P = .001*). The mortality rate reached 55% in children with ALI. The ceased group had significantly higher plasma sICAM-1 levels both at days 1 and 3 than the survived group ( P < .001*), and there was positive correlation between plasma sICAM-1 level and both duration of mechanical ventilation and the death rate, but more significant correlation was observed with plasma sICAM-1 levels at day 3 than day 1. Plasma sICAM-1 level served as a good predictor biomarker for both mechanical ventilation duration and the mortality risk in children with ALI.

  2. Detection of cancerous biological tissue areas by means of infrared absorption and SERS spectroscopy of intercellular fluid

    Science.gov (United States)

    Velicka, M.; Urboniene, V.; Ceponkus, J.; Pucetaite, M.; Jankevicius, F.; Sablinskas, V.

    2015-08-01

    We present a novel approach to the detection of cancerous kidney tissue areas by measuring vibrational spectra (IR absorption or SERS) of intercellular fluid taken from the tissue. The method is based on spectral analysis of cancerous and normal tissue areas in order to find specific spectral markers. The samples were prepared by sliding the kidney tissue over a substrate - surface of diamond ATR crystal in case of IR absorption or calcium fluoride optical window in case of SERS. For producing the SERS signal the dried fluid film was covered by silver nanoparticle colloidal solution. In order to suppress fluorescence background the measurements were performed in the NIR spectral region with the excitation wavelength of 1064 nm. The most significant spectral differences - spectral markers - were found in the region between 400 and 1800 cm-1, where spectral bands related to various vibrations of fatty acids, glycolipids and carbohydrates are located. Spectral markers in the IR and SERS spectra are different and the methods can complement each other. Both of them have potential to be used directly during surgery. Additionally, IR absorption spectroscopy in ATR mode can be combined with waveguide probe what makes this method usable in vivo.

  3. Communication Acoustics

    DEFF Research Database (Denmark)

    Blauert, Jens

    Communication Acoustics deals with the fundamentals of those areas of acoustics which are related to modern communication technologies. Due to the advent of digital signal processing and recording in acoustics, these areas have enjoyed an enormous upswing during the last 4 decades. The book...... the book a source of valuable information for those who want to improve or refresh their knowledge in the field of communication acoustics - and to work their way deeper into it. Due to its interdisciplinary character Communication Acoustics is bound to attract readers from many different areas, such as......: acoustics, cognitive science, speech science, and communication technology....

  4. Molecular cloning and tissue-specific expression analysis of mouse spinesin, a type II transmembrane serine protease 5

    International Nuclear Information System (INIS)

    Watanabe, Yoshihisa; Okui, Akira; Mitsui, Shinichi; Kawarabuki, Kentaro; Yamaguchi, Tatsuyuki; Uemura, Hidetoshi; Yamaguchi, Nozomi

    2004-01-01

    We have previously reported novel serine proteases isolated from cDNA libraries of the human and mouse central nervous system (CNS) by PCR using degenerate oligodeoxyribonucleotide primers designed on the basis of the serine protease motifs, AAHC and DSGGP. Here we report a newly isolated serine protease from the mouse CNS. This protease is homologous (77.9% identical) to human spinesin type II transmembrane serine protease 5. Mouse spinesin (m-spinesin) is also composed of (from the N-terminus) a short cytoplasmic domain, a transmembrane domain, a stem region containing a scavenger-receptor-like domain, and a serine protease domain, as is h-spinesin. We also isolated type 1, type 2, and type 3 variant cDNAs of m-spinesin. Full-length spinesin (type 4) and type 3 contain all the domains, whereas type 1 and type 2 variants lack the cytoplasmic, transmembrane, and scavenger-receptor-like domains. Subcellular localization of the variant forms was analyzed using enhanced green fluorescent protein (EGFP) fusion proteins. EGFP-type 4 fusion protein was predominantly localized to the ER, Golgi apparatus, and plasma membrane, whereas EGFP-type 1 was localized to the cytoplasm, reflecting differential classification of m-spinesin variants into transmembrane and cytoplasmic types. We analyzed the distribution of m-spinesin variants in mouse tissues, using RT-PCR with variant-specific primer sets. Interestingly, transmembrane-type spinesin, types 3 and 4, was specifically expressed in the spinal cord, whereas cytoplasmic type, type 1, was expressed in multiple tissues, including the cerebrum and cerebellum. Therefore, m-spinesin variants may have distinct biological functions arising from organ-specific variant expression

  5. Data communications

    International Nuclear Information System (INIS)

    Ann, Byeong Ho; Baek, Jeong Hun

    1998-01-01

    The contents of this book are notion of data communications : summary on data communication, data transmission, data communications system, data transmission technology, data conversion, data link control and control over error of data transmission and exchange of data communications network in the first part, computer communications network architecture : data communications architecture, OSI model, lower layer of OSI model, upper layer of OSI model and distributed surroundings in the second part, data information networking : LAN, FDDI, 100 Base T, DQDB and Frame Relay in the third part, Public Network : PSDN, N-ISDN, B-ISDN in the fourth part, internet and PC communication : emulator program, Binary file, BBS, E-mail service and user on-line service in the last part.

  6. Hydrophobic interaction between contiguous residues in the S6 transmembrane segment acts as a stimuli integration node in the BK channel

    Science.gov (United States)

    Carrasquel-Ursulaez, Willy; Contreras, Gustavo F.; Sepúlveda, Romina V.; Aguayo, Daniel; González-Nilo, Fernando

    2015-01-01

    Large-conductance Ca2+- and voltage-activated K+ channel (BK) open probability is enhanced by depolarization, increasing Ca2+ concentration, or both. These stimuli activate modular voltage and Ca2+ sensors that are allosterically coupled to channel gating. Here, we report a point mutation of a phenylalanine (F380A) in the S6 transmembrane helix that, in the absence of internal Ca2+, profoundly hinders channel opening while showing only minor effects on the voltage sensor active–resting equilibrium. Interpretation of these results using an allosteric model suggests that the F380A mutation greatly increases the free energy difference between open and closed states and uncouples Ca2+ binding from voltage sensor activation and voltage sensor activation from channel opening. However, the presence of a bulky and more hydrophobic amino acid in the F380 position (F380W) increases the intrinsic open–closed equilibrium, weakening the coupling between both sensors with the pore domain. Based on these functional experiments and molecular dynamics simulations, we propose that F380 interacts with another S6 hydrophobic residue (L377) in contiguous subunits. This pair forms a hydrophobic ring important in determining the open–closed equilibrium and, like an integration node, participates in the communication between sensors and between the sensors and pore. Moreover, because of its effects on open probabilities, the F380A mutant can be used for detailed voltage sensor experiments in the presence of permeant cations. PMID:25548136

  7. Exosomal Heat Shock Proteins as New Players in Tumour Cell-to-cell Communication

    Directory of Open Access Journals (Sweden)

    Claudia Campanella

    2014-06-01

    Full Text Available Exosomes have recently been proposed as novel elements in the study of intercellular communication in normal and pathological conditions. The biomolecular composition of exosomes reflects the specialized functions of the original cells. Heat shock proteins (Hsps are a group of chaperone proteins with diverse biological roles. In recent years, many studies have focused on the extracellular roles played by Hsps that appear to be involved in cancer development and immune system stimulation. Hsps localized on the surface of exosomes, secreted by normal and tumour cells, could be key players in intercellular cross-talk, particularly during the course of different diseases, such as cancer. Exosomal Hsps offer significant opportunities for clinical applications, including their use as potential novel biomarkers for the diagnoses or prognoses of different diseases, or for therapeutic applications and drug delivery.

  8. Exosomal Heat Shock Proteins as New Players in Tumour Cell-to-Cell Communication

    Directory of Open Access Journals (Sweden)

    Claudia Campanella

    2014-06-01

    Full Text Available Exosomes have recently been proposed as novel elements in the study of intercellular communication in normal and pathological conditions. The biomolecular composition of exosomes reflects the specialized functions of the original cells. Heat shock proteins (Hsps are a group of chaperone proteins with diverse biological roles. In recent years, many studies have focused on the extracellular roles played by Hsps that appear to be involved in cancer development and immune system stimulation. Hsps localized on the surface of exosomes, secreted by normal and tumour cells, could be key players in intercellular cross-talk, particularly during the course of different diseases, such as cancer. Exosomal Hsps offer significant opportunities for clinical applications, including their use as potential novel biomarkers for the diagnoses or prognoses of different diseases, or for therapeutic applications and drug delivery.

  9. Transmembrane transporter expression regulated by the glucosylceramide pathway in Cryptococcus neoformans.

    Science.gov (United States)

    Singh, Arpita; Rella, Antonella; Schwacke, John; Vacchi-Suzzi, Caterina; Luberto, Chiara; Del Poeta, Maurizio

    2015-11-16

    disrupt transmembrane signaling complex, which in turn contributes to cryptococcal osmotic, pH, ion homeostasis and its pathobiology. Six genes identified from gene expression microarrays by gene set enrichment analysis and validated by RT-PCR, are membrane located and associated with the growth defect at neutral-alkaline pH due to the absence and or presence of a structurally modified GlcCer. They may be involved in the transmembrane signaling network in Cryptococcus neoformans, and therefore the pathobiology of the fungus in these conditions.

  10. Gap junction protein connexin-43 interacts directly with microtubules

    NARCIS (Netherlands)

    Giepmans, B N; Verlaan, I; Hengeveld, T; Janssen, H; Calafat, J; Falk, M M; Moolenaar, W H

    2001-01-01

    Gap junctions are specialized cell-cell junctions that mediate intercellular communication. They are composed of connexin proteins, which form transmembrane channels for small molecules [1, 2]. The C-terminal tail of connexin-43 (Cx43), the most widely expressed connexin member, has been implicated

  11. Communication theory

    DEFF Research Database (Denmark)

    Stein, Irene F.; Stelter, Reinhard

    2011-01-01

    Communication theory covers a wide variety of theories related to the communication process (Littlejohn, 1999). Communication is not simply an exchange of information, in which we have a sender and a receiver. This very technical concept of communication is clearly outdated; a human being...... is not a data processing device. In this chapter, communication is understood as a process of shared meaning-making (Bruner, 1990). Human beings interpret their environment, other people, and themselves on the basis of their dynamic interaction with the surrounding world. Meaning is essential because people...... ascribe specific meanings to their experiences, their actions in life or work, and their interactions. Meaning is reshaped, adapted, and transformed in every communication encounter. Furthermore, meaning is cocreated in dialogues or in communities of practice, such as in teams at a workplace or in school...

  12. Benzo[a]pyrene induces intercellular adhesion molecule-1 through a caveolae and aryl hydrocarbon receptor mediated pathway

    International Nuclear Information System (INIS)

    Oesterling, Elizabeth; Toborek, Michal; Hennig, Bernhard

    2008-01-01

    Toxicologic and epidemiologic studies have linked benzo[a]pyrene (B[a]P) exposure with cardiovascular diseases such as atherosclerosis. The mechanisms of action leading to these diseases have not been fully understood. One key step in the development of atherosclerosis is vascular endothelial dysfunction, which is characterized by increased adhesiveness. To determine if B[a]P could lead to increased endothelial adhesiveness, the effects of B[a]P on human endothelial cell intercellular adhesion molecule-1 (ICAM-1) expression was investigated. B[a]P was able to increase ICAM-1 protein only after pretreatment with the aryl hydrocarbon receptor (AhR) agonist β-naphthoflavone (β-NF). Knockdown of AhR by siRNA or treatment with AhR antagonist α-naphthoflavone (α-NF) eliminated the induction of ICAM-1 from B[a]P, confirming the necessity of AhR in this process. Likewise, B[a]P only increased monocyte adhesion to the vascular endothelium when cells were pretreated with β-NF. Experiments were done to define a signaling mechanism. B[a]P increased phosphorylation of MEK and p38-MAPK, and inhibitors to these proteins blunted the ICAM-1 induction. B[a]P was also able to increase AP-1 DNA binding and phosphorylation of cJun. Phosphorylation of cJun was disrupted by MEK and p38-MAPK inhibitors linking the signaling cascade. Finally, the importance of membrane microdomains, caveolae, was demonstrated by knockdown of the structural protein caveolin-1. Disruption of caveolae eliminated the B[a]P-induced ICAM-1 expression. These data suggest a possible pro-inflammatory mechanism of action of B[a]P involving caveolae, leading to increased vascular endothelial adhesiveness, and this inflammation may be a critical step in the development of B[a]P-induced atherosclerosis

  13. Activated endothelial interleukin-1beta, -6, and -8 concentrations and intercellular adhesion molecule-1 expression are attenuated by lidocaine.

    LENUS (Irish Health Repository)

    Lan, Wei

    2012-02-03

    Endothelial cells play a key role in ischemia reperfusion injury. We investigated the effects of lidocaine on activated human umbilical vein endothelial cell (HUVEC) interleukin (IL)-1beta, IL-6, and IL-8 concentrations and intercellular adhesion molecule-1 (ICAM-1) expression. HUVECs were pretreated with different concentrations of lidocaine (0 to 0.5 mg\\/mL) for 60 min, thereafter tumor necrosis factor-alpha was added at a concentration of 2.5 ng\\/mL and the cells incubated for 4 h. Supernatants were harvested, and cytokine concentrations were analyzed by enzyme-linked immunosorbent assay. Endothelial ICAM-1 expression was analyzed by using flow cytometry. Differences were assessed using analysis of variance and post hoc unpaired Student\\'s t-test where appropriate. Lidocaine (0.5 mg\\/mL) decreased IL-1beta (1.89 +\\/- 0.11 versus 4.16 +\\/- 1.27 pg\\/mL; P = 0.009), IL-6 (65.5 +\\/- 5.14 versus 162 +\\/- 11.5 pg\\/mL; P < 0.001), and IL-8 (3869 +\\/- 785 versus 14,961 +\\/- 406 pg\\/mL; P < 0.001) concentrations compared with the control. IL-1beta, IL-6, and IL-8 concentrations in HUVECs treated with clinically relevant plasma concentrations of lidocaine (0.005 mg\\/mL) were similar to control. ICAM-1 expression on lidocaine-treated (0.05 mg\\/mL) HUVECs was less than on controls (198 +\\/- 52.7 versus 298 +\\/- 50.3; Mean Channel Fluorescence; P < 0.001). Activated endothelial IL-1beta, IL-6, and IL-8 concentrations and ICAM-1 expression are attenuated only by lidocaine at concentrations larger than clinically relevant concentrations.

  14. The mechanism of lauric acid-modified protein nanocapsules escape from intercellular trafficking vesicles and its implication for drug delivery.

    Science.gov (United States)

    Jiang, Lijuan; Liang, Xin; Liu, Gan; Zhou, Yun; Ye, Xinyu; Chen, Xiuli; Miao, Qianwei; Gao, Li; Zhang, Xudong; Mei, Lin

    2018-11-01

    Protein nanocapsules have exhibited promising potential applications in the field of protein drug delivery. A major issue with various promising nano-sized biotherapeutics including protein nanocapsules is that owing to their particle size they are subject to cellular uptake via endocytosis, and become entrapped and then degraded within endolysosomes, which can significantly impair their therapeutic efficacy. In addition, many nano-sized biotherapeutics could be also sequestered by autophagosomes and degraded through the autolysosomal pathway. Thus, a limiting step in achieving an effective protein therapy is to facilitate the endosomal escape and auto-lysosomal escape to ensure cytosolic delivery of the protein drugs. Here, we prepared a protein nanocapsule based on BSA (nBSA) and the BSA nanocapsules modified with a bilayer of lauric acid (LA-nBSA) to investigate the escape effects from the endosome and autophagosome. The size distribution of nBSA and LA-nBSA analyzed using DLS presents a uniform diameter centered at 10 nm and 16 nm. The data also showed that FITC-labeled nBSA and LA-nBSA were taken up by the cells mainly through Arf-6-dependent endocytosis and Rab34-mediated macropinocytosis. In addition, LA-nBSA could efficiently escape from endosomal before the degradation in endo-lysosomes. Autophagy could also sequester the LA-nBSA through p62 autophagosome vesicles. These two types of nanocapsules underwent different intracellular destinies and lauric acid (LA) coating played a vital role in intracellular particle retention. In conclusion, the protein nanocapsules modified with LA could enhance the protein nanocapsules escape from intercellular trafficking vesicles, and protect the protein from degradation by the lysosomes.

  15. Soluble intercellular adhesion molecule-1 and interleukin-6 levels reflect endothelial dysfunction in patients with primary hypercholesterolaemia treated with atorvastatin.

    Science.gov (United States)

    Nawawi, H; Osman, N S; Annuar, R; Khalid, B A K; Yusoff, K

    2003-08-01

    Adhesion molecules and cytokines are involved in the pathogenesis of intimal injury in atherosclerosis but their relationship with endothelial function remains unclear. The objectives of this study were to examine the effects of atorvastatin on soluble adhesion molecules, interleukin-6 (IL-6) and brachial artery endothelial-dependent flow mediated dilatation (FMD) in patients with familial (FH) and non-familial hypercholesterolaemia (NFH). A total of 74 patients (27 FH and 47 NFH) were recruited. Fasting lipid profiles, soluble intercellular adhesion molecule-1 (sICAM-1), soluble vascular-cellular adhesion molecule-1 (sVCAM-1), E-selectin, IL-6 and FMD were measured at baseline, 2 weeks, 3 and 9 months post-atorvastatin treatment (FH--80 mg/day, NFH--10 mg/day). In both groups, compared to baseline, sICAM-1 levels were significantly reduced at 2 weeks, further reduced at 3 months and maintained at 9 months (P<0.0001). The IL-6 levels were significantly reduced at 3 months and 9 months compared to baseline for FH (P<0.005) and NFH (P<0.0001). In both groups, the FMD at 2 weeks was higher than baseline (P<0.005), with progressive improvement up to 9 months. FMD was negatively correlated with sICAM-1 and IL-6. In conclusion, both low and high doses of atorvastatin lead to early progressive improvement in endothelial function in patients with primary hypercholesterolaemia. sICAM-1 and IL-6 levels reflect endothelial dysfunction in these patients.

  16. Purification and characterisation of an antifungal protein, MCha-Pr, from the intercellular fluid of bitter gourd (Momordica charantia) leaves.

    Science.gov (United States)

    Zhang, Beibei; Xie, Chengjian; Wei, Yunming; Li, Jing; Yang, Xingyong

    2015-03-01

    An antifungal protein, designated MCha-Pr, was isolated from the intercellular fluid of bitter gourd (Momordica charantia) leaves during a screen for potent antimicrobial proteins from plants. The isolation procedure involved a combination of extraction, ammonium sulphate precipitation, gel filtration on Bio-Gel P-6, ion exchange chromatography on CM-Sephadex, an additional gel filtration on HiLoad 16/60 Superdex 30, and finally, HPLC on a SOURCE 5RPC column. Matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry indicated that the protein had a molecular mass of 25733.46Da. Automated Edman degradation was used to determine the N-terminal sequence of MCha-Pr, and the amino acid sequence was identified as V-E-Y-T-I-T-G-N-A-G-N-T-P-G-G. The MCha-Pr protein has some similarity to the pathogenesis-related proteins from Atropa belladonna (deadly nightshade), Solanum tuberosum (potato), Ricinus communis (castor bean), and Nicotiana tabacum (tobacco). Analysis of the circular dichroism spectra indicated that MCha-Pr predominantly contains α-helix and β-sheet structures. MCha-Pr had inhibitory effects towards a variety of fungal species and the 50% inhibition of fungal growth (IC50) for Alternaria brassicae, Cercospora personata, Fusarium oxysporum, Mucor sp., and Rhizoctonia solani are 33 μM, 42 μM, 37 μM, 40 μM, and 48 μM, respectively. In addition, this antifungal protein can inhibit the germination of A. brassicae spores at 12.5 μM. These results suggest that MCha-Pr in bitter gourd leaves plays a protective role against phytopathogens and has a wide antimicrobial spectrum. Copyright © 2014 Elsevier Inc. All rights reserved.

  17. Increased Expression of Intercellular Adhesion Molecule-1, Vascular Cellular Adhesion Molecule-1 and Leukocyte Common Antigen in Diabetic Rat Retina

    Institute of Scientific and Technical Information of China (English)

    Ningyan Bai; Shibo Tang; Jing Ma; Yan Luo; Shaofeng Lin

    2003-01-01

    Purpose: To understand the expression and distribution of intercellular adhesion molecule- 1(ICAM- 1),vascular cellular adhesion molecule- 1 (VCAM- 1)and CD45 (Leukocyte Common Antigen) in the control nondiabetic and various courses of diabetic rats retina. To explore the role of adhesion molecules (Ams) and the adhesion of leukocytes to vascular endothelial cells via Ams in diabetic retinopathy(DR).Methods: Sixty healthy adult male Wistar rats were randomly divided into diabetic groups(induced by Streptozotocin, STZ) and normal control groups. Rats in these two groups were further randomly divided into 3, 7, 14, 30, 90 and 180 days-group,including 5 rats respectively. The immunohistochemical studies of ICAM-1, VCAM-1 and CD45 were carried out in the retinal digest preparations or retinal paraffin sections, and the results were analyzed qualitatively, semi-quantitatively.Results: No positive reaction of VCAM-1 was found, and weak reactions of ICAM-1,CD45 were found in nondiabetic rats retina. The difference of 6 control groups had no statistical significance(P > 0.05). The increased ICAM-1 and CD45 staining pattern were detectable 3 days after diabetes induction, and a few VCAM-1 positive cells were observed in the retinal blood capillaries. The difference of diabetes and control is significant( P < 0.05).Following the course, the expressions of ICAM-1, VCAM-1 and CD45 were increasingly enhanced, reaching a peak at the 14th day.Conclusion: Increased expression of ICAM-1, VCAM-1 and leukocytes adhering and stacking in retinal capillaries are the very early events in DR. Coherence of expression and distribution of the three further accounts for it is the key point for the onset of DR that Ams mediates leukocytes adhesion and endothelial cell injury.

  18. Anterograde axonal transport and intercellular transfer of WGA-HRP in trigeminal-innervated sensory receptors of rat incisive papilla.

    Science.gov (United States)

    Chan, K Y; Byers, M R

    1985-04-08

    The ultrastructure and identification of WGA-HRP-labeled sensory receptors in the rat incisive papilla (the most anterior part of hard palate) were studied using semiserial thin sections. Various sensory receptors were organized according to three locations: dome region (ventral), chemosensory corpuscle region (medial to orifice of incisive canal), and lateral labium (apposing the incisive canal). In the dome region, the sensory receptors were localized in three sensory zones that were associated with surface ridges (one medial and two lateral). In each of these zones, intraepithelial receptor axons and Merkel receptors occurred in the epithelium, while simple unencapsulated corpuscles, glomerular-Meissner corpuscles, and incisive (encapsulated) corpuscles occurred in the lamina propria. In the chemosensory corpuscle region, chemosensory corpuscles and intraepithelial receptor axons were located in the epithelium, and incisive corpuscles were present in the lamina propria. In the lateral labium, only intraepithelial receptor axons were prominent. In all these sensory receptors, the preterminal axons and axon terminals were labeled with the tracer protein. In addition, some nonneuronal cells closely associated with the axon terminals were selectively labeled, e.g., terminal Schwann cells, lamellar Schwann cells, Merkel cells, corpuscular basal cells and chemosensory cells. Other adjacent cells were not labeled, e.g., unspecialized epithelial cells, capsular cells, corpuscular sustentacular cells, and fibroblasts. In both labeled axons and cells, WGA-HRP was incorporated into vesicles, tubules, and vacuolar organelles. The specific intercellular transfer of tracer protein may indicate trophic interactions between axon terminals and support cells in sensory receptors. The specific organization of multiple sensory receptors in the rat incisive papilla may provide a useful alternative system for studying somatosensory physiology.

  19. Induction of mast cell accumulation by chymase via an enzymatic activity- and intercellular adhesion molecule-1-dependent mechanism.

    Science.gov (United States)

    Zhang, Huiyun; Wang, Junling; Wang, Ling; Zhan, Mengmeng; Li, Shigang; Fang, Zeman; Xu, Ciyan; Zheng, Yanshan; He, Shaoheng

    2018-02-01

    Chymase is a unique, abundant secretory product of mast cells and a potent chemoattractant for eosinophils, monocytes and neutrophils, but little is known of its influence on mast cell accumulation. A mouse peritoneal inflammation model, cell migration assay and flowcytometry analysis, were used to investigate the role of chymase in recruiting mast cells. Chymase increased, by up to 5.4-fold, mast cell numbers in mouse peritoneum. Inhibitors of chymase, heat-inactivation of the enzyme, sodium cromoglycate and terfenadine, and pretreatment of mice with anti-intercellular adhesion molecule 1, anti-L-selectin, anti-CD11a and anti-CD18 antibodies dramatically diminished the chymase-induced increase in mast cell accumulation. These findings indicate that this effect of chymase is dependent on its enzymatic activity and activation of adhesion molecules. In addition, chymase provoked a significant increase in 5-HT and eotaxin release (up to 1.8- and 2.2-fold, respectively) in mouse peritoneum. Since 5-HT, eotaxin and RANTES can induce marked mast cell accumulation, these indirect mechanisms may also contribute to chymase-induced mast cell accumulation. Moreover, chymase increased the trans-endothelium migration of mast cells in vitro indicating it also acts as a chemoattractant. The finding that mast cells accumulate in response to chymase implies further that chymase is a major pro-inflammatory mediator of mast cells. This effect of chymase, a major product of mast cell granules, suggests a novel self-amplification mechanism for mast cell accumulation in allergic inflammation. Mast cell stabilizers and inhibitors of chymase may have potential as a treatment of allergic disorders. © 2017 The British Pharmacological Society.

  20. Postcultural Communication?

    DEFF Research Database (Denmark)

    Jensen, Iben

    2015-01-01

    When we as scholars use the concept of intercultural communication in its classic definition, as communication between people with different cultural backgrounds, we perpetuate the notion that national differences influence communication more than other differences; in doing so, ethnic minorities...... is presented as a postcultural prism composed by practice theory (Schatzki 1996, Reckwitz 2002, Nicolini 2012, Kemmis 2012), Intersectionality (Brah, Phoenix, Collins Rahsack) and positioning theory (Harre & Langenhove 1998)....

  1. Indolo[3,2-b]carbazole inhibits gap junctional intercellular communication in rat primary hepatocytes and acts as a potential tumor promoter

    DEFF Research Database (Denmark)

    Herrmann, Susan; Seidelin, Michel; Bisgaard, Hanne Cathrine

    2002-01-01

    Indole-3-carbinol (I3C) is a naturally occurring substance that shows anti-carcinogenic properties in animal models. Besides its clear anti-carcinogenic effects, some studies indicate that I3C may sometimes act as a tumor promoter. Indolo[3,2-b]carbazole (ICZ), which is formed in the acidic...... environment of the stomach after intake of I3C, has a similar structure to, and shares biological effects with, the well-known tumor promoter 2,3,7,8-tetrachlorodibenzo-pdioxin (TCDD). Therefore, we hypothesized that ICZ could be responsible for the potential tumor-promoting activity of I3C. The aim...

  2. Evidence that the tri-cellular metabolism of N-acetylaspartate functions as the brain's "operating system": how NAA metabolism supports meaningful intercellular frequency-encoded communications.

    Science.gov (United States)

    Baslow, Morris H

    2010-11-01

    N-acetylaspartate (NAA), an acetylated derivative of L-aspartate (Asp), and N-acetylaspartylglutamate (NAAG), a derivative of NAA and L-glutamate (Glu), are synthesized by neurons in brain. However, neurons cannot catabolize either of these substances, and so their metabolism requires the participation of two other cell types. Neurons release both NAA and NAAG to extra-cellular fluid (ECF) upon stimulation, where astrocytes, the target cells for NAAG, hydrolyze it releasing NAA back into ECF, and oligodendrocytes, the target cells for NAA, hydrolyze it releasing Asp to ECF for recycling to neurons. This sequence is unique as it is the only known amino acid metabolic cycle in brain that requires three cell types for its completion. The results of this cycling are two-fold. First, neuronal metabolic water is transported to ECF for its removal from brain. Second, the rate of neuronal activity is coupled with focal hyperemia, providing stimulated neurons with the energy required for transmission of meaningful frequency-encoded messages. In this paper, it is proposed that the tri-cellular metabolism of NAA functions as the "operating system" of the brain, and is essential for normal cognitive and motor activities. Evidence in support of this hypothesis is provided by the outcomes of two human inborn errors in NAA metabolism.

  3. Inhibition of gap junctional intercellular communication by noncoplanar polychlorinated biphenyls: Inhibitory potencies and screening for potential mode(s) of action

    Czech Academy of Sciences Publication Activity Database

    Machala, M.; Bláha, L.; Vondráček, Jan; Trosko, J. E.; Scott, J.; Upham, B. L.

    2003-01-01

    Roč. 76, č. 1 (2003), s. 102-111 ISSN 1096-6080 R&D Projects: GA MZe QC0194; GA ČR GA525/00/D101 Grant - others:National Institute of Health(US) P42 ES04911-07 Institutional research plan: CEZ:AV0Z5004920 Keywords : WB-F344 cell line * gap junction * PCB congeners Subject RIV: BO - Biophysics Impact factor: 3.067, year: 2003

  4. Tumor promoting properties of a cigarette smoke prevalent polycyclic aromatic hydrocarbon as indicated by the inhibition of gap junctional intercellular communication via phosphatidylcholine-specific phospholipase C

    Czech Academy of Sciences Publication Activity Database

    Upham, B. L.; Bláha, L.; Babica, P.; Park, J.-S.; Sovadinová, I.; Pudrith, Ch.; Rummel, A.M.; Weis, L.M.; Sai, K.; Tithof, P.K.; Gužvič, M.; Vondráček, Jan; Machala, M.; Trosko, J.E.

    2008-01-01

    Roč. 99, č. 4 (2008), s. 696-705 ISSN 1347-9032 R&D Projects: GA ČR(CZ) GA524/05/0595 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : GJIC * phospholipases * tumor promotion Subject RIV: BO - Biophysics Impact factor: 3.471, year: 2008

  5. Calmodulin/CaMKII inhibition improves intercellular communication and impulse propagation in the heart and is antiarrhythmic under conditions when fibrosis is absent

    NARCIS (Netherlands)

    Takanari, Hiroki; Bourgonje, Vincent J A; Fontes, Magda S C; Raaijmakers, Antonia J A; Driessen, Helen; Jansen, John A.; Van Der Nagel, Roel; Kok, Bart; Van Stuijvenberg, Leonie; Boulaksil, Mohamed; Takemoto, Yoshio; Yamazaki, Masatoshi; Tsuji, Yukiomi; Honjo, Haruo; Kamiya, Kaichiro; Kodama, Itsuo; Anderson, Mark E.; Van Der Heyden, Marcel A G; Van Rijen, Harold V M; van Veen, AAB; Vos, Marc A.

    2016-01-01

    Aim In healthy hearts, ventricular gap junctions are mainly composed by connexin43 (Cx43) and localize in the intercalated disc, enabling appropriate electrical coupling. In diseased hearts, Cx43 is heterogeneously down-regulated, whereas activity of calmodulin/calcium-calmodulin protein kinase II

  6. Calmodulin/CaMKII inhibition improves intercellular communication and impulse propagation in the heart and is antiarrhythmic under conditions when fibrosis is absent

    NARCIS (Netherlands)

    Takanari, H.; Bourgonje, V.J.; Fontes, M.S.; Raaijmakers, A.J.; Driessen, H.; Jansen, JA; Nagel, R. van der; Kok, B; Stuijvenberg, L. van; Boulaksil, M.; Takemoto, Y.; Yamazaki, M.; Tsuji, Y.; Honjo, H.; Kamiya, K.; Kodama, I.; Anderson, M.E.; Heyden, M.A. van der; Rijen, H.V. van; Veen, T.A. van; Vos, M.A.

    2016-01-01

    AIM: In healthy hearts, ventricular gap junctions are mainly composed by connexin43 (Cx43) and localize in the intercalated disc, enabling appropriate electrical coupling. In diseased hearts, Cx43 is heterogeneously down-regulated, whereas activity of calmodulin/calcium-calmodulin protein kinase II

  7. Modulation of gap junctional intercellular communication between human smooth muscle cells by leukocyte-derived growth factors and cytokines in relation to atherogenesis

    NARCIS (Netherlands)

    Mensink, A.

    1997-01-01


    In this thesis, the effect of leukocyte-derived growth factors and cytokines on GJIC between SMC was investigated. GJIC is regarded as an important mechanism in the control of cell growth, cell differentiation and tissue homeostasis. Disturbance of SMC growth control is regarded to be a

  8. Accelerated SDS depletion from proteins by transmembrane electrophoresis: Impacts of Joule heating.

    Science.gov (United States)

    Unterlander, Nicole; Doucette, Alan Austin

    2018-02-08

    SDS plays a key role in proteomics workflows, including protein extraction, solubilization and mass-based separations (e.g. SDS-PAGE, GELFrEE). However, SDS interferes with mass spectrometry and so it must be removed prior to analysis. We recently introduced an electrophoretic platform, termed transmembrane electrophoresis (TME), enabling extensive depletion of SDS from proteins in solution with exceptional protein yields. However, our prior TME runs required 1 h to complete, being limited by Joule heating which causes protein aggregation at higher operating currents. Here, we demonstrate effective strategies to maintain lower TME sample temperatures, permitting accelerated SDS depletion. Among these strategies, the use of a magnetic stir bar to continuously agitate a model protein system (BSA) allows SDS to be depleted below 100 ppm (>98% removal) within 10 min of TME operations, while maintaining exceptional protein recovery (>95%). Moreover, these modifications allow TME to operate without any user intervention, improving throughput and robustness of the approach. Through fits of our time-course SDS depletion curves to an exponential model, we calculate SDS depletion half-lives as low as 1.2 min. This promising electrophoretic platform should provide proteomics researchers with an effective purification strategy to enable MS characterization of SDS-containing proteins. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Effects of centrifugation on transmembrane water loss from normal and pathologic erythrocytes

    Energy Technology Data Exchange (ETDEWEB)

    Kaperonis, A.A.; Chien, S.

    1989-02-01

    Plasma /sup 125/I-albumin was used as a marker of extracellular dilution in order to study the effect of high-speed centrifugation on transmembrane water distribution in several types of human red cells, including normal (AA), hemoglobin variants (beta A, AS, SC, beta S, and SS), and those from patients with hereditary spherocytosis. SS and AA erythrocytes were also examined for changes in intracellular hemoglobin concentration of three different density fractions and with increasing duration of spin. The minimum force and duration of centrifugation required to impair water permeability were found to vary with the red cell type, the anticoagulant used (heparin or EDTA), the initial hematocrit of the sample centrifuged, as well as among the individual erythrocyte fractions within the same sample. When subjecting pathologic erythrocytes to high-speed centrifugation, the /sup 125/I-albumin dilution technique can be used to determine whether the centrifugation procedure has led to an artifactual red cell water loss and to correct for this when it does occur. An abnormal membrane susceptibility to mechanical stress was demonstrated in erythrocytes from patients with hereditary spherocytosis and several hemoglobinopathies.

  10. Fat storage-inducing transmembrane protein 2 is required for normal fat storage in adipose tissue.

    Science.gov (United States)

    Miranda, Diego A; Kim, Ji-Hyun; Nguyen, Long N; Cheng, Wang; Tan, Bryan C; Goh, Vera J; Tan, Jolene S Y; Yaligar, Jadegoud; Kn, Bhanu Prakash; Velan, S Sendhil; Wang, Hongyan; Silver, David L

    2014-04-04

    Triglycerides within the cytosol of cells are stored in a phylogenetically conserved organelle called the lipid droplet (LD). LDs can be formed at the endoplasmic reticulum, but mechanisms that regulate the formation of LDs are incompletely understood. Adipose tissue has a high capacity to form lipid droplets and store triglycerides. Fat storage-inducing transmembrane protein 2 (FITM2/FIT2) is highly expressed in adipocytes, and data indicate that FIT2 has an important role in the formation of LDs in cells, but whether FIT2 has a physiological role in triglyceride storage in adipose tissue remains unproven. Here we show that adipose-specific deficiency of FIT2 (AF2KO) in mice results in progressive lipodystrophy of white adipose depots and metabolic dysfunction. In contrast, interscapular brown adipose tissue of AF2KO mice accumulated few but large LDs without changes in cellular triglyceride levels. High fat feeding of AF2KO mice or AF2KO mice on the genetically obese ob/ob background accelerated the onset of lipodystrophy. At the cellular level, primary adipocyte precursors of white and brown adipose tissue differentiated in vitro produced fewer but larger LDs without changes in total cellular triglyceride or triglyceride biosynthesis. These data support the conclusion that FIT2 plays an essential, physiological role in fat storage in vivo.

  11. Self-Assembling Organic Nanopores as Synthetic Transmembrane Channels with Tunable Functions

    Science.gov (United States)

    Wei, Xiaoxi

    A long-standing goal in the area of supramolecular self-assembly involves the development of synthetic ion/water channels capable of mimicking the mass-transport characteristics of biological channels and pores. Few examples of artificial transmembrane channels with large lumen, high conductivity and selectivity are known. A review of pronounced biological transmembrane protein channels and some representative synthetic models have been provided in Chapter 1, followed by our discovery and initial investigation of shape-persistent oligoamide and phenylene ethynylene macrocycles as synthetic ion/water channels. In Chapter 2, the systematic structural modification of oligoamide macrocycles 1, the so-called first-generation of these shape-persistent macrocycles, has led to third-generation macrocycles 3. The third generation was found to exhibit unprecedented, strong intermolecular association in both the solid state and solution via multiple techniques including X-ray diffraction (XRD), SEM, and 1H NMR. Fluorescence spectroscopy paired with dynamic light scattering (DLS) revealed that macrocycles 3 can assemble into a singly dispersed nanotubular structure in solution. The resultant self-assembling pores consisting of 3 were examined by HPTS-LUVs assays and BLM studies (Chapter 3) and found to form cation-selective (PK+/PCl- = 69:1) transmembrane ion channels with large conductance (200 ˜ 2000 pS for alkali cations) and high stability with open times reaching to 103 seconds. Tuning the aggregation state of macrocycles by choosing an appropriate polar solvent mixture (i.e., 3:1, THF:DMF, v/v) and concentration led to the formation of ion channels with well-defined square top behavior. A parallel study using DLS to examine the size of aggregates was used in conjunction with channel activity assays (LUVs/BLM) to reveal the effects of the aggregation state on channel activity. Empirical evidence now clearly indicates that a preassembled state, perhaps that of a

  12. The position of the Gly-xxx-Gly motif in transmembrane segments modulates dimer affinity.

    Science.gov (United States)

    Johnson, Rachel M; Rath, Arianna; Deber, Charles M

    2006-12-01

    Although the intrinsic low solubility of membrane proteins presents challenges to their high-resolution structure determination, insight into the amino acid sequence features and forces that stabilize their folds has been provided through study of sequence-dependent helix-helix interactions between single transmembrane (TM) helices. While the stability of helix-helix partnerships mediated by the Gly-xxx-Gly (GG4) motif is known to be generally modulated by distal interfacial residues, it has not been established whether the position of this motif, with respect to the ends of a given TM segment, affects dimer affinity. Here we examine the relationship between motif position and affinity in the homodimers of 2 single-spanning membrane protein TM sequences: glycophorin A (GpA) and bacteriophage M13 coat protein (MCP). Using the TOXCAT assay for dimer affinity on a series of GpA and MCP TM segments that have been modified with either 4 Leu residues at each end or with 8 Leu residues at the N-terminal end, we show that in each protein, centrally located GG4 motifs are capable of stronger helix-helix interactions than those proximal to TM helix ends, even when surrounding interfacial residues are maintained. The relative importance of GG4 motifs in stabilizing helix-helix interactions therefore must be considered not only in its specific residue context but also in terms of the location of the interactive surface relative to the N and C termini of alpha-helical TM segments.

  13. Cystic fibrosis transmembrane conductance regulator intracellular processing, trafficking, and opportunities for mutation-specific treatment.

    LENUS (Irish Health Repository)

    Rogan, Mark P

    2012-02-01

    Recent advances in basic science have greatly expanded our understanding of the cystic fibrosis (CF) transmembrane conductance regulator (CFTR), the chloride and bicarbonate channel that is encoded by the gene, which is mutated in patients with CF. We review the structure, function, biosynthetic processing, and intracellular trafficking of CFTR and discuss the five classes of mutations and their impact on the CF phenotype. The therapeutic discussion is focused on the significant progress toward CFTR mutation-specific therapies. We review the results of encouraging clinical trials examining orally administered therapeutics, including agents that promote read-through of class I mutations (premature termination codons); correctors, which overcome the CFTR misfolding that characterizes the common class II mutation F508del; and potentiators, which enhance the function of class III or IV mutated CFTR at the plasma membrane. Long-term outcomes from successful mutation-specific treatments could finally answer the question that has been lingering since and even before the CFTR gene discovery: Will therapies that specifically restore CFTR-mediated chloride secretion slow or arrest the deleterious cascade of events leading to chronic infection, bronchiectasis, and end-stage lung disease?

  14. Transmembrane collagen XVII modulates integrin dependent keratinocyte migration via PI3K/Rac1 signaling.

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    Stefanie Löffek

    Full Text Available The hemidesmosomal transmembrane component collagen XVII (ColXVII plays an important role in the anchorage of the epidermis to the underlying basement membrane. However, this adhesion protein seems to be also involved in the regulation of keratinocyte migration, since its expression in these cells is strongly elevated during reepithelialization of acute wounds and in the invasive front of squamous cell carcinoma, while its absence in ColXVII-deficient keratinocytes leads to altered cell motility. Using a genetic model of murine Col17a1⁻/⁻ keratinocytes we elucidated ColXVII mediated signaling pathways in cell adhesion and migration. Col17a1⁻/⁻ keratinocytes exhibited increased spreading on laminin 332 and accelerated, but less directed cell motility. These effects were accompanied by increased expression of the integrin subunits β4 and β1. The migratory phenotype, as evidenced by formation of multiple unstable lamellipodia, was associated with enhanced phosphoinositide 3-kinase (PI3K activity. Dissection of the signaling pathway uncovered enhanced phosphorylation of the β4 integrin subunit and the focal adhesion kinase (FAK as activators of PI3K. This resulted in elevated Rac1 activity as a downstream consequence. These results provide mechanistic evidence that ColXVII coordinates keratinocyte adhesion and directed motility by interfering integrin dependent PI3K activation and by stabilizing lamellipodia at the leading edge of reepithelializing wounds and in invasive squamous cell carcinoma.

  15. Transmembrane prostatic acid phosphatase (TMPAP interacts with snapin and deficient mice develop prostate adenocarcinoma.

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    Ileana B Quintero

    Full Text Available The molecular mechanisms underlying prostate carcinogenesis are poorly understood. Prostatic acid phosphatase (PAP, a prostatic epithelial secretion marker, has been linked to prostate cancer since the 1930's. However, the contribution of PAP to the disease remains controversial. We have previously cloned and described two isoforms of this protein, a secretory (sPAP and a transmembrane type-I (TMPAP. The goal in this work was to understand the physiological function of TMPAP in the prostate. We conducted histological, ultra-structural and genome-wide analyses of the prostate of our PAP-deficient mouse model (PAP(-/- with C57BL/6J background. The PAP(-/- mouse prostate showed the development of slow-growing non-metastatic prostate adenocarcinoma. In order to find out the mechanism behind, we identified PAP-interacting proteins byyeast two-hybrid assays and a clear result was obtained for the interaction of PAP with snapin, a SNARE-associated protein which binds Snap25 facilitating the vesicular membrane fusion process. We confirmed this interaction by co-localization studies in TMPAP-transfected LNCaP cells (TMPAP/LNCaP cells and in vivo FRET analyses in transient transfected LNCaP cells. The differential gene expression analyses revealed the dysregulation of the same genes known to be related to synaptic vesicular traffic. Both TMPAP and snapin were detected in isolated exosomes. Our results suggest that TMPAP is involved in endo-/exocytosis and disturbed vesicular traffic is a hallmark of prostate adenocarcinoma.

  16. Effects of centrifugation on transmembrane water loss from normal and pathologic erythrocytes

    International Nuclear Information System (INIS)

    Kaperonis, A.A.; Chien, S.

    1989-01-01

    Plasma 125 I-albumin was used as a marker of extracellular dilution in order to study the effect of high-speed centrifugation on transmembrane water distribution in several types of human red cells, including normal (AA), hemoglobin variants (beta A, AS, SC, beta S, and SS), and those from patients with hereditary spherocytosis. SS and AA erythrocytes were also examined for changes in intracellular hemoglobin concentration of three different density fractions and with increasing duration of spin. The minimum force and duration of centrifugation required to impair water permeability were found to vary with the red cell type, the anticoagulant used (heparin or EDTA), the initial hematocrit of the sample centrifuged, as well as among the individual erythrocyte fractions within the same sample. When subjecting pathologic erythrocytes to high-speed centrifugation, the 125 I-albumin dilution technique can be used to determine whether the centrifugation procedure has led to an artifactual red cell water loss and to correct for this when it does occur. An abnormal membrane susceptibility to mechanical stress was demonstrated in erythrocytes from patients with hereditary spherocytosis and several hemoglobinopathies

  17. A mathematical model of T lymphocyte calcium dynamics derived from single transmembrane protein properties

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    Christine Dorothee Schmeitz

    2013-09-01

    Full Text Available Fate decision processes of T lymphocytes are crucial for health and disease. Whether a T lymphocyte is activated, divides, gets anergic or initiates apoptosis depends on extracellular triggers and intracellular signalling. Free cytosolic calcium dynamics plays an important role in this context. The relative contributions of store-derived calcium entry and calcium entry from extracellular space to T lymphocyte activation are still a matter of debate. Here we develop a quantitative mathematical model of T lymphocyte calcium dynamics in order to establish a tool which allows to disentangle cause-effect relationships between ion fluxes and observed calcium time courses. The model is based on single transmembrane protein characteristics which have been determined in independent experiments. This reduces the number of unknown parameters in the model to a minimum and ensures the predictive power of the model. Simulation results are subsequently used for an analysis of whole cell calcium dynamics measured under various experimental conditions. The model accounts for a variety of these conditions, which supports the suitability of the modelling approach. The simulation results suggest a model in which calcium dynamics dominantly relies on the opening of channels in calcium stores while calcium entry through calcium-release activated channels (CRAC is more associated with the maintenance of the T lymphocyte calcium levels and prevents the cell from calcium depletion. Our findings indicate that CRAC guarantees a long-term stable calcium level which is required for cell survival and sustained calcium enhancement.

  18. Entamoeba histolytica phagocytosis of human erythrocytes involves PATMK, a member of the transmembrane kinase family.

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    Douglas R Boettner

    2008-01-01

    Full Text Available Entamoeba histolytica is the cause of amebic colitis and liver abscess. This parasite induces apoptosis in host cells and utilizes exposed ligands such as phosphatidylserine to ingest the apoptotic corpses and invade deeper into host tissue. The purpose of this work was to identify amebic proteins involved in the recognition and ingestion of dead cells. A member of the transmembrane kinase family, phagosome-associated TMK96 (PATMK, was identified in a proteomic screen for early phagosomal proteins. Anti-peptide affinity-purified antibody produced against PATMK demonstrated that it was a type I integral membrane protein that was expressed on the trophozoite surface, and that co-localized with human erythrocytes at the site of contact. The role of PATMK in erythrophagocytosis in vitro was demonstrated by: (i incubation of ameba with anti-PATMK antibodies; (ii PATMK mRNA knock-down using a novel shRNA expression system; and (iii expression of a carboxy-truncation of PATMK (PATMK(delta932. Expression of the carboxy-truncation of PATMK(delta932 also caused a specific reduction in the ability of E. histolytica to establish infection in the intestinal model of amebiasis, however these amebae retained the ability to cause hepatic abscesses when directly injected in the liver. In conclusion, PATMK was identified as a member of the TMK family that participates in erythrophagocytosis and is uniquely required for intestinal infection.

  19. Entamoeba histolytica phagocytosis of human erythrocytes involves PATMK, a member of the transmembrane kinase family.

    Science.gov (United States)

    Boettner, Douglas R; Huston, Christopher D; Linford, Alicia S; Buss, Sarah N; Houpt, Eric; Sherman, Nicholas E; Petri, William A

    2008-01-01

    Entamoeba histolytica is the cause of amebic colitis and liver abscess. This parasite induces apoptosis in host cells and utilizes exposed ligands such as phosphatidylserine to ingest the apoptotic corpses and invade deeper into host tissue. The purpose of this work was to identify amebic proteins involved in the recognition and ingestion of dead cells. A member of the transmembrane kinase family, phagosome-associated TMK96 (PATMK), was identified in a proteomic screen for early phagosomal proteins. Anti-peptide affinity-purified antibody produced against PATMK demonstrated that it was a type I integral membrane protein that was expressed on the trophozoite surface, and that co-localized with human erythrocytes at the site of contact. The role of PATMK in erythrophagocytosis in vitro was demonstrated by: (i) incubation of ameba with anti-PATMK antibodies; (ii) PATMK mRNA knock-down using a novel shRNA expression system; and (iii) expression of a carboxy-truncation of PATMK (PATMK(delta932)). Expression of the carboxy-truncation of PATMK(delta932) also caused a specific reduction in the ability of E. histolytica to establish infection in the intestinal model of amebiasis, however these amebae retained the ability to cause hepatic abscesses when directly injected in the liver. In conclusion, PATMK was identified as a member of the TMK family that participates in erythrophagocytosis and is uniquely required for intestinal infection.

  20. Basigin (CD147), a multifunctional transmembrane glycoprotein with various binding partners.

    Science.gov (United States)

    Muramatsu, Takashi

    2016-05-01

    Basigin, also called CD147 or EMMPRIN, is a transmembrane glycoprotein that belongs to the immunoglobulin superfamily. Basigin has isoforms; the common form (basigin or basigin-2) has two immunoglobulin domains, and the extended form (basigin-1) has three. Basigin is the receptor for cyclophilins, S100A9 and platelet glycoprotein VI, whereas basigin-1 serves as the receptor for the rod-derived cone viability factor. Basigin tightly associates with monocarboxylate transporters and is essential for their cell surface translocation and activities. In the same membrane plane, basigin also associates with other proteins including GLUT1, CD44 and CD98. The carbohydrate portion of basigin is recognized by lectins, such as galectin-3 and E-selectin. These molecular recognitions form the basis for the role of basigin in the transport of nutrients, migration of inflammatory leukocytes and induction of matrix metalloproteinases. Basigin is important in vision, spermatogenesis and other physiological phenomena, and plays significant roles in the pathogenesis of numerous diseases, including cancer. Basigin is also the receptor for an invasive protein RH5, which is present in malaria parasites. © The Authors 2015. Published by Oxford University Press on behalf of the Japanese Biochemical Society.

  1. Mutations in a novel gene with transmembrane domains underlie Usher syndrome type 3.

    Science.gov (United States)

    Joensuu, T; Hämäläinen, R; Yuan, B; Johnson, C; Tegelberg, S; Gasparini, P; Zelante, L; Pirvola, U; Pakarinen, L; Lehesjoki, A E; de la Chapelle, A; Sankila, E M

    2001-10-01

    Usher syndrome type 3 (USH3) is an autosomal recessive disorder characterized by progressive hearing loss, severe retinal degeneration, and variably present vestibular dysfunction, assigned to 3q21-q25. Here, we report on the positional cloning of the USH3 gene. By haplotype and linkage-disequilibrium analyses in Finnish carriers of a putative founder mutation, the critical region was narrowed to 250 kb, of which we sequenced, assembled, and annotated 207 kb. Two novel genes-NOPAR and UCRP-and one previously identified gene-H963-were excluded as USH3, on the basis of mutational analysis. USH3, the candidate gene that we identified, encodes a 120-amino-acid protein. Fifty-two Finnish patients were homozygous for a termination mutation, Y100X; patients in two Finnish families were compound heterozygous for Y100X and for a missense mutation, M44K, whereas patients in an Italian family were homozygous for a 3-bp deletion leading to an amino acid deletion and substitution. USH3 has two predicted transmembrane domains, and it shows no homology to known genes. As revealed by northern blotting and reverse-transcriptase PCR, it is expressed in many tissues, including the retina.

  2. Molecular Simulations of Sequence-Specific Association of Transmembrane Proteins in Lipid Bilayers

    Science.gov (United States)

    Doxastakis, Manolis; Prakash, Anupam; Janosi, Lorant

    2011-03-01

    Association of membrane proteins is central in material and information flow across the cellular membranes. Amino-acid sequence and the membrane environment are two critical factors controlling association, however, quantitative knowledge on such contributions is limited. In this work, we study the dimerization of helices in lipid bilayers using extensive parallel Monte Carlo simulations with recently developed algorithms. The dimerization of Glycophorin A is examined employing a coarse-grain model that retains a level of amino-acid specificity, in three different phospholipid bilayers. Association is driven by a balance of protein-protein and lipid-induced interactions with the latter playing a major role at short separations. Following a different approach, the effect of amino-acid sequence is studied using the four transmembrane domains of the epidermal growth factor receptor family in identical lipid environments. Detailed characterization of dimer formation and estimates of the free energy of association reveal that these helices present significant affinity to self-associate with certain dimers forming non-specific interfaces.

  3. Regulation of KV channel voltage-dependent activation by transmembrane β subunits

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    Xiaohui eSun

    2012-04-01

    Full Text Available Voltage-activated K+ (KV channels are important for shaping action potentials and maintaining resting membrane potential in excitable cells. KV channels contain a central pore-gate domain (PGD surrounded by four voltage-sensing domains (VSD. The VSDs will change conformation in response to alterations of the membrane potential thereby inducing the opening of the PGD. Many KV channels are heteromeric protein complexes containing auxiliary β subunits. These β subunits modulate channel expression and activity to increase functional diversity and render tissue specific phenotypes. This review focuses on the KV β subunits that contain transmembrane (TM segments including the KCNE family and the β subunits of large conductance, Ca2+- and voltage-activated K+ (BK channels. These TM β subunits affect the voltage-dependent activation of KV α subunits. Experimental and computational studies have described the structural location of these β subunits in the channel complexes and the biophysical effects on VSD activation, PGD opening and VSD-PGD coupling. These results reveal some common characteristics and mechanistic insights into KV channel modulation by TM β subunits.

  4. Effects of transmembrane hydraulic pressure on performance of forward osmosis membranes.

    Science.gov (United States)

    Coday, Bryan D; Heil, Dean M; Xu, Pei; Cath, Tzahi Y

    2013-03-05

    Forward osmosis (FO) is an emerging membrane separation process that continues to be tested and implemented in various industrial water and wastewater treatment applications. The growing interests in the technology have prompted laboratories and manufacturers to adopt standard testing methods to ensure accurate comparison of membrane performance under laboratory-controlled conditions; however, standardized methods might not capture specific operating conditions unique to industrial applications. Experiments with cellulose triacetate (CTA) and polyamide thin-film composite (TFC) FO membranes demonstrated that hydraulic transmembrane pressure (TMP), common in industrial operation of FO membrane elements, could affect membrane performance. Experiments were conducted with three FO membranes and with increasing TMP up to a maximum of 50 psi (3.45 bar). The feed solution was a mixture of salts and the draw solution was either a NaCl solution or concentrated seawater at similar osmotic pressure. Results revealed that TMP minimally affected water flux, reverse salt flux (RSF), and solute rejection of the CTA membrane. However, water flux through TFC membranes might slightly increase with increasing TMP, and RSF substantially declines with increasing TMP. It was observed that rejection of feed constituents was influenced by TMP and RSF.

  5. Brucella Intracellular Life Relies on the Transmembrane Protein CD98 Heavy Chain.

    Science.gov (United States)

    Keriel, Anne; Botella, Eric; Estrach, Soline; Bragagnolo, Gabriel; Vergunst, Annette C; Feral, Chloe C; O'Callaghan, David

    2015-06-01

    Brucella are intracellular bacterial pathogens that use a type IV secretion system (T4SS) to escape host defenses and create a niche in which they can multiply. Although the importance of Brucella T4SS is clear, little is known about its interactions with host cell structures. In this study, we identified the eukaryotic protein CD98hc as a partner for Brucella T4SS subunit VirB2. This transmembrane glycoprotein is involved in amino acid transport, modulation of integrin signaling, and cell-to-cell fusion. Knockdown of CD98hc expression in HeLa cells demonstrated that it is essential for Brucella infection. Using knockout dermal fibroblasts, we confirmed its role for Brucella but found that it is not required for Salmonella infection. CD98hc transiently accumulates around the bacteria during the early phases of infection and is required for both optimal bacterial uptake and intracellular multiplication of Brucella. These results provide new insights into the complex interplay between Brucella and its host. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  6. New Insights into Molecular Organization of Human Neuraminidase-1: Transmembrane Topology and Dimerization Ability

    Science.gov (United States)

    Maurice, Pascal; Baud, Stéphanie; Bocharova, Olga V.; Bocharov, Eduard V.; Kuznetsov, Andrey S.; Kawecki, Charlotte; Bocquet, Olivier; Romier, Beatrice; Gorisse, Laetitia; Ghirardi, Maxime; Duca, Laurent; Blaise, Sébastien; Martiny, Laurent; Dauchez, Manuel; Efremov, Roman G.; Debelle, Laurent

    2016-12-01

    Neuraminidase 1 (NEU1) is a lysosomal sialidase catalyzing the removal of terminal sialic acids from sialyloconjugates. A plasma membrane-bound NEU1 modulating a plethora of receptors by desialylation, has been consistently documented from the last ten years. Despite a growing interest of the scientific community to NEU1, its membrane organization is not understood and current structural and biochemical data cannot account for such membrane localization. By combining molecular biology and biochemical analyses with structural biophysics and computational approaches, we identified here two regions in human NEU1 - segments 139-159 (TM1) and 316-333 (TM2) - as potential transmembrane (TM) domains. In membrane mimicking environments, the corresponding peptides form stable α-helices and TM2 is suited for self-association. This was confirmed with full-size NEU1 by co-immunoprecipitations from membrane preparations and split-ubiquitin yeast two hybrids. The TM2 region was shown to be critical for dimerization since introduction of point mutations within TM2 leads to disruption of NEU1 dimerization and decrease of sialidase activity in membrane. In conclusion, these results bring new insights in the molecular organization of membrane-bound NEU1 and demonstrate, for the first time, the presence of two potential TM domains that may anchor NEU1 in the membrane, control its dimerization and sialidase activity.

  7. Residual mitochondrial transmembrane potential decreases unsaturated fatty acid level in sake yeast during alcoholic fermentation

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    Kazutaka Sawada

    2016-01-01

    Full Text Available Oxygen, a key nutrient in alcoholic fermentation, is rapidly depleted during this process. Several pathways of oxygen utilization have been reported in the yeast Saccharomyces cerevisiae during alcoholic fermentation, namely synthesis of unsaturated fatty acid, sterols and heme, and the mitochondrial electron transport chain. However, the interaction between these pathways has not been investigated. In this study, we showed that the major proportion of unsaturated fatty acids of ester-linked lipids in sake fermentation mash is derived from the sake yeast rather than from rice or koji (rice fermented with Aspergillus. Additionally, during alcoholic fermentation, inhibition of the residual mitochondrial activity of sake yeast increases the levels of unsaturated fatty acids of ester-linked lipids. These findings indicate that the residual activity of the mitochondrial electron transport chain reduces molecular oxygen levels and decreases the synthesis of unsaturated fatty acids, thereby increasing the synthesis of estery flavors by sake yeast. This is the first report of a novel link between residual mitochondrial transmembrane potential and the synthesis of unsaturated fatty acids by the brewery yeast during alcoholic fermentation.

  8. MemBrain: An Easy-to-Use Online Webserver for Transmembrane Protein Structure Prediction

    Science.gov (United States)

    Yin, Xi; Yang, Jing; Xiao, Feng; Yang, Yang; Shen, Hong-Bin

    2018-03-01

    Membrane proteins are an important kind of proteins embedded in the membranes of cells and play crucial roles in living organisms, such as ion channels, transporters, receptors. Because it is difficult to determinate the membrane protein's structure by wet-lab experiments, accurate and fast amino acid sequence-based computational methods are highly desired. In this paper, we report an online prediction tool called MemBrain, whose input is the amino acid sequence. MemBrain consists of specialized modules for predicting transmembrane helices, residue-residue contacts and relative accessible surface area of α-helical membrane proteins. MemBrain achieves a prediction accuracy of 97.9% of A TMH, 87.1% of A P, 3.2 ± 3.0 of N-score, 3.1 ± 2.8 of C-score. MemBrain-Contact obtains 62%/64.1% prediction accuracy on training and independent dataset on top L/5 contact prediction, respectively. And MemBrain-Rasa achieves Pearson correlation coefficient of 0.733 and its mean absolute error of 13.593. These prediction results provide valuable hints for revealing the structure and function of membrane proteins. MemBrain web server is free for academic use and available at www.csbio.sjtu.edu.cn/bioinf/MemBrain/. [Figure not available: see fulltext.

  9. Mice deficient in transmembrane prostatic acid phosphatase display increased GABAergic transmission and neurological alterations.

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    Heidi O Nousiainen

    Full Text Available Prostatic acid phosphatase (PAP, the first diagnostic marker and present therapeutic target for prostate cancer, modulates nociception at the dorsal root ganglia (DRG, but its function in the central nervous system has remained unknown. We studied expression and function of TMPAP (the transmembrane isoform of PAP in the brain by utilizing mice deficient in TMPAP (PAP-/- mice. Here we report that TMPAP is expressed in a subpopulation of cerebral GABAergic neurons, and mice deficient in TMPAP show multiple behavioral and neurochemical features linked to hyperdopaminergic dysregulation and altered GABAergic transmission. In addition to increased anxiety, disturbed prepulse inhibition, increased synthesis of striatal dopamine, and augmented response to amphetamine, PAP-deficient mice have enlarged lateral ventricles, reduced diazepam-induced loss of righting reflex, and increased GABAergic tone in the hippocampus. TMPAP in the mouse brain is localized presynaptically, and colocalized with SNARE-associated protein snapin, a protein involved in synaptic vesicle docking and fusion, and PAP-deficient mice display altered subcellular distribution of snapin. We have previously shown TMPAP to reside in prostatic exosomes and we propose that TMPAP is involved in the control of GABAergic tone in the brain also through exocytosis, and that PAP deficiency produces a distinct neurological phenotype.

  10. Tetraspanins and Transmembrane Adaptor Proteins As Plasma Membrane Organizers-Mast Cell Case.

    Science.gov (United States)

    Halova, Ivana; Draber, Petr

    2016-01-01

    The plasma membrane contains diverse and specialized membrane domains, which include tetraspanin-enriched domains (TEMs) and transmembrane adaptor protein (TRAP)-enriched domains. Recent biophysical, microscopic, and functional studies indicated that TEMs and TRAP-enriched domains are involved in compartmentalization of physicochemical events of such important processes as immunoreceptor signal transduction and chemotaxis. Moreover, there is evidence of a cross-talk between TEMs and TRAP-enriched domains. In this review we discuss the presence and function of such domains and their crosstalk using mast cells as a model. The combined data based on analysis of selected mast cell-expressed tetraspanins [cluster of differentiation (CD)9, CD53, CD63, CD81, CD151)] or TRAPs [linker for activation of T cells (LAT), non-T cell activation linker (NTAL), and phosphoprotein associated with glycosphingolipid-enriched membrane microdomains (PAG)] using knockout mice or specific antibodies point to a diversity within these two families and bring evidence of the important roles of these molecules in signaling events. An example of this diversity is physical separation of two TRAPs, LAT and NTAL, which are in many aspects similar but show plasma membrane location in different microdomains in both non-activated and activated cells. Although our understanding of TEMs and TRAP-enriched domains is far from complete, pharmaceutical applications of the knowledge about these domains are under way.

  11. Lysosomal-associated transmembrane protein 5 (LAPTM5 is a molecular partner of CD1e.

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    Catherine Angénieux

    Full Text Available The CD1e protein participates in the presentation of lipid antigens in dendritic cells. Its transmembrane precursor is transported to lysosomes where it is cleaved into an active soluble form. In the presence of bafilomycin, which inhibits vacuolar ATPase and consequently the acidification of endosomal compartments, CD1e associates with a 27 kD protein. In this work, we identified this molecular partner as LAPTM5. The latter protein and CD1e colocalize in trans-Golgi and late endosomal compartments. The quantity of LAPTM5/CD1e complexes increases when the cells are treated with bafilomycin, probably due to the protection of LAPTM5 from lysosomal proteases. Moreover, we could demonstrate that LAPTM5/CD1e association occurs under physiological conditions. Although LAPTM5 was previously shown to act as a platform recruiting ubiquitin ligases and facilitating the transport of receptors to lysosomes, we found no evidence that LATPM5 controls either CD1e ubiquitination or the generation of soluble lysosomal CD1e proteins. Notwithstanding these last observations, the interaction of LAPTM5 with CD1e and their colocalization in antigen processing compartments both suggest that LAPTM5 might influence the role of CD1e in the presentation of lipid antigens.

  12. The MARVEL transmembrane motif of occludin mediates oligomerization and targeting to the basolateral surface in epithelia.

    Science.gov (United States)

    Yaffe, Yakey; Shepshelovitch, Jeanne; Nevo-Yassaf, Inbar; Yeheskel, Adva; Shmerling, Hedva; Kwiatek, Joanna M; Gaus, Katharina; Pasmanik-Chor, Metsada; Hirschberg, Koret

    2012-08-01

    Occludin (Ocln), a MARVEL-motif-containing protein, is found in all tight junctions. MARVEL motifs are comprised of four transmembrane helices associated with the localization to or formation of diverse membrane subdomains by interacting with the proximal lipid environment. The functions of the Ocln MARVEL motif are unknown. Bioinformatics sequence- and structure-based analyses demonstrated that the MARVEL domain of Ocln family proteins has distinct evolutionarily conserved sequence features that are consistent with its basolateral membrane localization. Live-cell microscopy, fluorescence resonance energy transfer (FRET) and bimolecular fluorescence complementation (BiFC) were used to analyze the intracellular distribution and self-association of fluorescent-protein-tagged full-length human Ocln or the Ocln MARVEL motif excluding the cytosolic C- and N-termini (amino acids 60-269, FP-MARVEL-Ocln). FP-MARVEL-Ocln efficiently arrived at the plasma membrane (PM) and was sorted to the basolateral PM in filter-grown polarized MDCK cells. A series of conserved aromatic amino acids within the MARVEL domain were found to be associated with Ocln dimerization using BiFC. FP-MARVEL-Ocln inhibited membrane pore growth during Triton-X-100-induced solubilization and was shown to increase the membrane-ordered state using Laurdan, a lipid dye. These data demonstrate that the Ocln MARVEL domain mediates self-association and correct sorting to the basolateral membrane.

  13. Transmission of integrin β7 transmembrane domain topology enables gut lymphoid tissue development.

    Science.gov (United States)

    Sun, Hao; Lagarrigue, Frederic; Gingras, Alexandre R; Fan, Zhichao; Ley, Klaus; Ginsberg, Mark H

    2018-04-02

    Integrin activation regulates adhesion, extracellular matrix assembly, and cell migration, thereby playing an indispensable role in development and in many pathological processes. A proline mutation in the central integrin β3 transmembrane domain (TMD) creates a flexible kink that uncouples the topology of the inner half of the TMD from the outer half. In this study, using leukocyte integrin α4β7, which enables development of gut-associated lymphoid tissue (GALT), we examined the biological effect of such a proline mutation and report that it impairs agonist-induced talin-mediated activation of integrin α4β7, thereby inhibiting rolling lymphocyte arrest, a key step in transmigration. Furthermore, the α4β7(L721P) mutation blocks lymphocyte homing to and development of the GALT. These studies show that impairing the ability of an integrin β TMD to transmit talin-induced TMD topology inhibits agonist-induced physiological integrin activation and biological function in development. © 2018 Sun et al.

  14. The cystic fibrosis transmembrane recruiter the alter ego of CFTR as a multi-kinase anchor.

    Science.gov (United States)

    Mehta, Anil

    2007-11-01

    This review focuses on a newly discovered interaction between protein kinases involved in cellular energetics, a process that may be disturbed in cystic fibrosis for unknown reasons. I propose a new model where kinase-mediated cellular transmission of energy provides mechanistic insight to a latent role of the cystic fibrosis transmembrane conductance regulator (CFTR). I suggest that CFTR acts as a multi-kinase recruiter to the apical epithelial membrane. My group finds that, in the cytosol, two protein kinases involved in cell energy homeostasis, nucleoside diphosphate kinase (NDPK) and AMP-activated kinase (AMPK), bind one another. Preliminary data suggest that both can also bind CFTR (function unclear). The disrupted role of this CFTR-kinase complex as 'membrane transmitter to the cell' is proposed as an alternative paradigm to the conventional ion transport mediated and CFTR/chloride-centric view of cystic fibrosis pathogenesis. Chloride remains important, but instead, chloride-induced control of the phosphohistidine content of one kinase component (NDPK, via a multi-kinase complex that also includes a third kinase, CK2; formerly casein kinase 2). I suggest that this complex provides the necessary near-equilibrium conditions needed for efficient transmission of phosphate energy to proteins controlling cellular energetics. Crucially, a new role for CFTR as a kinase controller is proposed with ionic concentration acting as a signal. The model posits a regulatory control relay for energy sensing involving a cascade of protein kinases bound to CFTR.

  15. Dissection of seroreactivity against the tryptophan-rich motif of the feline immunodeficiency virus transmembrane glycoprotein

    International Nuclear Information System (INIS)

    Freer, Giulia; Giannecchini, Simone; Tissot, Alain; Bachmann, Martin F.; Rovero, Paolo; Serres, Pierre Francoise; Bendinelli, Mauro

    2004-01-01

    Immunogenicity of the tryptophan-rich motif (TrpM) in the membrane-proximal ectodomain of the transmembrane (TM) glycoprotein of feline immunodeficiency virus (FIV) was investigated. Peptide 59, a peptide containing the TrpM of the TM of FIV, was covalently coupled to Qβ phage virus-like particles (Qβ-59) in the attempt to induce potent anti-TrpM B cell responses in cats. All Qβ-59 immunized cats, but not cats that received a mixture of uncoupled Qβ and peptide 59, developed antibodies that reacted with a same epitope in extensive binding and binding competition assays. The epitope recognized was composed of three amino acids, two of which are adjacent. However, Qβ-59-immune sera failed to recognize whole FIV in all binding and neutralization assays performed. Furthermore, no reactivity against the TrpM was detected by screening sera from FIV-infected cats that had reacted with TM peptides, confirming that this epitope does not seem to be serologically functional in the FIV virion. The data suggest that TrpM may not be a suitable target for antiviral vaccine design

  16. Non-equilibrium dynamics of 2D liquid crystals driven by transmembrane gas flow.

    Science.gov (United States)

    Seki, Kazuyoshi; Ueda, Ken; Okumura, Yu-ichi; Tabe, Yuka

    2011-07-20

    Free-standing films composed of several layers of chiral smectic liquid crystals (SmC*) exhibited unidirectional director precession under various vapor transfers across the films. When the transferred vapors were general organic solvents, the precession speed linearly depended on the momentum of the transmembrane vapors, where the proportional constant was independent of the kind of vapor. In contrast, the same SmC* films under water transfer exhibited precession in the opposite direction. As a possible reason for the rotational inversion, we suggest the competition of two origins for the torques, one of which is microscopic and the other macroscopic. Next, we tried to move an external object by making use of the liquid crystal (LC) motion. When a solid or a liquid particle was set on a film under vapor transfer, the particle was rotated in the same direction as the LC molecules. Using home-made laser tweezers, we measured the force transmitted from the film to the particle, which we found to be several pN.

  17. Transmembrane carbonic anhydrase isozymes IX and XII in the female mouse reproductive organs

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    Tomas Eija

    2004-10-01

    Full Text Available Abstract Background Carbonic anhydrase (CA classically catalyses the reversible hydration of dissolved CO2 to form bicarbonate ions and protons. The twelve active CA isozymes are thought to regulate a variety of cellular functions including several processes in the reproductive systems. Methods The present study was designed to investigate the expression of transmembrane CAs, CA IX and XII, in the mouse uterus, ovary and placenta. The expression of CA IX and XII was examined by immunoperoxidase staining method and western blotting. CA II and XIII served as positive controls since they are known to be present in the mouse reproductive tract. Results The data of our study indicated that CA XII is expressed in the mouse endometrium. Only very faint signal was observed in the corpus luteum of the ovary and the placenta remained mainly negative. CA IX showed weak reaction in the endometrial epithelium, while it was completely absent in the ovary and placenta. Conclusion The conservation of CA XII expression in both mouse and human endometrium suggests a role for this isozyme in reproductive physiology.

  18. Tetraspanins and Transmembrane Adaptor Proteins as Plasma Membrane Organizers – Mast Cell Case

    Directory of Open Access Journals (Sweden)

    Ivana eHalova

    2016-05-01

    Full Text Available The plasma membrane contains diverse and specialized membrane domains, which include tetraspanin-enriched domains (TEMs and transmembrane adaptor protein (TRAP-enriched domains. Recent biophysical, microscopic and functional studies indicated that TEMs and TRAP-enriched domains are involved in compartmentalization of physicochemical events of such important processes as immunoreceptor signal transduction and chemotaxis. Moreover, there is evidence of a cross-talk between TEMs and TRAP-enriched domains. In this review we discuss the presence and function of such domains and their crosstalk using mast cells as a model. The combined data based on analysis of selected mast cell-expressed tetraspanins [cluster of differentiation (CD9, CD53, CD63, CD81, CD151] or TRAPs [linker for activation of T cells (LAT, non-T cell activation linker (NTAL, and phosphoprotein associated with glycosphingolipid-enriched membrane microdomains (PAG] using knockout mice or specific antibodies point to a diversity within these two families and bring evidence of the important roles of these molecules in signaling events. An example of this diversity is physical separation of two TRAPs, LAT and NTAL, which are in many aspects similar but show plasma membrane location in different microdomains in both non-activated and activated cells. Although our understanding of TEMs and TRAP-enriched domains is far from complete, pharmaceutical applications of the knowledge about these domains are under way.

  19. Simulations of skin barrier function: free energies of hydrophobic and hydrophilic transmembrane pores in ceramide bilayers.

    Science.gov (United States)

    Notman, Rebecca; Anwar, Jamshed; Briels, W J; Noro, Massimo G; den Otter, Wouter K

    2008-11-15

    Transmembrane pore formation is central to many biological processes such as ion transport, cell fusion, and viral infection. Furthermore, pore formation in the ceramide bilayers of the stratum corneum may be an important mechanism by which penetration enhancers such as dimethylsulfoxide (DMSO) weaken the barrier function of the skin. We have used the potential of mean constraint force (PMCF) method to calculate the free energy of pore formation in ceramide bilayers in both the innate gel phase and in the DMSO-induced fluidized state. Our simulations show that the fluid phase bilayers form archetypal water-filled hydrophilic pores similar to those observed in phospholipid bilayers. In contrast, the rigid gel-phase bilayers develop hydrophobic pores. At the relatively small pore diameters studied here, the hydrophobic pores are empty rather than filled with bulk water, suggesting that they do not compromise the barrier function of ceramide membranes. A phenomenological analysis suggests that these vapor pores are stable, below a critical radius, because the penalty of creating water-vapor and tail-vapor interfaces is lower than that of directly exposing the strongly hydrophobic tails to water. The PMCF free energy profile of the vapor pore supports this analysis. The simulations indicate that high DMSO concentrations drastically impair the barrier function of the skin by strongly reducing the free energy required for pore opening.

  20. Poxvirus-encoded TNF decoy receptors inhibit the biological activity of transmembrane TNF.

    Science.gov (United States)

    Pontejo, Sergio M; Alejo, Ali; Alcami, Antonio

    2015-10-01

    Poxviruses encode up to four different soluble TNF receptors, named cytokine response modifier B (CrmB), CrmC, CrmD and CrmE. These proteins mimic the extracellular domain of the cellular TNF receptors to bind and inhibit the activity of TNF and, in some cases, other TNF superfamily ligands. Most of these ligands are released after the enzymic cleavage of a membrane precursor. However, transmembrane TNF (tmTNF) is not only a precursor of soluble TNF but also exerts specific pro-inflammatory and immunological activities. Here, we report that viral TNF receptors bound and inhibited tmTNF and describe some interesting differences in their activity against the soluble cytokine. Thus, CrmE, which does not inhibit mouse soluble TNF, could block murine tmTNF-induced cytotoxicity. We propose that this anti-tmTNF effect should be taken into consideration when assessing the role of viral TNF decoy receptors in the pathogenesis of poxvirus.

  1. A combination of compositional index and genetic algorithm for predicting transmembrane helical segments.

    Directory of Open Access Journals (Sweden)

    Nazar Zaki

    Full Text Available Transmembrane helix (TMH topology prediction is becoming a focal problem in bioinformatics because the structure of TM proteins is difficult to determine using experimental methods. Therefore, methods that can computationally predict the topology of helical membrane proteins are highly desirable. In this paper we introduce TMHindex, a method for detecting TMH segments using only the amino acid sequence information. Each amino acid in a protein sequence is represented by a Compositional Index, which is deduced from a combination of the difference in amino acid occurrences in TMH and non-TMH segments in training protein sequences and the amino acid composition information. Furthermore, a genetic algorithm was employed to find the optimal threshold value for the separation of TMH segments from non-TMH segments. The method successfully predicted 376 out of the 378 TMH segments in a dataset consisting of 70 test protein sequences. The sensitivity and specificity for classifying each amino acid in every protein sequence in the dataset was 0.901 and 0.865, respectively. To assess the generality of TMHindex, we also tested the approach on another standard 73-protein 3D helix dataset. TMHindex correctly predicted 91.8% of proteins based on TM segments. The level of the accuracy achieved using TMHindex in comparison to other recent approaches for predicting the topology of TM proteins is a strong argument in favor of our proposed method.The datasets, software together with supplementary materials are available at: http://faculty.uaeu.ac.ae/nzaki/TMHindex.htm.

  2. Contribution of Kunitz protease inhibitor and transmembrane domains to amyloid precursor protein homodimerization.

    Science.gov (United States)

    Ben Khalifa, N; Tyteca, D; Courtoy, P J; Renauld, J C; Constantinescu, S N; Octave, J N; Kienlen-Campard, P

    2012-01-01

    The two major isoforms of the human amyloid precursor protein (APP) are APP695 and APP751. They differ by the insertion of a Kunitz-type protease inhibitor (KPI) sequence in the extracellular domain of APP751. APP-KPI isoforms are increased in Alzheimer's disease brains, and they could be associated with disease progression. Recent studies have shown that APP processing to Aβ is regulated by homodimerization, which involves both extracellular and juxtamembrane/transmembrane (JM/TM) regions. Our aim is to understand the mechanisms controlling APP dimerization and the contribution of the ectodomain and JM/TM regions to this process. We used bimolecular fluorescence complementation approaches coupled to fluorescence-activated cell sorting analysis to measure the dimerization level of different APP isoforms and APP C-terminal fragments (C99) mutated in their JM/TM region. APP751 was found to form significantly more homodimers than APP695. Mutation of dimerization motifs in the TM domain of APP or C99 did not significantly affect fluorescence complementation. These findings indicate that the KPI domain plays a major role in APP dimerization. They set the basis for further investigation of the relation between dimerization, metabolism and function of APP. Copyright © 2012 S. Karger AG, Basel.

  3. Targeting a genetic defect: cystic fibrosis transmembrane conductance regulator modulators in cystic fibrosis

    Directory of Open Access Journals (Sweden)

    Nico Derichs

    2013-03-01

    Full Text Available Cystic fibrosis (CF is caused by genetic mutations that affect the cystic fibrosis transmembrane conductance regulator (CFTR protein. These mutations can impact the synthesis and transfer of the CFTR protein to the apical membrane of epithelial cells, as well as influencing the gating or conductance of chloride and bicarbonate ions through the channel. CFTR dysfunction results in ionic imbalance of epithelial secretions in several organ systems, such as the pancreas, gastrointestinal tract, liver and the respiratory system. Since discovery of the CFTR gene in 1989, research has focussed on targeting the underlying genetic defect to identify a disease-modifying treatment for CF. Investigated management strategies have included gene therapy and the development of small molecules that target CFTR mutations, known as CFTR modulators. CFTR modulators are typically identified by high-throughput screening assays, followed by preclinical validation using cell culture systems. Recently, one such modulator, the CFTR potentiator ivacaftor, was approved as an oral therapy for CF patients with the G551D-CFTR mutation. The clinical development of ivacaftor not only represents a breakthrough in CF care but also serves as a noteworthy example of personalised medicine.

  4. Interaction of protein C inhibitor with the type II transmembrane serine protease enteropeptidase.

    Directory of Open Access Journals (Sweden)

    Thomas A Prohaska

    Full Text Available The serine protease inhibitor protein C inhibitor (PCI is expressed in many human tissues and exhibits broad protease reactivity. PCI binds glycosaminoglycans and certain phospholipids, which modulate its inhibitory activity. Enteropeptidase (EP is a type II transmembrane serine protease mainly found on the brush border membrane of epithelial cells in the duodenum, where it activates trypsinogen to initiate the digestion of food proteins. Some active EP is also present in duodenal fluid and has been made responsible for causing pancreatitis in case of duodeno-pancreatic reflux. Together with its substrate trypsinogen, EP is furthermore present in the epidermis and in some cancer cells. In this report, we show that PCI inhibited EP with an apparent 2nd order rate constant of 4.48 × 10(4 M(-1 s(-1. Low molecular weight (LMWH and unfractionated heparin (UFH slightly reduced the inhibitory effect of PCI. The SI (stoichiometry of inhibition value for the inhibition of EP by PCI was 10.8 in the absence and 17.9 in the presence of UFH (10 U/ml. By inhibiting trypsin, chymotrypsin, and additionally EP, PCI might play a role in the protection of the pancreas from autodigestion. Furthermore the interaction of PCI with EP may influence the regulation of epithelial differentiation.

  5. System and methods for predicting transmembrane domains in membrane proteins and mining the genome for recognizing G-protein coupled receptors

    Science.gov (United States)

    Trabanino, Rene J; Vaidehi, Nagarajan; Hall, Spencer E; Goddard, William A; Floriano, Wely

    2013-02-05

    The invention provides computer-implemented methods and apparatus implementing a hierarchical protocol using multiscale molecular dynamics and molecular modeling methods to predict the presence of transmembrane regions in proteins, such as G-Protein Coupled Receptors (GPCR), and protein structural models generated according to the protocol. The protocol features a coarse grain sampling method, such as hydrophobicity analysis, to provide a fast and accurate procedure for predicting transmembrane regions. Methods and apparatus of the invention are useful to screen protein or polynucleotide databases for encoded proteins with transmembrane regions, such as GPCRs.

  6. Championship Communication

    Science.gov (United States)

    VanDerveer, Beth; Butterick, Betsy

    2017-01-01

    This article discusses team development and focuses specifically on championship communication and team-building activities. Team development takes time and the process typically occurs in stages. Regardless of the sport or what the competitive field may look like, communication is an often overlooked, yet vital element in cultivating a…

  7. Effective communication

    International Nuclear Information System (INIS)

    Kuntz, B.S.

    1989-01-01

    At the Waste Isolation Pilot Plant (WIPP) the responsibilities assigned to public affairs (PA) include communications to two main groups: institutional representatives and the general public. Research data indicates that these two populations perceive risk in different fashions. This paper discusses these distinct perceptions and how the communication programs at WIPP have been designed to accommodate these two differences

  8. Satellite Communications

    Indian Academy of Sciences (India)

    First page Back Continue Last page Overview Graphics. Satellite Communications. Arthur C Clarke wrote a seminal paper in 1945 in wireless world. Use three satellites in geo-synchronous orbit to enable intercontinental communications. System could be realised in '50 to 100 years'

  9. Short length transmembrane domains having voluminous exoplasmic halves determine retention of Type II membrane proteins in the Golgi complex

    OpenAIRE

    Quiroga, Rodrigo; Trenchi, Alejandra; Gonzalez Montoro, Ayelén; Valdez, Javier Esteban; Maccioni, Hugo Jose Fernando

    2017-01-01

    It is still unclear why some proteins that travel along the secretory pathway are retained in the Golgi complex whereas others make their way to the plasma membrane. Recent bioinformatic analyses on a large number of single-spanning membrane proteins support the hypothesis that specific features of the transmembrane domain (TMD) are relevant to the sorting of these proteins to particular organelles. Here we experimentally test this hypothesis for Golgi and plasma membrane proteins. Using the ...

  10. Vitamin A transport and the transmembrane pore in the cell-surface receptor for plasma retinol binding protein.

    Directory of Open Access Journals (Sweden)

    Ming Zhong

    Full Text Available Vitamin A and its derivatives (retinoids play diverse and crucial functions from embryogenesis to adulthood and are used as therapeutic agents in human medicine for eye and skin diseases, infections and cancer. Plasma retinol binding protein (RBP is the principal and specific vitamin A carrier in the blood and binds vitamin A at 1:1 ratio. STRA6 is the high-affinity membrane receptor for RBP and mediates cellular vitamin A uptake. STRA6 null mice have severely depleted vitamin A reserves for vision and consequently have vision loss, even under vitamin A sufficient conditions. STRA6 null humans have a wide range of severe pathological phenotypes in many organs including the eye, brain, heart and lung. Known membrane transport mechanisms involve transmembrane pores that regulate the transport of the substrate (e.g., the gating of ion channels. STRA6 represents a new type of membrane receptor. How this receptor interacts with its transport substrate vitamin A and the functions of its nine transmembrane domains are still completely unknown. These questions are critical to understanding the molecular basis of STRA6's activities and its regulation. We employ acute chemical modification to introduce chemical side chains to STRA6 in a site-specific manner. We found that modifications with specific chemicals at specific positions in or near the transmembrane domains of this receptor can almost completely suppress its vitamin A transport activity. These experiments provide the first evidence for the existence of a transmembrane pore, analogous to the pore of ion channels, for this new type of cell-surface receptor.

  11. Corruption of host seven-transmembrane proteins by pathogenic microbes: a common theme in animals and plants?

    Science.gov (United States)

    Panstruga, Ralph; Schulze-Lefert, Paul

    2003-04-01

    Human diseases like AIDS, malaria, and pneumonia are caused by pathogens that corrupt host chemokine G-protein coupled receptors for molecular docking. Comparatively, little is known about plant host factors that are required for pathogenesis and that may serve as receptors for the entry of pathogenic microbes. Here, we review potential analogies between human chemokine receptors and the plant seven-transmembrane MLO protein, a candidate serving a dual role as docking molecule and defence modulator for the phytopathogenic powdery mildew fungus.

  12. Biologically Complex Planar Cell Plasma Membranes Supported on Polyelectrolyte Cushions Enhance Transmembrane Protein Mobility and Retain Native Orientation.

    Science.gov (United States)

    Liu, Han-Yuan; Chen, Wei-Liang; Ober, Christopher K; Daniel, Susan

    2018-01-23

    Reconstituted supported lipid bilayers (SLB) are widely used as in vitro cell-surface models because they are compatible with a variety of surface-based analytical techniques. However, one of the challenges of using SLBs as a model of the cell surface is the limited complexity in membrane composition, including the incorporation of transmembrane proteins and lipid diversity that may impact the activity of those proteins. Additionally, it is challenging to preserve the transmembrane protein native orientation, function, and mobility in SLBs. Here, we leverage the interaction between cell plasma membrane vesicles and polyelectrolyte brushes to create planar bilayers from cell plasma membrane vesicles that have budded from the cell surface. This approach promotes the direct incorporation of membrane proteins and other species into the planar bilayer without using detergent or reconstitution and preserves membrane constituents. Furthermore, the structure of the polyelectrolyte brush serves as a cushion between the planar bilayer and rigid supporting surface, limiting the interaction of the cytosolic domains of membrane proteins with this surface. Single particle tracking was used to analyze the motion of GPI-linked yellow fluorescent proteins (GPI-YFP) and neon-green fused transmembrane P2X2 receptors (P2X2-neon) and shows that this platform retains over 75% mobility of multipass transmembrane proteins in its native membrane environment. An enzyme accessibility assay confirmed that the protein orientation is preserved and results in the extracellular domain facing toward the bulk phase and the cytosolic side facing the support. Because the platform presented here retains the complexity of the cell plasma membrane and preserves protein orientation and mobility, it is a better representative mimic of native cell surfaces, which may find many applications in biological assays aimed at understanding cell membrane phenomena.

  13. [Application of Brownian dynamics to the description of transmembrane ion flow as exemplified by the chloride channel of glycine receptor].

    Science.gov (United States)

    Boronovskiĭ, S E; Nartsissov, Ia R

    2009-01-01

    Using the Brownian dynamics of the movement of hydrated ion in a viscous water solution, a mathematical model has been built, which describes the transport of charged particles through a single protein pore in a lipid membrane. The dependences of transmembrane ion currents on ion concentrations in solution have been obtained. It was shown that, if the geometry of a membrane pore is identical to that of the inner part of the glycine receptor channel and there is no ion selectivity, then the values of both chloride and sodium currents are not greater than 0.5 pA at the physiological concentrations of these ions. If local charge heterogeneity caused by charged amino acid residues of transmembrane protein segments is included into the model calculations, the chloride current increases to about 3.7 pA, which exceeds more than seven times the value for sodium ions under the conditions of the complex channel geometry in the range of physiological concentrations of ions in the solution. The model takes changes in the density of charge distribution both inside the channel and near the protein surface into account. The alteration of pore geometry can be also considered as a parameter at the researcher's option. Thus, the model appears as an effective tool for the description of transmembrane currents for other types of membrane channels.

  14. Transmembrane START domain proteins: in silico identification, characterization and expression analysis under stress conditions in chickpea (Cicer arietinum L.).

    Science.gov (United States)

    Satheesh, Viswanathan; Chidambaranathan, Parameswaran; Jagannadham, Prasanth Tejkumar; Kumar, Vajinder; Jain, Pradeep K; Chinnusamy, Viswanathan; Bhat, Shripad R; Srinivasan, R

    2016-01-01

    Steroidogenic acute regulatory related transfer (StART) proteins that are involved in transport of lipid molecules, play a myriad of functions in insects, mammals and plants. These proteins consist of a modular START domain of approximately 200 amino acids which binds and transfers the lipids. In the present study we have performed a genome-wide search for all START domain proteins in chickpea. The search identified 36 chickpea genes belonging to the START domain family. Through a phylogenetic tree reconstructed with Arabidopsis, rice, chickpea, and soybean START proteins, we were able to identify four transmembrane START (TM-START) proteins in chickpea. These four proteins are homologous to the highly conserved mammalian phosphatidylcholine transfer proteins. Multiple sequence alignment of all the transmembrane containing START proteins from Arabidopsis, rice, chickpea, and soybean revealed that the amino acid residues to which phosphatidylcholine binds in mammals, is also conserved in all these plant species, implying an important functional role and a very similar mode of action of all these proteins across dicots and monocots. This study characterizes a few of the not so well studied transmembrane START superfamily genes that may be involved in stress signaling. Expression analysis in various tissues showed that these genes are predominantly expressed in flowers and roots of chickpea. Three of the chickpea TM-START genes showed induced expression in response to drought, salt, wound and heat stress, suggesting their role in stress response.

  15. The nectin-1α transmembrane domain, but not the cytoplasmic tail, influences cell fusion induced by HSV-1 glycoproteins

    International Nuclear Information System (INIS)

    Subramanian, Ravi P.; Dunn, Jennifer E.; Geraghty, Robert J.

    2005-01-01

    Nectin-1 is a receptor for herpes simplex virus (HSV), a member of the immunoglobulin superfamily, and a cellular adhesion molecule. To study domains of nectin-1α involved in cell fusion, we measured the ability of nectin-1α/nectin-2α chimeras, nectin-1α/CD4 chimeras, and transmembrane domain and cytoplasmic tail mutants of nectin-1α to promote cell fusion induced by HSV-1 glycoproteins. Our results demonstrate that only chimeras and mutants containing the entire V-like domain and a link to the plasma membrane conferred cell-fusion activity. The transmembrane domain and cytoplasmic tail of nectin-1 were not required for any viral receptor or cell adhesion function tested. Cellular cytoplasmic factors that bind to the nectin-1α cytoplasmic tail, therefore, did not influence virus entry or cell fusion. Interestingly, the efficiency of cell fusion was reduced when membrane-spanning domains of nectin-1α and gD were replaced by glycosylphosphatidylinositol tethers, indicating that transmembrane domains may play a modulatory role in the gD/nectin-1α interaction in fusion

  16. Pyridoxal phosphate as a probe of the cytoplasmic domains of transmembrane proteins: Application to the nicotinic acetylcholine receptor

    International Nuclear Information System (INIS)

    Perez-Ramirez, B.; Martinez-Carrion, M.

    1989-01-01

    A novel procedure has been developed to specifically label the cytoplasmic domains of transmembrane proteins with the aldehyde pyridoxal 5-phosphate (PLP). Torpedo californica acetylcholine receptor (AcChR) vesicles were loaded with [ 3 H]pyridoxine 5-phosphate ([ 3 H]PNP) and pyridoxine-5-phosphate oxidase, followed by intravesicular enzymatic oxidation of [ 3 H]PNP at 37 degree C in the presence of externally added cytochrome c as a scavenger of possible leaking PLP product. The four receptor subunits were labeled whether the reaction was carried out on the internal surface or separately designed to mark the external one. On the other hand, the relative pyridoxylation of the subunits differed in both cases, reflecting differences in accessible lysyl residues in each side of the membrane. Even though there are no large differences in the total lysine content among the subunits and there are two copies of the α-subunit, internal surface labeling by PLP was greatest for the highest molecular weight (δ) subunit, reinforcing the concept that the four receptor subunits are transmembranous and may protrude into the cytoplasmic face in a fashion that is proportional to their subunit molecular weight. Yet, the labeling data do not fit well to any of the models proposed for AcChR subunit folding. The method described can be used for selective labeling of the cytoplasmic domains of transmembrane proteins in sealed membrane vesicles

  17. Effects of Chinese yellow wine on nitric oxide synthase and intercellular adhesion molecule-1 expressions in rat vascular endothelial cells.

    Science.gov (United States)

    Zhao, Fei; Ji, Zheng; Chi, Jufang; Tang, Weiliang; Zhai, Xiaoya; Meng, Liping; Guo, Hangyuan

    2016-02-01

    The objective of this study was to determine similarities in the effect of yellow wine as compared to statin and the possibility that yellow wine inhibits tumour necrosis factor-α (TNF-α)-induced nitric oxide (NO) synthesis, endothelial nitric oxide synthase (eNOS), inducible nitric oxide synthase (iNOS), and intercellular adhesion molecule-1 (ICAM-1) in cultured rat vascular endothelial cells (VECs). We isolated VECs, and cultivated and purified Sprague Dawley (SD) rat thoracic aortas in vitro. We selected the optimal wine concentration using clonogenic and MTT assays to measure cell survival. Next, we divided the cells into 9 groups: (1) control, (2) TNF-α, (3) TNF-α + rosuvastatin (10 μmol/L), (4) TNF-α + ethanol 0.5%, (5) TNF-α + yellow wine 0.5%, (6) TNF-α + ethanol 1.0%, (7) TNF-α + yellow wine 1.0%, (8) TNF-α + ethanol 1.5%, and (9) TNF-α + yellow wine 1.5% and they were given the corresponding treatment for 24 h. We determined NO production with nitrate reductase. We then measured eNOS activity, and detected eNOS, iNOS, and ICAM-1 protein levels by Western blotting. Compared with the TNF-α group, NO production, eNOS activity, and eNOS protein expression in the rosuvastatin, and yellow wine 1.0%, and 1.5% groups were significantly increased. Protein expression of iNOS and ICAM-1 in the rosuvastatin, yellow wine 1.0%, and 1.5% groups were significantly decreased. Compared with the rosuvastatin group, eNOS, iNOS, and ICAM-1 protein expression in the yellow wine (0.5% -1.5%) groups were significantly different. Treatment with yellow wine increased NO production, eNOS activity, and eNOS protein expression, which decreases iNOS and ICAM-1 protein expression. We conclude that yellow wine may have similar beneficial effects as rosuvastatin on the cardiovascular system. These effects may be attributed to their anti-atherosclerotic actions.

  18. Phenotype characterization of embryoid body structures generated by a crystal comet effect tail in an intercellular cancer collision scenario

    Directory of Open Access Journals (Sweden)

    Diaz JA

    2012-01-01

    Full Text Available Jairo A Diaz, Mauricio F MurilloDepartment of Pathology, Hospital Departmental Villavicencio, Hospital Departmental Granada, Medicine School, University Cooperative of Colombia, Villavicencio, Meta, ColombiaAbstract: Cancer is, by definition, the uncontrolled growth of autonomous cells that eventually destroy adjacent tissues and generate architectural disorder. However, this concept cannot be totally true. In three well documented studies, we have demonstrated that cancer tissues produce order zones that evolve over time and generate embryoid body structures in a space-time interval. The authors decided to revise the macroscopic and microscopic material in well-developed malignant tumors in which embryoid bodies were identified to determine the phenotype characterization that serves as a guideline for easy recognition. The factors responsible for this morphogenesis are physical, bioelectric, and magnetic susceptibilities produced by crystals that act as molecular designers for the topographic gradients that guide the surrounding silhouette and establish tissue head-tail positional identities. The structures are located in amniotic-like cavities and show characteristic somite-like embryologic segmentation. Immunophenotypic study has demonstrated exclusion factor positional identity in relation to enolase-immunopositive expression of embryoid body and human chorionic gonadotropin immunopositivity exclusion factor expression in the surrounding tissues. The significance of these observations is that they can also be predicted by experimental image data collected by the Large Hadron Collider (LHC accelerator at the European Organization for Nuclear Research, in which two-beam subatomic collision particles in the resulting debris show hyperorder domains similar to those identified by us in intercellular cancer collisions. Our findings suggest that we are dealing with true reverse biologic system information in an activated collective cancer stem cell

  19. Unprecedented multiplicity of Ig transmembrane and secretory mRNA forms in the cartilaginous fish.

    Science.gov (United States)

    Rumfelt, Lynn L; Diaz, Marilyn; Lohr, Rebecca L; Mochon, Evonne; Flajnik, Martin F

    2004-07-15

    In most jawed vertebrates including cartilaginous fish, membrane-bound IgM is expressed as a five Ig superfamily (Igsf)-domain H chain attached to a transmembrane (Tm) region. Heretofore, bony fish IgM was the one exception with IgM mRNA spliced to produce a four-domain Tm H chain. We now demonstrate that the Tm and secretory (Sec) mRNAs of the novel cartilaginous fish Ig isotypes, IgW and IgNAR, are present in multiple forms, most likely generated by alternative splicing. In the nurse shark, Ginglymostoma cirratum, and horn shark, Heterodontus francisci, alternative splicing of Tm exons to the second or the fourth constant (C(H)) exons produces two distinct IgW Tm cDNAs. Although the seven-domain IgW Sec cDNA form contains a canonical secretory tail shared with IgM, IgNAR, and IgA, we report a three-domain cDNA form of shark IgW (IgW(short)) having an unusual Sec tail, which is orthologous to skate IgX(short) cDNA. The IgW and IgW(short) Sec transcripts are restricted in their tissue distribution and expression levels vary among individual sharks, with all forms expressed early in ontogeny. IgNAR mRNA is alternatively spliced to produce a truncated four-domain Tm cDNA and a second Tm cDNA is expressed identical in Igsf domains as the Sec form. PBL is enriched in the Tm cDNA of these Igs. These molecular data suggest that cartilaginous fish have augmented their humoral immune repertoire by diversifying the sizes of their Ig isotypes. Furthermore, these Tm cDNAs are prototypical and the truncated variants may translate as more stable protein at the cell surface.

  20. Modulating Transmembrane α-Helix Interactions through pH-Sensitive Boundary Residues.

    Science.gov (United States)

    Ng, Derek P; Deber, Charles M

    2016-08-09

    Changes in pH can alter the structure and activity of proteins and may be used by the cell to control molecular function. This coupling can also be used in non-native applications through the design of pH-sensitive biomolecules. For example, the pH (low) insertion peptide (pHLIP) can spontaneously insert into a lipid bilayer when the pH decreases. We have previously shown that the α-helicity and helix-helix interactions of the TM2 α-helix of the proteolipid protein (PLP) are sensitive to the local hydrophobicity at its C-terminus. Given that there is an ionizable residue (Glu-88) at the C-terminus of this transmembrane (TM) segment, we hypothesized that changing the ionization state of this residue through pH may alter the local hydrophobicity of the peptide enough to affect both its secondary structure and helix-helix interactions. To examine this phenomenon, we synthesized peptide analogues of the PLP TM2 α-helix (wild-type sequence (66)AFQYVIYGTASFFFLYGALLLAEGF(90)). Using circular dichroism and Förster resonance energy transfer in the membrane-mimetic detergent sodium dodecyl sulfate, we found that a decrease in pH increases both peptide α-helicity and the extent of self-association. This pH-dependent effect is due specifically to the presence of Glu-88 at the C-terminus. Additional experiments in which Phe-90 was mutated to residues of varying hydrophobicities indicated that the strength of this effect is dependent on the local hydrophobicity near Glu-88. Our results have implications for the design of TM peptide switches and improve our understanding of how membrane protein structure and activity can be regulated through local molecular environmental changes.

  1. Structural fragment clustering reveals novel structural and functional motifs in α-helical transmembrane proteins

    Directory of Open Access Journals (Sweden)

    Vassilev Boris

    2010-04-01

    Full Text Available Abstract Background A large proportion of an organism's genome encodes for membrane proteins. Membrane proteins are important for many cellular processes, and several diseases can be linked to mutations in them. With the tremendous growth of sequence data, there is an increasing need to reliably identify membrane proteins from sequence, to functionally annotate them, and to correctly predict their topology. Results We introduce a technique called structural fragment clustering, which learns sequential motifs from 3D structural fragments. From over 500,000 fragments, we obtain 213 statistically significant, non-redundant, and novel motifs that are highly specific to α-helical transmembrane proteins. From these 213 motifs, 58 of them were assigned to function and checked in the scientific literature for a biological assessment. Seventy percent of the motifs are found in co-factor, ligand, and ion binding sites, 30% at protein interaction interfaces, and 12% bind specific lipids such as glycerol or cardiolipins. The vast majority of motifs (94% appear across evolutionarily unrelated families, highlighting the modularity of functional design in membrane proteins. We describe three novel motifs in detail: (1 a dimer interface motif found in voltage-gated chloride channels, (2 a proton transfer motif found in heme-copper oxidases, and (3 a convergently evolved interface helix motif found in an aspartate symporter, a serine protease, and cytochrome b. Conclusions Our findings suggest that functional modules exist in membrane proteins, and that they occur in completely different evolutionary contexts and cover different binding sites. Structural fragment clustering allows us to link sequence motifs to function through clusters of structural fragments. The sequence motifs can be applied to identify and characterize membrane proteins in novel genomes.

  2. Delivering Transmembrane Peptide Complexes to the Gas Phase Using Nanodiscs and Electrospray Ionization

    Science.gov (United States)

    Li, Jun; Richards, Michele R.; Kitova, Elena N.; Klassen, John S.

    2017-10-01

    The gas-phase conformations of dimers of the channel-forming membrane peptide gramicidin A (GA), produced from isobutanol or aqueous solutions of GA-containing nanodiscs (NDs), are investigated using electrospray ionization-ion mobility separation-mass spectrometry (ESI-IMS-MS) and molecular dynamics (MD) simulations. The IMS arrival times measured for (2GA + 2Na)2+ ions from isobutanol reveal three different conformations, with collision cross-sections (Ω) of 683 Å2 (conformation 1, C1), 708 Å2 (C2), and 737 Å2 (C3). The addition of NH4CH3CO2 produced (2GA + 2Na)2+ and (2GA + H + Na)2+ ions, with Ω similar to those of C1, C2, and C3, as well as (2GA + 2H)2+, (2GA + 2NH4)2+, and (2GA + H + NH4)2+ ions, which adopt a single conformation with a Ω similar to that of C2. These results suggest that the nature of the charging agents, imparted by the ESI process, can influence dimer conformation in the gas phase. Notably, the POPC NDs produced exclusively (2GA + 2NH4)2+ dimer ions; the DMPC NDs produced both (2GA + 2H)2+ and (2GA + 2NH4)2+ dimer ions. While the Ω of (2GA + 2H)2+ is similar to that of C2, the (2GA + 2NH4)2+ ions from NDs adopt a more compact structure, with a Ω of 656 Å2. It is proposed that this compact structure corresponds to the ion conducting single stranded head-to-head helical GA dimer. These findings highlight the potential of NDs, combined with ESI, for transferring transmembrane peptide complexes directly from lipid bilayers to the gas phase. [Figure not available: see fulltext.

  3. Transmembrane Prostatic Acid Phosphatase (TMPAP) Interacts with Snapin and Deficient Mice Develop Prostate Adenocarcinoma

    Science.gov (United States)

    Quintero, Ileana B.; Herrala, Annakaisa M.; Araujo, César L.; Pulkka, Anitta E.; Hautaniemi, Sampsa; Ovaska, Kristian; Pryazhnikov, Evgeny; Kulesskiy, Evgeny; Ruuth, Maija K.; Soini, Ylermi; Sormunen, Raija T.; Khirug, Leonard; Vihko, Pirkko T.

    2013-01-01

    The molecular mechanisms underlying prostate carcinogenesis are poorly understood. Prostatic acid phosphatase (PAP), a prostatic epithelial secretion marker, has been linked to prostate cancer since the 1930's. However, the contribution of PAP to the disease remains controversial. We have previously cloned and described two isoforms of this protein, a secretory (sPAP) and a transmembrane type-I (TMPAP). The goal in this work was to understand the physiological function of TMPAP in the prostate. We conducted histological, ultra-structural and genome-wide analyses of the prostate of our PAP-deficient mouse model (PAP−/−) with C57BL/6J background. The PAP−/− mouse prostate showed the development of slow-growing non-metastatic prostate adenocarcinoma. In order to find out the mechanism behind, we identified PAP-interacting proteins byyeast two-hybrid assays and a clear result was obtained for the interaction of PAP with snapin, a SNARE-associated protein which binds Snap25 facilitating the vesicular membrane fusion process. We confirmed this interaction by co-localization studies in TMPAP-transfected LNCaP cells (TMPAP/LNCaP cells) and in vivo FRET analyses in transient transfected LNCaP cells. The differential gene expression analyses revealed the dysregulation of the same genes known to be related to synaptic vesicular traffic. Both TMPAP and snapin were detected in isolated exosomes. Our results suggest that TMPAP is involved in endo-/exocytosis and disturbed vesicular traffic is a hallmark of prostate adenocarcinoma. PMID:24039861

  4. Cystic fibrosis transmembrane regulator haplotypes in households of patients with cystic fibrosis.

    Science.gov (United States)

    Furgeri, Daniela Tenório; Marson, Fernando Augusto Lima; Correia, Cyntia Arivabeni Araújo; Ribeiro, José Dirceu; Bertuzzo, Carmen Sílvia

    2018-01-30

    Nearly 2000 mutations in the cystic fibrosis transmembrane regulator (CFTR) gene have been reported. The F508del mutation occurs in approximately 50-65% of patients with cystic fibrosis (CF). However, molecular diagnosis is not always possible. Therefore, silent polymorphisms can be used to label the mutant allele in households of patients with CF. To verify the haplotypes of four polymorphisms at the CFTR locus in households of patients with CF for pre-fertilization, pre-implantation, and prenatal indirect mutation diagnosis to provide better genetic counseling for families and patients with CF and to associate the genotypes/haplotypes with the F508del mutation screening. GATT polymorphism analysis was performed using direct polymerase chain reaction amplification, and the MP6-D9, TUB09 and TUB18 polymorphism analyses were performed using restriction fragment length polymorphism. Nine haplotypes were found in 37 CFTR alleles, and of those, 24 were linked with the F508del mutation and 13 with other CFTR mutations. The 6 (GATT), C (MP6-D9), G (TUB09), and C (TUB18) haplotypes showed the highest prevalence (48%) of the mutant CFTR allele and were linked to the F508del mutation (64%). In 43% of households analyzed, at least one informative polymorphism can be used for the indirect diagnostic test. CFTR polymorphisms are genetic markers that are useful for identifying the mutant CFTR alleles in households of patients with CF when it is not possible to establish the complete CFTR genotype. Moreover, the polymorphisms can be used for indirect CFTR mutation identification in cases of pre-fertilization, pre-implantation and prenatal analysis. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Determination of Six Transmembrane Protein of Prostate 2 Gene Expression and Intracellular Localization in Prostate Cancer

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    Bora İrer

    2017-12-01

    Full Text Available Objective: In this study, we aimed to determine the relationship between the RNA and protein expression profile of six transmembrane protein of prostate 2 (STAMP2 gene and androgen and the intracellular localization of STAMP2. Materials and Methods: RNA and protein were obtained from androgen treated lymph node carcinoma of the prostate (LNCaP cells, untreated LNCaP cells, DU145 cells with no androgen receptor, and STAMP2 transfected COS-7 cells. The expression profile of STAMP2 gene and the effect of androgenes on the expression was shown in RNA and protein levels by using Northern and Western blotting methods. In addition, intracellular localization of the naturally synthesized STAMP2 protein and the transfected STAMP2 protein in COS-7 cells after androgen administration in both LNCaP cells was determined by immunofluorescence microscopy. Results: We found that the RNA and protein expression of STAMP2 gene in LNCaP cells are regulated by androgenes, the power of expression is increased with the duration of androgen treatment and there is no STAMP2 expression in DU145 cells which has no androgen receptor. As a result of the immunofluorescence microscopy study we observed that STAMP2 protein was localized at golgi complex and cell membrane. Conclusion: In conclusion, we have demonstrated that STAMP2 may play an important role in the pathogenesis of the prostate cancer and in the androgen-dependent androgen-independent staging of prostate cancer. In addition, STAMP2 protein, which is localized in the intracellular golgi complex and cell membrane, may be a new target molecule for prostate cancer diagnosis and treatment.

  6. Function and expression of cystic fibrosis transmembrane conductance regulator after small intestinal transplantation in mice.

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    Penghong Song

    Full Text Available The secretion function of intestinal graft is one of the most important factors for successful intestinal transplantation. Cystic fibrosis transmembrane conductance regulator (CFTR mediates HCO3(- and Cl(- secretions in intestinal epithelial cells. In this study, we made investigation on the expression and function of CFTR in an experimental model of murine small intestinal transplantation. Heterotopic intestinal transplantations were performed in syngeneic mice. The mRNA and protein expressions of CFTR were analyzed by real time PCR and western blot. Murine intestinal mucosal HCO3(- and Cl(- secretions were examined in vitro in Ussing chambers by the pH stat and short circuit current (I(sc techniques. The results showed that forskolin, an activator of CFTR, stimulated jejunal mucosal epithelial HCO3(- and Cl(- secretions in mice, but forskolin-stimulated HCO3(- and Cl(- secretions in donor and recipient jejunal mucosae of mice after heterotopic jejunal transplantation were markedly decreased, compared with controls (P<0.001. The mRNA and protein expression levels of CFTR in donor and recipient jejunal mucosae of mice were also markedly lower than those in controls (P<0.001, and the mRNA and protein expression levels of tumor necrosis factor α (TNFα were markedly increased in donor jejunal mucosae of mice (P<0.001, compared with controls. Further experiments showed that TNFα down-regulated the expression of CFTR mRNA in murine jejunal mucosa. In conclusion, after intestinal transplantation, the function of CFTR was impaired, and its mRNA and protein expressions were down-regulated, which may be induced by TNFα.

  7. Sphingosine-1-Phosphate Is a Novel Regulator of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR Activity.

    Directory of Open Access Journals (Sweden)

    Firhan A Malik

    Full Text Available The cystic fibrosis transmembrane conductance regulator (CFTR attenuates sphingosine-1-phosphate (S1P signaling in resistance arteries and has emerged as a prominent regulator of myogenic vasoconstriction. This investigation demonstrates that S1P inhibits CFTR activity via adenosine monophosphate-activated kinase (AMPK, establishing a potential feedback link. In Baby Hamster Kidney (BHK cells expressing wild-type human CFTR, S1P (1μmol/L attenuates forskolin-stimulated, CFTR-dependent iodide efflux. S1P's inhibitory effect is rapid (within 30 seconds, transient and correlates with CFTR serine residue 737 (S737 phosphorylation. Both S1P receptor antagonism (4μmol/L VPC 23019 and AMPK inhibition (80μmol/L Compound C or AMPK siRNA attenuate S1P-stimluated (i AMPK phosphorylation, (ii CFTR S737 phosphorylation and (iii CFTR activity inhibition. In BHK cells expressing the ΔF508 CFTR mutant (CFTRΔF508, the most common mutation causing cystic fibrosis, both S1P receptor antagonism and AMPK inhibition enhance CFTR activity, without instigating discernable correction. In summary, we demonstrate that S1P/AMPK signaling transiently attenuates CFTR activity. Since our previous work positions CFTR as a negative S1P signaling regulator, this signaling link may positively reinforce S1P signals. This discovery has clinical ramifications for the treatment of disease states associated with enhanced S1P signaling and/or deficient CFTR activity (e.g. cystic fibrosis, heart failure. S1P receptor/AMPK inhibition could synergistically enhance the efficacy of therapeutic strategies aiming to correct aberrant CFTR trafficking.

  8. Epithelial Cell–Derived Secreted and Transmembrane 1a Signals to Activated Neutrophils during Pneumococcal Pneumonia

    Science.gov (United States)

    Kamata, Hirofumi; Yamamoto, Kazuko; Wasserman, Gregory A.; Zabinski, Mary C.; Yuen, Constance K.; Lung, Wing Yi; Gower, Adam C.; Belkina, Anna C.; Ramirez, Maria I.; Deng, Jane C.; Quinton, Lee J.; Jones, Matthew R.

    2016-01-01

    Airway epithelial cell responses are critical to the outcome of lung infection. In this study, we aimed to identify unique contributions of epithelial cells during lung infection. To differentiate genes induced selectively in epithelial cells during pneumonia, we compared genome-wide expression profiles from three sorted cell populations: epithelial cells from uninfected mouse lungs, epithelial cells from mouse lungs with pneumococcal pneumonia, and nonepithelial cells from those same infected lungs. Of 1,166 transcripts that were more abundant in epithelial cells from infected lungs compared with nonepithelial cells from the same lungs or from epithelial cells of uninfected lungs, 32 genes were identified as highly expressed secreted products. Especially strong signals included two related secreted and transmembrane (Sectm) 1 genes, Sectm1a and Sectm1b. Refinement of sorting strategies suggested that both Sectm1 products were induced predominantly in conducting airway epithelial cells. Sectm1 was induced during the early stages of pneumococcal pneumonia, and mutation of NF-κB RelA in epithelial cells did not diminish its expression. Instead, type I IFN signaling was necessary and sufficient for Sectm1 induction in lung epithelial cells, mediated by signal transducer and activator of transcription 1. For target cells, Sectm1a bound to myeloid cells preferentially, in particular Ly6GbrightCD11bbright neutrophils in the infected lung. In contrast, Sectm1a did not bind to neutrophils from uninfected lungs. Sectm1a increased expression of the neutrophil-attracting chemokine CXCL2 by neutrophils from the infected lung. We propose that Sectm1a is an epithelial product that sustains a positive feedback loop amplifying neutrophilic inflammation during pneumococcal pneumonia. PMID:27064756

  9. Epithelial Cell-Derived Secreted and Transmembrane 1a Signals to Activated Neutrophils during Pneumococcal Pneumonia.

    Science.gov (United States)

    Kamata, Hirofumi; Yamamoto, Kazuko; Wasserman, Gregory A; Zabinski, Mary C; Yuen, Constance K; Lung, Wing Yi; Gower, Adam C; Belkina, Anna C; Ramirez, Maria I; Deng, Jane C; Quinton, Lee J; Jones, Matthew R; Mizgerd, Joseph P

    2016-09-01

    Airway epithelial cell responses are critical to the outcome of lung infection. In this study, we aimed to identify unique contributions of epithelial cells during lung infection. To differentiate genes induced selectively in epithelial cells during pneumonia, we compared genome-wide expression profiles from three sorted cell populations: epithelial cells from uninfected mouse lungs, epithelial cells from mouse lungs with pneumococcal pneumonia, and nonepithelial cells from those same infected lungs. Of 1,166 transcripts that were more abundant in epithelial cells from infected lungs compared with nonepithelial cells from the same lungs or from epithelial cells of uninfected lungs, 32 genes were identified as highly expressed secreted products. Especially strong signals included two related secreted and transmembrane (Sectm) 1 genes, Sectm1a and Sectm1b. Refinement of sorting strategies suggested that both Sectm1 products were induced predominantly in conducting airway epithelial cells. Sectm1 was induced during the early stages of pneumococcal pneumonia, and mutation of NF-κB RelA in epithelial cells did not diminish its expression. Instead, type I IFN signaling was necessary and sufficient for Sectm1 induction in lung epithelial cells, mediated by signal transducer and activator of transcription 1. For target cells, Sectm1a bound to myeloid cells preferentially, in particular Ly6G(bright)CD11b(bright) neutrophils in the infected lung. In contrast, Sectm1a did not bind to neutrophils from uninfected lungs. Sectm1a increased expression of the neutrophil-attracting chemokine CXCL2 by neutrophils from the infected lung. We propose that Sectm1a is an epithelial product that sustains a positive feedback loop amplifying neutrophilic inflammation during pneumococcal pneumonia.

  10. Cystic Fibrosis Transmembrane Regulator Modulators: Implications for the Management of Depression and Anxiety in Cystic Fibrosis.

    Science.gov (United States)

    Talwalkar, Jaideep S; Koff, Jonathan L; Lee, Hochang B; Britto, Clemente J; Mulenos, Arielle M; Georgiopoulos, Anna M

    Individuals with cystic fibrosis (CF) are at high risk for depression and anxiety, which are associated with worse medical outcomes. Novel therapies for CF hold great promise for improving physical health, but the effects of these therapies on mental health remain poorly understood. This review aims to familiarize psychiatrists with the potential effect of novel CF therapies on depression and anxiety. We discuss novel therapies that directly target the mutant CF protein, the CF transmembrane regulator (CFTR), which are called CFTR modulators. We summarize depression and anxiety screening and treatment guidelines under implementation in accredited CF centers. Case vignettes highlight the complexities of caring for individuals with CF with comorbid depression and anxiety, including patients experiencing worsening depression and anxiety proximate to initiation of CFTR modulator therapy, and management of drug-drug interactions. Although CFTR modulator therapies provide hope for improving clinical outcomes, worsening depression and anxiety occurs in some patients when starting these novel agents. This phenomenon may be multifactorial, with hypothesized contributions from CFTR modulator-psychotropic medication interactions, direct effects of CFTR modulators on central nervous system function, the psychologic effect of starting a potentially life-altering drug, and typical triggers of depression and anxiety such as stress, pain, and inflammation. The medical and psychiatric complexity of many individuals with CF warrants more direct involvement of mental health specialists on the multidisciplinary CF team. Inclusion of mental health variables in patients with CF registries will facilitate further examination at an epidemiologic level. Copyright © 2017 The Academy of Psychosomatic Medicine. Published by Elsevier Inc. All rights reserved.

  11. Cystic fibrosis transmembrane conductance regulator contributes to reacidification of alkalinized lysosomes in RPE cells.

    Science.gov (United States)

    Liu, Ji; Lu, Wennan; Guha, Sonia; Baltazar, Gabriel C; Coffey, Erin E; Laties, Alan M; Rubenstein, Ronald C; Reenstra, William W; Mitchell, Claire H

    2012-07-15

    The role of the cystic fibrosis transmembrane conductance regulator (CFTR) in lysosomal acidification has been difficult to determine. We demonstrate here that CFTR contributes more to the reacidification of lysosomes from an elevated pH than to baseline pH maintenance. Lysosomal alkalinization is increasingly recognized as a factor in diseases of accumulation, and we previously showed that cAMP reacidified alkalinized lysosomes in retinal pigmented epithelial (RPE) cells. As the influx of anions to electrically balance proton accumulation may enhance lysosomal acidification, the contribution of the cAMP-activated anion channel CFTR to lysosomal reacidification was probed. The antagonist CFTR(inh)-172 had little effect on baseline levels of lysosomal pH in cultured human RPE cells but substantially reduced the reacidification of compromised lysosomes by cAMP. Likewise, CFTR activators had a bigger impact on cells whose lysosomes had been alkalinized. Knockdown of CFTR with small interfering RNA had a larger effect on alkalinized lysosomes than on baseline levels. Inhibition of CFTR in isolated lysosomes altered pH. While CFTR and Lamp1 were colocalized, treatment with cAMP did not increase targeting of CFTR to the lysosome. The inhibition of CFTR slowed lysosomal degradation of photoreceptor outer segments while activation of CFTR enhanced their clearance from compromised lysosomes. Activation of CFTR acidified RPE lysosomes from the ABCA4(-/-) mouse model of recessive Stargardt's disease, whose lysosomes are considerably alkalinized. In summary, CFTR contributes more to reducing lysosomal pH from alkalinized levels than to maintaining baseline pH. Treatment to activate CFTR may thus be of benefit in disorders of accumulation associated with lysosomal alkalinization.

  12. Influence of magnesium sulfate on HCO3/Cl transmembrane exchange rate in human erythrocytes.

    Science.gov (United States)

    Chernyshova, Ekaterina S; Zaikina, Yulia S; Tsvetovskaya, Galina A; Strokotov, Dmitry I; Yurkin, Maxim A; Serebrennikova, Elena S; Volkov, Leonid; Maltsev, Valeri P; Chernyshev, Andrei V

    2016-03-21

    Magnesium sulfate (MgSO4) is widely used in medicine but molecular mechanisms of its protection through influence on erythrocytes are not fully understood and are considerably controversial. Using scanning flow cytometry, in this work for the first time we observed experimentally (both in situ and in vitro) a significant increase of HCO3(-)/Cl(-) transmembrane exchange rate of human erythrocytes in the presence of MgSO4 in blood. For a quantitative analysis of the obtained experimental data, we introduced and verified a molecular kinetic model, which describes activation of major anion exchanger Band 3 (or AE1) by its complexation with free intracellular Mg(2+) (taking into account Mg(2+) membrane transport and intracellular buffering). Fitting the model to our in vitro experimental data, we observed a good correspondence between theoretical and experimental kinetic curves that allowed us to evaluate the model parameters and to estimate for the first time the association constant of Mg(2+) with Band 3 as KB~0.07mM, which is in agreement with known values of the apparent Mg(2+) dissociation constant (from 0.01 to 0.1mM) that reflects experiments on enrichment of Mg(2+) at the inner erythrocyte membrane (Gunther, 2007). Results of this work partly clarify the molecular mechanisms of MgSO4 action in human erythrocytes. The method developed allows one to estimate quantitatively a perspective of MgSO4 treatment for a patient. It should be particularly helpful in prenatal medicine for early detection of pathologies associated with the risk of fetal hypoxia. Copyright © 2016 Elsevier Ltd. All rights reserved.

  13. Relevance of lysine snorkeling in the outer transmembrane domain of small viral potassium ion channels.

    Science.gov (United States)

    Gebhardt, Manuela; Henkes, Leonhard M; Tayefeh, Sascha; Hertel, Brigitte; Greiner, Timo; Van Etten, James L; Baumeister, Dirk; Cosentino, Cristian; Moroni, Anna; Kast, Stefan M; Thiel, Gerhard

    2012-07-17

    Transmembrane domains (TMDs) are often flanked by Lys or Arg because they keep their aliphatic parts in the bilayer and their charged groups in the polar interface. Here we examine the relevance of this so-called "snorkeling" of a cationic amino acid, which is conserved in the outer TMD of small viral K(+) channels. Experimentally, snorkeling activity is not mandatory for Kcv(PBCV-1) because K29 can be replaced by most of the natural amino acids without any corruption of function. Two similar channels, Kcv(ATCV-1) and Kcv(MT325), lack a cytosolic N-terminus, and neutralization of their equivalent cationic amino acids inhibits their function. To understand the variable importance of the cationic amino acids, we reanalyzed molecular dynamics simulations of Kcv(PBCV-1) and N-terminally truncated mutants; the truncated mutants mimic Kcv(ATCV-1) and Kcv(MT325). Structures were analyzed with respect to membrane positioning in relation to the orientation of K29. The results indicate that the architecture of the protein (including the selectivity filter) is only weakly dependent on TMD length and protonation of K29. The penetration depth of Lys in a given protonation state is independent of the TMD architecture, which leads to a distortion of shorter proteins. The data imply that snorkeling can be important for K(+) channels; however, its significance depends on the architecture of the entire TMD. The observation that the most severe N-terminal truncation causes the outer TMD to move toward the cytosolic side suggests that snorkeling becomes more relevant if TMDs are not stabilized in the membrane by other domains.

  14. Distinct neurobehavioural effects of cannabidiol in transmembrane domain neuregulin 1 mutant mice.

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    Leonora E Long

    Full Text Available The cannabis constituent cannabidiol (CBD possesses anxiolytic and antipsychotic properties. We have previously shown that transmembrane domain neuregulin 1 mutant (Nrg1 TM HET mice display altered neurobehavioural responses to the main psychoactive constituent of cannabis, Δ(9-tetrahydrocannabinol. Here we investigated whether Nrg1 TM HET mice respond differently to CBD and whether CBD reverses schizophrenia-related phenotypes expressed by these mice. Adult male Nrg1 TM HET and wild type-like littermates (WT received vehicle or CBD (1, 50 or 100 mg/kg i.p. for 21 days. During treatment and 48 h after withdrawal we measured behaviour, whole blood CBD concentrations and autoradiographic receptor binding. Nrg1 HET mice displayed locomotor hyperactivity, PPI deficits and reduced 5-HT(2A receptor binding density in the substantia nigra, but these phenotypes were not reversed by CBD. However, long-term CBD (50 and 100 mg/kg selectively enhanced social interaction in Nrg1 TM HET mice. Furthermore, acute CBD (100 mg/kg selectively increased PPI in Nrg1 TM HET mice, although tolerance to this effect was manifest upon repeated CBD administration. Long-term CBD (50 mg/kg also selectively increased GABA(A receptor binding in the granular retrosplenial cortex in Nrg1 TM HET mice and reduced 5-HT(2A binding in the substantia nigra in WT mice. Nrg1 appears necessary for CBD-induced anxiolysis since only WT mice developed decreased anxiety-related behaviour with repeated CBD treatment. Altered pharmacokinetics in mutant mice could not explain our findings since no genotype differences existed in CBD blood concentrations. Here we demonstrate that Nrg1 modulates acute and long-term neurobehavioural effects of CBD, which does not reverse the schizophrenia-relevant phenotypes.

  15. SAXS analysis of a soluble cytosolic NgBR construct including extracellular and transmembrane domains.

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    Joshua Holcomb

    Full Text Available The Nogo-B receptor (NgBR is involved in oncogenic Ras signaling through directly binding to farnesylated Ras. It recruits farnesylated Ras to the non-lipid-raft membrane for interaction with downstream effectors. However, the cytosolic domain of NgBR itself is only partially folded. The lack of several conserved secondary structural elements makes this domain unlikely to form a complete farnesyl binding pocket. We find that inclusion of the extracellular and transmembrane domains that contain additional conserved residues to the cytosolic region results in a well folded protein with a similar size and shape to the E.coli cis-isoprenyl transferase (UPPs. Small Angle X-ray Scattering (SAXS analysis reveals the radius of gyration (Rg of our NgBR construct to be 18.2 Å with a maximum particle dimension (Dmax of 61.0 Å. Ab initio shape modeling returns a globular molecular envelope with an estimated molecular weight of 23.0 kD closely correlated with the calculated molecular weight. Both Kratky plot and pair distribution function of NgBR scattering reveal a bell shaped peak which is characteristic of a single globularly folded protein. In addition, circular dichroism (CD analysis reveals that our construct has the secondary structure contents similar to the UPPs. However, this result does not agree with the currently accepted topological orientation of NgBR which might partition this construct into three separate domains. This discrepancy suggests another possible NgBR topology and lends insight into a potential molecular basis of how NgBR facilitates farnesylated Ras recruitment.

  16. Secreted and transmembrane 1A is a novel co-stimulatory ligand.

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    Duncan Howie

    Full Text Available Most T cell responses to pathogens or self antigens are modulated through the action of regulatory T cells and tissue-specific inhibitory mechanisms. To this end, several receptor-ligand pairs have evolved which either augment or diminish T cell function. Here we describe the tissue ligand SECTM1A (Secreted and transmembrane1A as an alternative murine CD7 ligand. We show that SECTM1A, like SECTM1B, binds strongly to CD7, and that SECTM1B was able to compete with SECTM1A for CD7 binding. SECTM1A is ubiquitously expressed and has two major alternative transcripts which differ in expression between tissues. Both immobilised soluble forms of SECTM1A and SECTM1B and cell surface anchored forms demonstrated opposing effects on CD4+ T cell activation. Whereas SECTM1A acted as a co-stimulator of T cells, enhancing IL-2 production and proliferation, SECTM1B proved inhibitory to TCR mediated T cell activation. Surprisingly, both functional outcomes proved to be CD7-independent, indicating the existence of alternative receptors for both ligands. We used a SECTM1A-Fc fusion protein to immunoprecipitate potential alternative ligands from detergent lysates of CD7(-/- T cells and, using mass spectrometry, identified GITR as a SECTM1A binder. SECTM1A was found to bind to activated CD4+ and CD8+ T cells as well as to CHO cells expressing cell surface GITR. Binding of SECTM1A to activated primary T cells was inhibited by either GITRL-Fc or anti GITR antibodies. Thus SECTM1A and SECTM1B represent novel reciprocal alternative ligands which may function to modulate the activation of effector and regulatory T cells. The ability of SECTM1A to activate T cells may be explained by its ability to bind to GITR.

  17. Science communication as political communication

    Science.gov (United States)

    Scheufele, Dietram A.

    2014-01-01

    Scientific debates in modern societies often blur the lines between the science that is being debated and the political, moral, and legal implications that come with its societal applications. This manuscript traces the origins of this phenomenon to professional norms within the scientific discipline and to the nature and complexities of modern science and offers an expanded model of science communication that takes into account the political contexts in which science communication takes place. In a second step, it explores what we know from empirical work in political communication, public opinion research, and communication research about the dynamics that determine how issues are debated and attitudes are formed in political environments. Finally, it discusses how and why it will be increasingly important for science communicators to draw from these different literatures to ensure that the voice of the scientific community is heard in the broader societal debates surrounding science. PMID:25225389

  18. Science communication as political communication.

    Science.gov (United States)

    Scheufele, Dietram A

    2014-09-16

    Scientific debates in modern societies often blur the lines between the science that is being debated and the political, moral, and legal implications that come with its societal applications. This manuscript traces the origins of this phenomenon to professional norms within the scientific discipline and to the nature and complexities of modern science and offers an expanded model of science communication that takes into account the political contexts in which science communication takes place. In a second step, it explores what we know from empirical work in political communication, public opinion research, and communication research about the dynamics that determine how issues are debated and attitudes are formed in political environments. Finally, it discusses how and why it will be increasingly important for science communicators to draw from these different literatures to ensure that the voice of the scientific community is heard in the broader societal debates surrounding science.

  19. Satellite Communications

    CERN Document Server

    Pelton, Joseph N

    2012-01-01

    The field of satellite communications represents the world's largest space industry. Those who are interested in space need to understand the fundamentals of satellite communications, its technology, operation, business, economic, and regulatory aspects. This book explains all this along with key insights into the field's future growth trends and current strategic challenges. Fundamentals of Satellite Communications is a concise book that gives all of the key facts and figures as well as a strategic view of where this dynamic industry is going. Author Joseph N. Pelton, PhD, former Dean of the International Space University and former Director of Strategic Policy at Intelstat, presents a r

  20. Short communication

    African Journals Online (AJOL)

    abp

    2017-09-04

    Sep 4, 2017 ... Face-to-face interviews were conducted using a standardized ... Short communication. Open Access ... clinic during the time of the study and were invited to participate in the study. .... consume them. This is another ...