Shaping the tracks : Regulation of microtubule dynamics by kinesins KIF21A and KIF21B

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van Riel, W.E.|info:eu-repo/dai/nl/338772634

2016-01-01

Control of microtubule dynamics is important for cell morphogenesis. Kinesins, motor proteins known to function in cargo transport, were recently also implicated in altering the microtubule network. Several kinesins are described to cause microtubule network reorganization or stabilization, either

Intracellular Transport and Kinesin Superfamily Proteins: Structure, Function and Dynamics

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Hirokawa, N.; Takemura, R.

Using various molecular cell biological and molecular genetic approaches, we identified kinesin superfamily proteins (KIFs) and characterized their significant functions in intracellular transport, which is fundamental for cellular morphogenesis, functioning, and survival. We showed that KIFs not only transport various membranous organelles, proteins complexes and mRNAs fundamental for cellular functions but also play significant roles in higher brain functions such as memory and learning, determination of important developmental processes such as left-right asymmetry formation and brain wiring. We also elucidated that KIFs recognize and bind to their specific cargoes using scaffolding or adaptor protein complexes. Concerning the mechanism of motility, we discovered the simplest unique monomeric motor KIF1A and determined by molecular biophysics, cryoelectron microscopy and X-ray crystallography that KIF1A can move on a microtubule processively as a monomer by biased Brownian motion and by hydolyzing ATP.

Identification of two novel KIF5A mutations in hereditary spastic paraplegia associated with mild peripheral neuropathy.

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López, Eva; Casasnovas, Carlos; Giménez, Javier; Santamaría, Raúl; Terrazas, Jesús M; Volpini, Víctor

2015-11-15

Spastic paraplegia type 10 (SPG10) is a rare form of autosomal dominant hereditary spastic paraplegia (AD-HSP) due to mutations in KIF5A, a gene encoding the neuronal kinesin heavy-chain involved in axonal transport. KIF5A mutations have been associated with a wide clinical spectrum, ranging from pure HSP to isolated peripheral nerve involvement or complicated HSP phenotypes. Most KIF5A mutations are clustered in the motor domain of the protein that is necessary for microtubule interaction. Here we describe two Spanish families with an adult onset complicated AD-HSP in which neurological studies revealed a mild sensory neuropathy. Intention tremor was also present in both families. Molecular genetic analysis identified two novel mutations c.773 C>T and c.833 C>T in the KIF5A gene resulting in the P258L and P278L substitutions respectively. Both were located in the highly conserved kinesin motor domain of the protein which has previously been identified as a hot spot for KIF5A mutations. This study adds to the evidence associating the known occurrence of mild peripheral neuropathy in the adult onset SPG10 type of AD-HSP. Copyright © 2015 Elsevier B.V. All rights reserved.

Interdependency and phosphorylation of KIF4 and condensin I are essential for organization of chromosome scaffold.

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Rawin Poonperm

Full Text Available Kinesin family member 4 (KIF4 and condensins I and II are essential chromosomal proteins for chromosome organization by locating primarily to the chromosome scaffold. However, the mechanism of how KIF4 and condensins localize to the chromosome scaffold is poorly understood. Here, we demonstrate a close relationship between the chromosome localization of KIF4 and condensin I, but not condensin II, and show that KIF4 and condensin I assist each other for stable scaffold formation by forming a stable complex. Moreover, phosphorylation of KIF4 and condensin I by Aurora B and polo-like kinase 1 (Plk1 is important for KIF4 and condensin I localization to the chromosome. Aurora B activity facilitates the targeting of KIF4 and condensin I to the chromosome, whereas Plk1 activity promotes the dissociation of these proteins from the chromosome. Thus, the interdependency between KIF4 and condensin I, and their phosphorylation states play important roles in chromosome scaffold organization during mitosis.

Interdependency and phosphorylation of KIF4 and condensin I are essential for organization of chromosome scaffold.

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Poonperm, Rawin; Takata, Hideaki; Uchiyama, Susumu; Fukui, Kiichi

2017-01-01

Kinesin family member 4 (KIF4) and condensins I and II are essential chromosomal proteins for chromosome organization by locating primarily to the chromosome scaffold. However, the mechanism of how KIF4 and condensins localize to the chromosome scaffold is poorly understood. Here, we demonstrate a close relationship between the chromosome localization of KIF4 and condensin I, but not condensin II, and show that KIF4 and condensin I assist each other for stable scaffold formation by forming a stable complex. Moreover, phosphorylation of KIF4 and condensin I by Aurora B and polo-like kinase 1 (Plk1) is important for KIF4 and condensin I localization to the chromosome. Aurora B activity facilitates the targeting of KIF4 and condensin I to the chromosome, whereas Plk1 activity promotes the dissociation of these proteins from the chromosome. Thus, the interdependency between KIF4 and condensin I, and their phosphorylation states play important roles in chromosome scaffold organization during mitosis.

Stable knockdown of Kif5b in MDCK cells leads to epithelial–mesenchymal transition

International Nuclear Information System (INIS)

Cui, Ju; Jin, Guoxiang; Yu, Bin; Wang, Zai; Lin, Raozhou; Huang, Jian-Dong

2015-01-01

Polarization of epithelial cells requires vectorial sorting and transport of polarity proteins to apical or basolateral domains. Kif5b is the mouse homologue of the human ubiquitous Kinesin Heavy Chain (uKHC). To investigate the function of Kif5b in epithelial cells, we examined the phenotypes of Kif5b-deficient MDCK cells. Stable knockdown of Kif5b in MDCK cells resulted in reduced cell proliferation rate, profound changes in cell morphology, loss of epithelial cell marker, and gain of mesenchymal marker, as well as increased cell migration, invasion, and tumorigenesis abilities. E-cadherin and NMMIIA could interact with Kif5b in polarized MDCK cells, and their expression levels were decreased in Kif5b-deficient MDCK cells. Overexpression of E-cadherin and NMMIIA in Kif5b depleted MDCK cells could decrease mesenchymal marker expression and cell migration ability. These results indicate that stable knockdown of Kif5b in MDCK cells can lead to epithelial–mesenchymal transition, which is mediated by defective E-cadherin and NMMIIA expression. - Highlights: • Knockdown of Kif5b in MDCK cells resulted in reduced cell proliferation rate. • Kif5b deficient MDCK cells underwent epithelial–mesenchymal transition. • E-cadherin and NMMIIA could interact with Kif5b in polarized MDCK cells. • Decreased E-cadherin and NMMIIA levels mediate EMT in Kif5b deficient MDCK cells. • Overexpression of E-cadherin and NMMIIA reverse the effects of Kif5b knockdown

Stable knockdown of Kif5b in MDCK cells leads to epithelial–mesenchymal transition

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Cui, Ju, E-mail: juzi.cui@gmail.com [The Key Laboratory of Geriatrics, Beijing Hospital & Beijing Institute of Geriatrics, Ministry of Health, Beijing (China); Department of Biochemistry, LKS Faculty of Medicine, The University of Hong Kong, Hong Kong SAR (China); Jin, Guoxiang; Yu, Bin [Department of Biochemistry, LKS Faculty of Medicine, The University of Hong Kong, Hong Kong SAR (China); Wang, Zai [Department of Biochemistry, LKS Faculty of Medicine, The University of Hong Kong, Hong Kong SAR (China); Institute of Clinical Medical Sciences, China-Japan Friendship Hospital, Beijing (China); Lin, Raozhou [Department of Biochemistry, LKS Faculty of Medicine, The University of Hong Kong, Hong Kong SAR (China); Huang, Jian-Dong, E-mail: jdhuang@hku.hk [Department of Biochemistry, LKS Faculty of Medicine, The University of Hong Kong, Hong Kong SAR (China); The Centre for Synthetic Biology Engineering Research, Shenzhen Institutes of Advanced Technology, Shenzhen (China)

2015-07-17

Polarization of epithelial cells requires vectorial sorting and transport of polarity proteins to apical or basolateral domains. Kif5b is the mouse homologue of the human ubiquitous Kinesin Heavy Chain (uKHC). To investigate the function of Kif5b in epithelial cells, we examined the phenotypes of Kif5b-deficient MDCK cells. Stable knockdown of Kif5b in MDCK cells resulted in reduced cell proliferation rate, profound changes in cell morphology, loss of epithelial cell marker, and gain of mesenchymal marker, as well as increased cell migration, invasion, and tumorigenesis abilities. E-cadherin and NMMIIA could interact with Kif5b in polarized MDCK cells, and their expression levels were decreased in Kif5b-deficient MDCK cells. Overexpression of E-cadherin and NMMIIA in Kif5b depleted MDCK cells could decrease mesenchymal marker expression and cell migration ability. These results indicate that stable knockdown of Kif5b in MDCK cells can lead to epithelial–mesenchymal transition, which is mediated by defective E-cadherin and NMMIIA expression. - Highlights: • Knockdown of Kif5b in MDCK cells resulted in reduced cell proliferation rate. • Kif5b deficient MDCK cells underwent epithelial–mesenchymal transition. • E-cadherin and NMMIIA could interact with Kif5b in polarized MDCK cells. • Decreased E-cadherin and NMMIIA levels mediate EMT in Kif5b deficient MDCK cells. • Overexpression of E-cadherin and NMMIIA reverse the effects of Kif5b knockdown.

MLL/WDR5 Complex Regulates Kif2A Localization to Ensure Chromosome Congression and Proper Spindle Assembly during Mitosis.

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Ali, Aamir; Veeranki, Sailaja Naga; Chinchole, Akash; Tyagi, Shweta

2017-06-19

Mixed-lineage leukemia (MLL), along with multisubunit (WDR5, RbBP5, ASH2L, and DPY30) complex catalyzes the trimethylation of H3K4, leading to gene activation. Here, we characterize a chromatin-independent role for MLL during mitosis. MLL and WDR5 localize to the mitotic spindle apparatus, and loss of function of MLL complex by RNAi results in defects in chromosome congression and compromised spindle formation. We report interaction of MLL complex with several kinesin and dynein motors. We further show that the MLL complex associates with Kif2A, a member of the Kinesin-13 family of microtubule depolymerase, and regulates the spindle localization of Kif2A during mitosis. We have identified a conserved WDR5 interaction (Win) motif, so far unique to the MLL family, in Kif2A. The Win motif of Kif2A engages in direct interactions with WDR5 for its spindle localization. Our findings highlight a non-canonical mitotic function of MLL complex, which may have a direct impact on chromosomal stability, frequently compromised in cancer. Copyright © 2017 Elsevier Inc. All rights reserved.

Diabetes alters KIF1A and KIF5B motor proteins in the hippocampus.

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Filipa I Baptista

Full Text Available Diabetes mellitus is the most common metabolic disorder in humans. Diabetic encephalopathy is characterized by cognitive and memory impairments, which have been associated with changes in the hippocampus, but the mechanisms underlying those impairments triggered by diabetes, are far from being elucidated. The disruption of axonal transport is associated with several neurodegenerative diseases and might also play a role in diabetes-associated disorders affecting nervous system. We investigated the effect of diabetes (2 and 8 weeks duration on KIF1A, KIF5B and dynein motor proteins, which are important for axonal transport, in the hippocampus. The mRNA expression of motor proteins was assessed by qRT-PCR, and also their protein levels by immunohistochemistry in hippocampal slices and immunoblotting in total extracts of hippocampus from streptozotocin-induced diabetic and age-matched control animals. Diabetes increased the expression and immunoreactivity of KIF1A and KIF5B in the hippocampus, but no alterations in dynein were detected. Since hyperglycemia is considered a major player in diabetic complications, the effect of a prolonged exposure to high glucose on motor proteins, mitochondria and synaptic proteins in hippocampal neurons was also studied, giving particular attention to changes in axons. Hippocampal cell cultures were exposed to high glucose (50 mM or mannitol (osmotic control; 25 mM plus 25 mM glucose for 7 days. In hippocampal cultures incubated with high glucose no changes were detected in the fluorescence intensity or number of accumulations related with mitochondria in the axons of hippocampal neurons. Nevertheless, high glucose increased the number of fluorescent accumulations of KIF1A and synaptotagmin-1 and decreased KIF5B, SNAP-25 and synaptophysin immunoreactivity specifically in axons of hippocampal neurons. These changes suggest that anterograde axonal transport mediated by these kinesins may be impaired in hippocampal

Characterization of KIF11 as a novel prognostic biomarker and therapeutic target for oral cancer.

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Daigo, Kayo; Takano, Atsushi; Thang, Phung Manh; Yoshitake, Yoshihiro; Shinohara, Masanori; Tohnai, Iwau; Murakami, Yoshinori; Maegawa, Jiro; Daigo, Yataro

2018-01-01

Oral cancer has a high mortality rate, and its incidence is increasing gradually worldwide. As the effectiveness of standard treatments is still limited, the development of new therapeutic strategies is eagerly awaited. Kinesin family member 11 (KIF11) is a motor protein required for establishing a bipolar spindle in cell division. The role of KIF11 in oral cancer is unclear. Therefore, the present study aimed to assess the role of KIF11 in oral cancer and evaluate its role as a prognostic biomarker and therapeutic target for treating oral cancer. Immunohistochemical analysis demonstrated that KIF11 was expressed in 64 of 99 (64.6%) oral cancer tissues but not in healthy oral epithelia. Strong KIF11 expression was significantly associated with poor prognosis among oral cancer patients (P=0.034), and multivariate analysis confirmed its independent prognostic value. In addition, inhibition of KIF11 expression by transfection of siRNAs into oral cancer cells or treatment of cells with a KIF11 inhibitor significantly suppressed cell proliferation, probably through G2/M arrest and subsequent induction of apoptosis. These results suggest that KIF11 could be a potential prognostic biomarker and therapeutic target for oral cancer.

Molecular characterization of a KIF3B-like kinesin gene in the testis of Octopus tankahkeei (Cephalopoda, Octopus).

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Dang, Ran; Zhu, Jun-Quan; Tan, Fu-Qing; Wang, Wei; Zhou, Hong; Yang, Wan-Xi

2012-05-01

KIF3B is known for maintaining and assembling cilia and flagellum. To date, the function of KIF3B and its relationship with KIF3A during spermiogenesis in the cephalopod Octopus tankahkeei remains unknown. In the present study, we characterized a gene encoding a homologue of rat KIF3B in the O. tankahkeei testis and examined its temporal and spatial expression pattern during spermiogenesis. The cDNA of KIF3B was obtained with degenerate and RACE PCR and the distribution pattern of ot-kif3b were observed with RT-PCR. The morphological development during spermiogenesis was illustrated by histological and transmission electron microscopy and mRNA expression of ot-kif3b was observed by in situ hybridization. The 2,365 nucleotides cDNA consisted of a 102 bp 5' untranslated region (UTR), a 2,208 bp open reading frame (ORF) encoding a protein of 736 amino acids, and a 55 bp 3' UTR. Multiple alignments revealed that the putative Ot-KIF3B shared 68, 68, 69, 68, and 67% identity with that of Homo sapiens, Mus musculus, Gallus gallus, Danio rerio, and Xenopus laevis, respectively, along with high identities with Ot-KIF3A in fundamental structures. Ot-kif3b transcripts appeared gradually in early spermatids, increased in intermediate spermatids and maximized in drastically remodeled and final spermatids. The kif3b gene is identified and its expression pattern is demonstrated for the first time in O. tankahkeei. Compared to ot-kif3a reported by our laboratory before, our data suggested that the putative heterodimeric motor proteins Ot-KIF3A/B may be involved in intraspermatic transport and might contribute to structural changes during spermiogenesis.

A novel role of KIF3b in the seminoma cell cycle

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Shen, Hao-Qing; Xiao, Yu-Xi; She, Zhen-Yu [The Sperm Laboratory, College of Life Sciences, Zhejiang University, 866 Yu Hang Tang Road, Hangzhou 310058 (China); Tan, Fu-Qing [The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou 310003 (China); Yang, Wan-Xi, E-mail: wxyang@spermlab.org [The Sperm Laboratory, College of Life Sciences, Zhejiang University, 866 Yu Hang Tang Road, Hangzhou 310058 (China)

2017-03-01

KIF3b is a protein of the kinesin-2 family which plays an important role in intraflagellar transport. Testis cancer is a common cancer among young men. Its diagnostic rate is increasing and over half of the cases are seminomas. Many aspects of the mechanism and gene expression background of this cancer remain unclear. Using western-blotting and semi-quantitative PCR we found high protein levels of KIF3b enrichment in seminoma tissue despite the mRNA levels remaining equivalent to that of normal testicular tissues. The distribution of KIF3b was mainly in cells with division potential. Wound-healing assays and cell counting kit assays showed that the knockdown of KIF3b significantly suppressed cell migration ability, viability and number in HeLa cells. Immunofluorescence images during the cell cycle revealed that KIF3b tended to gather at the spindles and was enriched at the central spindle. This indicated that KIF3b may also have direct impacts upon spindle formation and cytokinesis. By counting the numbers of nuclei, spindles and cells, we found that the rates of multipolar division and multi-nucleation were raised in KIF3b-knockdown cells. In this way we demonstrate that KIF3b functions importantly in mitosis and may be essential to seminoma cell division and proliferation as well as being necessary for normal cell division. - Highlights: • A significant upregulation of KIF3b is detected in seminoma. • Knockdown of KIF3b impacts on cell proliferation and migration. • KIF3b may have direct impacts upon spindle formation and cytokinesis.

KIF14 Promotes AKT Phosphorylation and Contributes to Chemoresistance in Triple-Negative Breast Cancer

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Stina M. Singel

2014-03-01

Full Text Available Despite evidence that kinesin family member 14 (KIF14 can serve as a prognostic biomarker in various solid tumors, how it contributes to tumorigenesis remains unclear. We observed that experimental decrease in KIF14 expression increases docetaxel chemosensitivity in estrogen receptor–negative/progesterone receptor–negative/human epidermal growth factor receptor 2-negative, “triple-negative” breast cancers (TNBC. To investigate the oncogenic role of KIF14, we used noncancerous human mammary epithelial cells and ectopically expressed KIF14 and found increased proliferative capacity, increased anchorage-independent grown in vitro, and increased resistance to docetaxel but not to doxorubicin, carboplatin, or gemcitabine. Seventeen benign breast biopsies of BRCA1 or BRCA2 mutation carriers showed increased KIF14 mRNA expression by fluorescence in situ hybridization compared to controls with no known mutations in BRCA1 or BRCA2, suggesting increased KIF14 expression as a biomarker of high-risk breast tissue. Evaluation of 34 cases of locally advanced TNBC showed that KIF14 expression significantly correlates with chemotherapy-resistant breast cancer. KIF14 knockdown also correlates with decreased AKT phosphorylation and activity. Live-cell imaging confirmed an insulin-induced temporal colocalization of KIF14 and AKT at the plasma membrane, suggesting a potential role of KIF14 in promoting activation of AKT. An experimental small-molecule inhibitor of KIF14 was then used to evaluate the potential anticancer benefits of downregulating KIF14 activity. Inhibition of KIF14 shows a chemosensitizing effect and correlates with decreasing activation of AKT. Together, these findings show an early and critical role for KIF14 in the tumorigenic potential of TNBC, and therapeutic targeting of KIF14 is feasible and effective for TNBC.

Identification of Orch3, a locus controlling dominant resistance to autoimmune orchitis, as kinesin family member 1C.

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Roxana del Rio

Full Text Available Experimental autoimmune orchitis (EAO, the principal model of non-infectious testicular inflammatory disease, can be induced in susceptible mouse strains by immunization with autologous testicular homogenate and appropriate adjuvants. As previously established, the genome of DBA/2J mice encodes genes that are capable of conferring dominant resistance to EAO, while the genome of BALB/cByJ mice does not and they are therefore susceptible to EAO. In a genome scan, we previously identified Orch3 as the major quantitative trait locus controlling dominant resistance to EAO and mapped it to chromosome 11. Here, by utilizing a forward genetic approach, we identified kinesin family member 1C (Kif1c as a positional candidate for Orch3 and, using a transgenic approach, demonstrated that Kif1c is Orch3. Mechanistically, we showed that the resistant Kif1c(D2 allele leads to a reduced antigen-specific T cell proliferative response as a consequence of decreased MHC class II expression by antigen presenting cells, and that the L(578 → P(578 and S(1027 → P(1027 polymorphisms distinguishing the BALB/cByJ and DBA/2J alleles, respectively, can play a role in transcriptional regulation. These findings may provide mechanistic insight into how polymorphism in other kinesins such as KIF21B and KIF5A influence susceptibility and resistance to human autoimmune diseases.

KIF26B, a novel oncogene, promotes proliferation and metastasis by activating the VEGF pathway in gastric cancer.

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Zhang, H; Ma, R-R; Wang, X-J; Su, Z-X; Chen, X; Shi, D-B; Guo, X-Y; Liu, H-T; Gao, P

2017-10-05

Tumor metastasis is the main reason of cancer-related death for gastric cancer (GC) patients and gene expression microarray data indicate that kinesin family member 26B (KIF26B) is one of the most upregulated genes in metastatic GC samples. Specifically, KIF26B expression was upregulated in a stepwise manner from non-tumorous gastric mucosa, primary GC tissues without metastasis, via primary GC tissues with metastasis, to secondary lymph node metastatic (LNM) foci. Increased expression of KIF26B was correlated with tumor size, positive LNM or distant metastases and poor prognosis. KIF26B, negatively regulated by miR-372, promoted GC cell proliferation and metastasis in vitro and in vivo. Mechanistic investigations confirmed that the main target of KIF26B was the vascular endothelial growth factor (VEGF) signaling pathway, particularly by inhibition or overexpression of VEGFA, PXN, FAK, PIK3CA, BCL2 and CREB1. Thus, KIF26B, a novel oncogene regulated by miR-372, promotes proliferation and metastasis through the VEGF pathway in GC.

A coordinated molecular 'fishing' mechanism in heterodimeric kinesin

International Nuclear Information System (INIS)

Hou, Ruizheng; Wang, Zhisong

2010-01-01

Kar3 is a kinesin motor that facilitates chromosome segregation during cell division. Unlike many members of the kinesin superfamily, Kar3 forms a heterodimer with non-motor protein Vik1 or Cik1 in vivo. The heterodimers show ATP-driven minus-end directed motility along a microtubule (MT) lattice, and also serve as depolymerase at the MT ends. The molecular mechanisms behind this dual functionality remain mysterious. Here, a molecular mechanical model for the Kar3/Vik1 heterodimer based on structural, kinetic and motility data reveals a long-range chemomechanical transmission mechanism that resembles a familiar fishing tactic. By this molecular 'fishing', ATP-binding to Kar3 dissociates catalytically inactive Vik1 off MT to facilitate minus-end sliding of the dimer on the MT lattice. When the dimer binds the frayed ends of MT, the fishing channels ATP hydrolysis energy into MT deploymerization by a mechanochemical effect. The molecular fishing thus provides a unified mechanistic ground for Kar3's dual functionality. The fishing-promoted depolymerization differs from the depolymerase mechanisms found in homodimeric kinesins. The fishing also enables intermolecular coordination with a chemomechanical coupling feature different from the paradigmatic pattern of homodimeric motors. This study rationalizes some puzzling experimental observation, and suggests new experiments for further elucidation of the fishing mechanism

KIF7 Controls the Proliferation of Cells of the Respiratory Airway through Distinct Microtubule Dependent Mechanisms.

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Garry L Coles

2015-10-01

Full Text Available The cell cycle must be tightly coordinated for proper control of embryonic development and for the long-term maintenance of organs such as the lung. There is emerging evidence that Kinesin family member 7 (Kif7 promotes Hedgehog (Hh signaling during embryonic development, and its misregulation contributes to diseases such as ciliopathies and cancer. Kif7 encodes a microtubule interacting protein that controls Hh signaling through regulation of microtubule dynamics within the primary cilium. However, whether Kif7 has a function in nonciliated cells remains largely unknown. The role Kif7 plays in basic cell biological processes like cell proliferation or cell cycle progression also remains to be elucidated. Here, we show that Kif7 is required for coordination of the cell cycle, and inactivation of this gene leads to increased cell proliferation in vivo and in vitro. Immunostaining and transmission electron microscopy experiments show that Kif7dda/dda mutant lungs are hyperproliferative and exhibit reduced alveolar epithelial cell differentiation. KIF7 depleted C3H10T1/2 fibroblasts and Kif7dda/dda mutant mouse embryonic fibroblasts have increased growth rates at high cellular densities, suggesting that Kif7 may function as a general regulator of cellular proliferation. We ascertained that in G1, Kif7 and microtubule dynamics regulate the expression and activity of several components of the cell cycle machinery known to control entry into S phase. Our data suggest that Kif7 may function to regulate the maintenance of the respiratory airway architecture by controlling cellular density, cell proliferation, and cycle exit through its role as a microtubule associated protein.

The regulation of MS-KIF18A expression and cross talk with estrogen receptor.

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Margalit Zusev

2009-07-01

Full Text Available This study provides a novel view on the interactions between the MS-KIF18A, a kinesin protein, and estrogen receptor alpha (ERalpha which were studied in vivo and in vitro. Additionally, the regulation of MS-KIF18A expression by estrogen was investigated at the gene and protein levels. An association between recombinant proteins; ERalpha and MS-KIF18A was demonstrated in vitro in a pull down assay. Such interactions were proven also for endogenous proteins in MBA-15 cells were detected prominently in the cytoplasm and are up-regulated by estrogen. Additionally, an association between these proteins and the transcription factor NF-kappaB was identified. MS-KIF18A mRNA expression was measured in vivo in relation to age and estrogen level in mice and rats models. A decrease in MS-KIF18A mRNA level was measured in old and in OVX-estrogen depleted rats as compared to young animals. The low MS-KIF18A mRNA expression in OVX rats was restored by estrogen treatment. We studied the regulation of MS-KIF18A transcription by estrogen using the luciferase reporter gene and chromatin immuno-precipitation (ChIP assays. The luciferase reporter gene assay demonstrated an increase in MS-KIF18A promoter activity in response to 10(-8 M estrogen and 10(-7M ICI-182,780. Complimentary, the ChIP assay quantified the binding of ERalpha and pcJun to the MS-KIF18A promoter that was enhanced in cells treated by estrogen and ICI-182,780. In addition, cells treated by estrogen expressed higher levels of MS-KIF18A mRNA and protein and the protein turnover in MBA-15 cells was accelerated. Presented data demonstrated that ERalpha is a defined cargo of MS-KIF18A and added novel insight on the role of estrogen in regulation of MS-KIF18A expression both in vivo and in vitro.

Genome-wide Analyses Identify KIF5A as a Novel ALS Gene.

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Nicolas, Aude; Kenna, Kevin P; Renton, Alan E; Ticozzi, Nicola; Faghri, Faraz; Chia, Ruth; Dominov, Janice A; Kenna, Brendan J; Nalls, Mike A; Keagle, Pamela; Rivera, Alberto M; van Rheenen, Wouter; Murphy, Natalie A; van Vugt, Joke J F A; Geiger, Joshua T; Van der Spek, Rick A; Pliner, Hannah A; Shankaracharya; Smith, Bradley N; Marangi, Giuseppe; Topp, Simon D; Abramzon, Yevgeniya; Gkazi, Athina Soragia; Eicher, John D; Kenna, Aoife; Mora, Gabriele; Calvo, Andrea; Mazzini, Letizia; Riva, Nilo; Mandrioli, Jessica; Caponnetto, Claudia; Battistini, Stefania; Volanti, Paolo; La Bella, Vincenzo; Conforti, Francesca L; Borghero, Giuseppe; Messina, Sonia; Simone, Isabella L; Trojsi, Francesca; Salvi, Fabrizio; Logullo, Francesco O; D'Alfonso, Sandra; Corrado, Lucia; Capasso, Margherita; Ferrucci, Luigi; Moreno, Cristiane de Araujo Martins; Kamalakaran, Sitharthan; Goldstein, David B; Gitler, Aaron D; Harris, Tim; Myers, Richard M; Phatnani, Hemali; Musunuri, Rajeeva Lochan; Evani, Uday Shankar; Abhyankar, Avinash; Zody, Michael C; Kaye, Julia; Finkbeiner, Steven; Wyman, Stacia K; LeNail, Alex; Lima, Leandro; Fraenkel, Ernest; Svendsen, Clive N; Thompson, Leslie M; Van Eyk, Jennifer E; Berry, James D; Miller, Timothy M; Kolb, Stephen J; Cudkowicz, Merit; Baxi, Emily; Benatar, Michael; Taylor, J Paul; Rampersaud, Evadnie; Wu, Gang; Wuu, Joanne; Lauria, Giuseppe; Verde, Federico; Fogh, Isabella; Tiloca, Cinzia; Comi, Giacomo P; Sorarù, Gianni; Cereda, Cristina; Corcia, Philippe; Laaksovirta, Hannu; Myllykangas, Liisa; Jansson, Lilja; Valori, Miko; Ealing, John; Hamdalla, Hisham; Rollinson, Sara; Pickering-Brown, Stuart; Orrell, Richard W; Sidle, Katie C; Malaspina, Andrea; Hardy, John; Singleton, Andrew B; Johnson, Janel O; Arepalli, Sampath; Sapp, Peter C; McKenna-Yasek, Diane; Polak, Meraida; Asress, Seneshaw; Al-Sarraj, Safa; King, Andrew; Troakes, Claire; Vance, Caroline; de Belleroche, Jacqueline; Baas, Frank; Ten Asbroek, Anneloor L M A; Muñoz-Blanco, José Luis; Hernandez, Dena G; Ding, Jinhui; Gibbs, J Raphael; Scholz, Sonja W; Floeter, Mary Kay; Campbell, Roy H; Landi, Francesco; Bowser, Robert; Pulst, Stefan M; Ravits, John M; MacGowan, Daniel J L; Kirby, Janine; Pioro, Erik P; Pamphlett, Roger; Broach, James; Gerhard, Glenn; Dunckley, Travis L; Brady, Christopher B; Kowall, Neil W; Troncoso, Juan C; Le Ber, Isabelle; Mouzat, Kevin; Lumbroso, Serge; Heiman-Patterson, Terry D; Kamel, Freya; Van Den Bosch, Ludo; Baloh, Robert H; Strom, Tim M; Meitinger, Thomas; Shatunov, Aleksey; Van Eijk, Kristel R; de Carvalho, Mamede; Kooyman, Maarten; Middelkoop, Bas; Moisse, Matthieu; McLaughlin, Russell L; Van Es, Michael A; Weber, Markus; Boylan, Kevin B; Van Blitterswijk, Marka; Rademakers, Rosa; Morrison, Karen E; Basak, A Nazli; Mora, Jesús S; Drory, Vivian E; Shaw, Pamela J; Turner, Martin R; Talbot, Kevin; Hardiman, Orla; Williams, Kelly L; Fifita, Jennifer A; Nicholson, Garth A; Blair, Ian P; Rouleau, Guy A; Esteban-Pérez, Jesús; García-Redondo, Alberto; Al-Chalabi, Ammar; Rogaeva, Ekaterina; Zinman, Lorne; Ostrow, Lyle W; Maragakis, Nicholas J; Rothstein, Jeffrey D; Simmons, Zachary; Cooper-Knock, Johnathan; Brice, Alexis; Goutman, Stephen A; Feldman, Eva L; Gibson, Summer B; Taroni, Franco; Ratti, Antonia; Gellera, Cinzia; Van Damme, Philip; Robberecht, Wim; Fratta, Pietro; Sabatelli, Mario; Lunetta, Christian; Ludolph, Albert C; Andersen, Peter M; Weishaupt, Jochen H; Camu, William; Trojanowski, John Q; Van Deerlin, Vivianna M; Brown, Robert H; van den Berg, Leonard H; Veldink, Jan H; Harms, Matthew B; Glass, Jonathan D; Stone, David J; Tienari, Pentti; Silani, Vincenzo; Chiò, Adriano; Shaw, Christopher E; Traynor, Bryan J; Landers, John E

2018-03-21

To identify novel genes associated with ALS, we undertook two lines of investigation. We carried out a genome-wide association study comparing 20,806 ALS cases and 59,804 controls. Independently, we performed a rare variant burden analysis comparing 1,138 index familial ALS cases and 19,494 controls. Through both approaches, we identified kinesin family member 5A (KIF5A) as a novel gene associated with ALS. Interestingly, mutations predominantly in the N-terminal motor domain of KIF5A are causative for two neurodegenerative diseases: hereditary spastic paraplegia (SPG10) and Charcot-Marie-Tooth type 2 (CMT2). In contrast, ALS-associated mutations are primarily located at the C-terminal cargo-binding tail domain and patients harboring loss-of-function mutations displayed an extended survival relative to typical ALS cases. Taken together, these results broaden the phenotype spectrum resulting from mutations in KIF5A and strengthen the role of cytoskeletal defects in the pathogenesis of ALS. Copyright © 2018 Elsevier Inc. All rights reserved.

The Drosophila KIF1A homolog unc-104 is important for site-specific active zone maturation

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Yao V. Zhang

2016-09-01

Full Text Available Abstract Mutations in the kinesin-3 family member KIF1A have been associated with hereditary spastic paraplegia, hereditary sensory and autonomic neuropathy type 2 and intellectual disability. Both autosomal recessive and autosomal dominant forms of inheritance have been reported. Loss of KIF1A or its homolog unc-104 causes early postnatal or embryonic lethality in mice and Drosophila, respectively. In this study we use a previously described hypomorphic allele of unc-104, unc-104bris, to investigate the impact of partial loss-of-function of kinesin-3 function on active zone formation at the Drosophila neuromuscular junction. unc-104bris mutants exhibit synaptic defects where a subset of synapses at the neuromuscular junction lack the key active zone organizer protein Bruchpilot. Modulating synaptic Bruchpilot levels by ectopic overexpression or RNAi-mediated knockdown suggests that the loss of active zone components such as Ca2+ channel and Liprin-α from these synapses is caused by impaired kinesin-3 transport rather than due to the absence of Bruchpilot at these synapses. In addition to defects in active zone maturation, unc-104bris mutants display impaired transport of dense core vesicles and synaptic vesicle associated proteins, among which Rab3 has been shown to regulate the distribution of Bruchpilot to active zones. Overexpression of Rab3 partially ameliorates synaptic phenotypes of unc-104bris neuromuscular junction, suggesting that lack of presynaptic Rab3 may contribute to defects in synapse maturation.

Hot-spot KIF5A mutations cause familial ALS

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Yilmaz, Rüstem; Müller, Kathrin; Grehl, Torsten; Petri, Susanne; Meyer, Thomas; Grosskreutz, Julian; Weydt, Patrick; Ruf, Wolfgang; Neuwirth, Christoph; Weber, Markus; Pinto, Susana; Claeys, Kristl G; Schrank, Berthold; Jordan, Berit; Knehr, Antje; Günther, Kornelia; Hübers, Annemarie; Zeller, Daniel; Kubisch, Christian; Jablonka, Sibylle; Klopstock, Thomas; de Carvalho, Mamede; Sperfeld, Anne; Borck, Guntram; Volk, Alexander E; Dorst, Johannes; Weis, Joachim; Otto, Markus; Schuster, Joachim; Del Tredici, Kelly; Braak, Heiko; Danzer, Karin M; Freischmidt, Axel; Meitinger, Thomas; Strom, Tim M; Ludolph, Albert C; Andersen, Peter M; Weishaupt, Jochen H; Weyen, Ute; Hermann, Andreas; Hagenacker, Tim; Koch, Jan Christoph; Lingor, Paul; Göricke, Bettina; Zierz, Stephan; Baum, Petra; Wolf, Joachim; Winkler, Andrea; Young, Peter; Bogdahn, Ulrich; Prudlo, Johannes; Kassubek., Jan

2018-01-01

Abstract Heterozygous missense mutations in the N-terminal motor or coiled-coil domains of the kinesin family member 5A (KIF5A) gene cause monogenic spastic paraplegia (HSP10) and Charcot-Marie-Tooth disease type 2 (CMT2). Moreover, heterozygous de novo frame-shift mutations in the C-terminal domain of KIF5A are associated with neonatal intractable myoclonus, a neurodevelopmental syndrome. These findings, together with the observation that many of the disease genes associated with amyotrophic lateral sclerosis disrupt cytoskeletal function and intracellular transport, led us to hypothesize that mutations in KIF5A are also a cause of amyotrophic lateral sclerosis. Using whole exome sequencing followed by rare variant analysis of 426 patients with familial amyotrophic lateral sclerosis and 6137 control subjects, we detected an enrichment of KIF5A splice-site mutations in amyotrophic lateral sclerosis (2/426 compared to 0/6137 in controls; P = 4.2 × 10−3), both located in a hot-spot in the C-terminus of the protein and predicted to affect splicing exon 27. We additionally show co-segregation with amyotrophic lateral sclerosis of two canonical splice-site mutations in two families. Investigation of lymphoblast cell lines from patients with KIF5A splice-site mutations revealed the loss of mutant RNA expression and suggested haploinsufficiency as the most probable underlying molecular mechanism. Furthermore, mRNA sequencing of a rare non-synonymous missense mutation (predicting p.Arg1007Gly) located in the C-terminus of the protein shortly upstream of the splice donor of exon 27 revealed defective KIF5A pre-mRNA splicing in respective patient-derived cell lines owing to abrogation of the donor site. Finally, the non-synonymous single nucleotide variant rs113247976 (minor allele frequency = 1.00% in controls, n = 6137), also located in the C-terminal region [p.(Pro986Leu) in exon 26], was significantly enriched in familial amyotrophic lateral sclerosis patients (minor

KIF11 inhibition for glioblastoma treatment: reason to hope or a struggle with the brain?

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Valensin, Silvia; Ghiron, Chiara; Lamanna, Claudia; Kremer, Andreas; Rossi, Marco; Ferruzzi, Pietro; Nievo, Marco; Bakker, Annette

2009-01-01

Glioblastomas (GBM) are typically comprised of morphologically diverse cells. Despite current advances in therapy, including surgical resection followed by radiation and chemotherapy, the prognosis for patients with GBM remains poor. Unfortunately, most patients die within 2 years of diagnosis of their disease. Molecular abnormalities vary among individual patients and also within each tumor. Indeed, one of the distinguishing features of GBM is its marked genetic heterogeneity. Due to the brain location of the tumor, the potential target inhibition for anticancer therapy must exhibit a manageable neurotoxicity profile in the concentration range in which the compounds show anti-proliferative activity. Kinesin KIF11 inhibition by small molecules such as Monastrol or Ispinesib is currently under investigation in the field of malignant tumors. In the current study we have assessed the relevance of the anti-mitotic Kinesin-like protein KIF11 in human GBM cell-lines. In this study the target was validated using a set of well characterised and potentially specific small molecule inhibitors of KIF11: an ispinesib analog, Monastrol, a Merck compound and 3 simplified derivatives of the Merck compound. Following an in silico selection, those compounds predicted to bear a favorable BBB permeation profile were assessed for their phenotypic effect on cell lines derived both from primary (U87MG) as well as treated (DBTRG-05-MG) glioblastomas. For some compounds, these data could be compared to their effect on normal human astrocytes, as well as their neurotoxicity on primary rat cortical neurons. The ispinesib analogue 1 showed an anti-proliferative effect on GBM cell lines by blocking them in the G2/M phase in a concentration range which was shown to be harmless to primary rat cortical neurons. Furthermore, ispinesib analog increased caspase 3/7-induced apoptosis in U87MG cells. In the area of cell cycle inhibition, KIF11 is critical for proper spindle assembly and represents an

Hot-spot KIF5A mutations cause familial ALS.

Science.gov (United States)

Brenner, David; Yilmaz, Rüstem; Müller, Kathrin; Grehl, Torsten; Petri, Susanne; Meyer, Thomas; Grosskreutz, Julian; Weydt, Patrick; Ruf, Wolfgang; Neuwirth, Christoph; Weber, Markus; Pinto, Susana; Claeys, Kristl G; Schrank, Berthold; Jordan, Berit; Knehr, Antje; Günther, Kornelia; Hübers, Annemarie; Zeller, Daniel; Kubisch, Christian; Jablonka, Sibylle; Sendtner, Michael; Klopstock, Thomas; de Carvalho, Mamede; Sperfeld, Anne; Borck, Guntram; Volk, Alexander E; Dorst, Johannes; Weis, Joachim; Otto, Markus; Schuster, Joachim; Del Tredici, Kelly; Braak, Heiko; Danzer, Karin M; Freischmidt, Axel; Meitinger, Thomas; Strom, Tim M; Ludolph, Albert C; Andersen, Peter M; Weishaupt, Jochen H

2018-01-12

Heterozygous missense mutations in the N-terminal motor or coiled-coil domains of the kinesin family member 5A (KIF5A) gene cause monogenic spastic paraplegia (HSP10) and Charcot-Marie-Tooth disease type 2 (CMT2). Moreover, heterozygous de novo frame-shift mutations in the C-terminal domain of KIF5A are associated with neonatal intractable myoclonus, a neurodevelopmental syndrome. These findings, together with the observation that many of the disease genes associated with amyotrophic lateral sclerosis disrupt cytoskeletal function and intracellular transport, led us to hypothesize that mutations in KIF5A are also a cause of amyotrophic lateral sclerosis. Using whole exome sequencing followed by rare variant analysis of 426 patients with familial amyotrophic lateral sclerosis and 6137 control subjects, we detected an enrichment of KIF5A splice-site mutations in amyotrophic lateral sclerosis (2/426 compared to 0/6137 in controls; P = 4.2 × 10-3), both located in a hot-spot in the C-terminus of the protein and predicted to affect splicing exon 27. We additionally show co-segregation with amyotrophic lateral sclerosis of two canonical splice-site mutations in two families. Investigation of lymphoblast cell lines from patients with KIF5A splice-site mutations revealed the loss of mutant RNA expression and suggested haploinsufficiency as the most probable underlying molecular mechanism. Furthermore, mRNA sequencing of a rare non-synonymous missense mutation (predicting p.Arg1007Gly) located in the C-terminus of the protein shortly upstream of the splice donor of exon 27 revealed defective KIF5A pre-mRNA splicing in respective patient-derived cell lines owing to abrogation of the donor site. Finally, the non-synonymous single nucleotide variant rs113247976 (minor allele frequency = 1.00% in controls, n = 6137), also located in the C-terminal region [p.(Pro986Leu) in exon 26], was significantly enriched in familial amyotrophic lateral sclerosis patients (minor allele

KIF20A-Mediated RNA Granule Transport System Promotes the Invasiveness of Pancreatic Cancer Cells

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Keisuke Taniuchi

2014-12-01

Full Text Available Pancreatic cancers are aggressive because they are highly invasive and highly metastatic; moreover, effective treatments for aggressive pancreatic cancers are lacking. Here, we report that the motor kinesin protein KIF20A promoted the motility and invasiveness of pancreatic cancer cells through transporting the RNA-binding protein IGF2BP3 and IGF2BP3-bound transcripts toward cell protrusions along microtubules. We previously reported that IGF2BP3 and its target transcripts are assembled into cytoplasmic stress granules of pancreatic cancer cells, and that IGF2BP3 promotes the motility and invasiveness of pancreatic cancer cells through regulation of localized translation of IGF2BP3-bound transcripts in cell protrusions. We show that knockdown of KIF20A inhibited accumulation of IGF2BP3-containing stress granules in cell protrusions and suppressed local protein expression from specific IGF2BP3-bound transcripts, ARF6 and ARHGEF4, in the protrusions. Our results provide insight into the link between regulation of KIF20A-mediated trafficking of IGF2BP3-containing stress granules and modulation of the motility and invasiveness in pancreatic cancers.

HBV-related hepatocellular carcinoma susceptibility gene KIF1B is not associated with development of chronic hepatitis B.

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Rong Zhong

Full Text Available A recent genome-wide association study has identified a new susceptibility locus, kinesin family member 1B gene (KIF1B, strongly associated with progression from chronic hepatitis B (CHB to hepatitis B virus-related hepatocellular carcinoma (HCC in Chinese population, this study was carried out to explore the role of the genetic variants in KIF1B in the development of chronic hepatitis B.Three KIF1B polymorphisms (rs8019, rs17401924, and rs17401966 were selected and genotyped in 473 CHB patients and 580 controls with no history of CHB. Odds ratios (ORs and 95% confidence intervals (CIs were calculated by logistic regression model. None of these three SNPs showed association with CHBs after adjusting for age and gender. Equivalence-based method analysis confirmed the absence of association. In the further haplotype analysis, three common haplotypes were observed in this study population, but no significant effect was also found for haplotypes in the progression to CHB.This study showed the new locus identified for HCC, KIF1B, was not associated with progression to CHB, implying distinct genetic susceptibility factor contributes to the progression from hepatitis B virus infection to HCC. Nevertheless, further comprehensive analyses are warranted to dissect the mechanism.

Kinesin-73 is a processive motor that localizes to Rab5-containing organelles.

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Huckaba, Thomas M; Gennerich, Arne; Wilhelm, James E; Chishti, Athar H; Vale, Ronald D

2011-03-04

Drosophila Kinesin-73 (Khc-73), which plays a role in mitotic spindle polarity in neuroblasts, is a metazoan-specific member of the Kinesin-3 family of motors, which includes mammalian KIF1A and Caenorhabditis elegans Unc-104. The mechanism of Kinesin-3 motors has been controversial because some studies have reported that they transport cargo as monomers whereas other studies have suggested a dimer mechanism. Here, we have performed single-molecule motility and cell biological studies of Khc-73. We find that constructs containing the motor and the conserved short stretches of putative coiled-coil-forming regions are predominantly monomeric in vitro, but that dimerization allows for fast, processive movement and high force production (7 piconewtons). In Drosophila cell lines, we present evidence that Khc-73 can dimerize in vivo. We also show that Khc-73 is recruited specifically to Rab5-containing endosomes through its "tail" domain. Our results suggest that the N-terminal half of Khc-73 can undergo a monomer-dimer transition to produce a fast processive motor and that its C-terminal half possesses a specific Rab5-vesicle binding domain.

Identification of KIF3A as a Novel Candidate Gene for Childhood Asthma Using RNA Expression and Population Allelic Frequencies Differences

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Butsch Kovacic, Melinda; Biagini Myers, Jocelyn M.; Wang, Ning; Martin, Lisa J.; Lindsey, Mark; Ericksen, Mark B.; He, Hua; Patterson, Tia L.; Baye, Tesfaye M.; Torgerson, Dara; Roth, Lindsey A.; Gupta, Jayanta; Sivaprasad, Umasundari; Gibson, Aaron M.; Tsoras, Anna M.; Hu, Donglei; Eng, Celeste; Chapela, Rocío; Rodríguez-Santana, José R.; Rodríguez-Cintrón, William; Avila, Pedro C.; Beckman, Kenneth; Seibold, Max A.; Gignoux, Chris; Musaad, Salma M.; Chen, Weiguo; Burchard, Esteban González; Khurana Hershey, Gurjit K.

2011-01-01

Background Asthma is a chronic inflammatory disease with a strong genetic predisposition. A major challenge for candidate gene association studies in asthma is the selection of biologically relevant genes. Methodology/Principal Findings Using epithelial RNA expression arrays, HapMap allele frequency variation, and the literature, we identified six possible candidate susceptibility genes for childhood asthma including ADCY2, DNAH5, KIF3A, PDE4B, PLAU, SPRR2B. To evaluate these genes, we compared the genotypes of 194 predominantly tagging SNPs in 790 asthmatic, allergic and non-allergic children. We found that SNPs in all six genes were nominally associated with asthma (pasthma (OR = 2.3, pasthma population attributable risk of 18.5%. The association between KIF3A rs7737031 and asthma was validated in 3 independent populations, further substantiating the validity of our gene selection approach. Conclusions/Significance Our study demonstrates that KIF3A, a member of the kinesin superfamily of microtubule associated motors that are important in the transport of protein complexes within cilia, is a novel candidate gene for childhood asthma. Polymorphisms in KIF3A may in part be responsible for poor mucus and/or allergen clearance from the airways. Furthermore, our study provides a promising framework for the identification and evaluation of novel candidate susceptibility genes. PMID:21912604

Structural analysis of intermolecular interactions in the kinesin adaptor complex fasciculation and elongation protein zeta 1/ short coiled-coil protein (FEZ1/SCOCO.

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Marcos Rodrigo Alborghetti

Full Text Available Cytoskeleton and protein trafficking processes, including vesicle transport to synapses, are key processes in neuronal differentiation and axon outgrowth. The human protein FEZ1 (fasciculation and elongation protein zeta 1 / UNC-76, in C. elegans, SCOCO (short coiled-coil protein / UNC-69 and kinesins (e.g. kinesin heavy chain / UNC116 are involved in these processes. Exploiting the feature of FEZ1 protein as a bivalent adapter of transport mediated by kinesins and FEZ1 protein interaction with SCOCO (proteins involved in the same path of axonal growth, we investigated the structural aspects of intermolecular interactions involved in this complex formation by NMR (Nuclear Magnetic Resonance, cross-linking coupled with mass spectrometry (MS, SAXS (Small Angle X-ray Scattering and molecular modelling. The topology of homodimerization was accessed through NMR (Nuclear Magnetic Resonance studies of the region involved in this process, corresponding to FEZ1 (92-194. Through studies involving the protein in its monomeric configuration (reduced and dimeric state, we propose that homodimerization occurs with FEZ1 chains oriented in an anti-parallel topology. We demonstrate that the interaction interface of FEZ1 and SCOCO defined by MS and computational modelling is in accordance with that previously demonstrated for UNC-76 and UNC-69. SAXS and literature data support a heterotetrameric complex model. These data provide details about the interaction interfaces probably involved in the transport machinery assembly and open perspectives to understand and interfere in this assembly and its involvement in neuronal differentiation and axon outgrowth.

Effects of the KIF2C neck peptide on microtubules: lateral disintegration of microtubules and β-structure formation.

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Shimizu, Youské; Shimizu, Takashi; Nara, Masayuki; Kikumoto, Mahito; Kojima, Hiroaki; Morii, Hisayuki

2013-04-01

Members of the kinesin-13 sub-family, including KIF2C, depolymerize microtubules. The positive charge-rich 'neck' region extending from the N-terminus of the catalytic head is considered to be important in the depolymerization activity. Chemically synthesized peptides, covering the basic region (A182-E200), induced a sigmoidal increase in the turbidity of a microtubule suspension. The increase was suppressed by salt addition or by reduction of basicity by amino acid substitutions. Electron microscopic observations revealed ring structures surrounding the microtubules at high peptide concentrations. Using the peptide A182-D218, we also detected free thin straight filaments, probably protofilaments disintegrated from microtubules. Therefore, the neck region, even without the catalytic head domain, may induce lateral disintegration of microtubules. With microtubules lacking anion-rich C-termini as a result of subtilisin treatment, addition of the peptide induced only a moderate increase in turbidity, and rings and protofilaments were rarely detected, while aggregations, also thought to be caused by lateral disintegration, were often observed in electron micrographs. Thus, the C-termini are not crucial for the action of the peptides in lateral disintegration but contribute to structural stabilization of the protofilaments. Previous structural studies indicated that the neck region of KIF2C is flexible, but our IR analysis suggests that the cation-rich region (K190-A204) forms β-structure in the presence of microtubules, which may be of significance with regard to the action of the neck region. Therefore, the neck region of KIF2C is sufficient to cause disintegration of microtubules into protofilaments, and this may contribute to the ability of KIF2C to cause depolymerization of microtubules. © 2013 The Authors Journal compilation © 2013 FEBS.

Polymorphic locus rs10492972 of the KIF1B gene association with multiple sclerosis in Russia: case control study.

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Kudryavtseva, Ekaterina A; Rozhdestvenskii, Aleksei S; Kakulya, Anastasia V; Khanokh, Elena V; Delov, Roman A; Malkova, Nadezhda A; Korobko, Denis S; Platonov, Fedor A; Aref Eva, Elena G; Zagorskaya, Natalia N; Aliferova, Valentina M; Titova, Marina A; Babenko, Sergei A; Smagina, Inna V; El Chaninova, Svetlana A; Zolovkina, Anna G; Lifshits, G I; Puzyrev, Valerii P; Filipenko, Maxim L

2011-11-01

Axonal degeneration is responsible for the progression of the irreversible destruction caused by multiple sclerosis (MS) resulting ultimately in permanent disability. The KIF1B protein, a member of the kinesin family, is necessary for axon growth and myelination in vertebrates. In the recent paper, Aulchenko et al. suggested that the rs10492972[C] variant of KIF1B increases susceptibility to MS, but three following replication study didn't confirm this association. We studied the association of the polymorphic locus rs10492972 present in the KIF1B gene with genetic predisposition and its occurrence in clinical presentations of MS patients resident in western Siberia and the Sakha Republic (Yakutia), Russia. rs10492972 has been genotype in 833 samples of MS patient and 689 healthy controls. Distribution of rs10492972 genotypes corresponded with a Hardy-Weinberg distribution in both the MS patient and control groups, with the frequency of the C allele being the same in both groups (33%). Frequencies of occurrence of the genotypes were not shown to be associated with different disease courses or other characteristics of the disease, such as age at onset or duration. A complete meta-analysis of all analogous studies published to date showed that the protective effect of the rs10492972[C] allele is statistically significant (OR=0.95, C.I.95% [0.90-0.99], p=0.02). Copyright © 2011. Published by Elsevier Inc.

No evidence for cardiac dysfunction in Kif6 mutant mice.

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Abdul Hameed

Full Text Available A KIF6 variant in man has been reported to be associated with adverse cardiovascular outcomes after myocardial infarction. No clear biological or physiological data exist for Kif6. We sought to investigate the impact of a deleterious KIF6 mutation on cardiac function in mice. Kif6 mutant mice were generated and verified. Cardiac function was assessed by serial echocardiography at baseline, after ageing and after exercise. Lipid levels were also measured. No discernable adverse lipid or cardiac phenotype was detected in Kif6 mutant mice. These data suggest that dysfunction of Kif6 is linked to other more complex biological/biochemical parameters or is unlikely to be of material consequence in cardiac function.

Tau excess impairs mitosis and kinesin-5 function, leading to aneuploidy and cell death.

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Bougé, Anne-Laure; Parmentier, Marie-Laure

2016-03-01

In neurodegenerative diseases such as Alzheimer's disease (AD), cell cycle defects and associated aneuploidy have been described. However, the importance of these defects in the physiopathology of AD and the underlying mechanistic processes are largely unknown, in particular with respect to the microtubule (MT)-binding protein Tau, which is found in excess in the brain and cerebrospinal fluid of affected individuals. Although it has long been known that Tau is phosphorylated during mitosis to generate a lower affinity for MTs, there is, to our knowledge, no indication that an excess of this protein could affect mitosis. Here, we studied the effect of an excess of human Tau (hTau) protein on cell mitosis in vivo. Using the Drosophila developing wing disc epithelium as a model, we show that an excess of hTau induces a mitotic arrest, with the presence of monopolar spindles. This mitotic defect leads to aneuploidy and apoptotic cell death. We studied the mechanism of action of hTau and found that the MT-binding domain of hTau is responsible for these defects. We also demonstrate that the effects of hTau occur via the inhibition of the function of the kinesin Klp61F, the Drosophila homologue of kinesin-5 (also called Eg5 or KIF11). We finally show that this deleterious effect of hTau is also found in other Drosophila cell types (neuroblasts) and tissues (the developing eye disc), as well as in human HeLa cells. By demonstrating that MT-bound Tau inhibits the Eg5 kinesin and cell mitosis, our work provides a new framework to consider the role of Tau in neurodegenerative diseases. © 2016. Published by The Company of Biologists Ltd.

Glycogen synthase kinase 3 phosphorylates kinesin light chains and negatively regulates kinesin-based motility

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Morfini, Gerardo; Szebenyi, Gyorgyi; Elluru, Ravindhra; Ratner, Nancy; Brady, Scott T.

2002-01-01

Membrane-bounded organelles (MBOs) are delivered to different domains in neurons by fast axonal transport. The importance of kinesin for fast antero grade transport is well established, but mechanisms for regulating kinesin-based motility are largely unknown. In this report, we provide biochemical and in vivo evidence that kinesin light chains (KLCs) interact with and are in vivo substrates for glycogen synthase kinase 3 (GSK3). Active GSK3 inhibited anterograde, but not retrograde, transport in squid axoplasm and reduced the amount of kinesin bound to MBOs. Kinesin microtubule binding and microtubule-stimulated ATPase activities were unaffected by GSK3 phosphorylation of KLCs. Active GSK3 was also localized preferentially to regions known to be sites of membrane delivery. These data suggest that GSK3 can regulate fast anterograde axonal transport and targeting of cargos to specific subcellular domains in neurons.

KIF1Bβ and Neuroblastoma: Failure to Divide and Cull

OpenAIRE

Blackstone, Craig

2016-01-01

Neuroblastomas are associated with KIF1Bβ mutations within tumor suppressor region 1p36. In this issue of Developmental Cell, Li et al. (2016) show that KIF1Bβ binding releases calcineurin autoinhibition, leading to dephosphorylation of the DRP1 GTPase and subsequent mitochondrial fragmentation. KIF1Bβ impairment causes mitochondrial hyperfusion, impairing developmental apoptosis and promoting tumorigenesis.

Kinesin expands and stabilizes the GDP-microtubule lattice

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Peet, Daniel R.; Burroughs, Nigel J.; Cross, Robert A.

2018-05-01

Kinesin-1 is a nanoscale molecular motor that walks towards the fast-growing (plus) ends of microtubules, hauling molecular cargo to specific reaction sites in cells. Kinesin-driven transport is central to the self-organization of eukaryotic cells and shows great promise as a tool for nano-engineering1. Recent work hints that kinesin may also play a role in modulating the stability of its microtubule track, both in vitro2,3 and in vivo4, but the results are conflicting5-7 and the mechanisms are unclear. Here, we report a new dimension to the kinesin-microtubule interaction, whereby strong-binding state (adenosine triphosphate (ATP)-bound and apo) kinesin-1 motor domains inhibit the shrinkage of guanosine diphosphate (GDP) microtubules by up to two orders of magnitude and expand their lattice spacing by 1.6%. Our data reveal an unexpected mechanism by which the mechanochemical cycles of kinesin and tubulin interlock, and so allow motile kinesins to influence the structure, stability and mechanics of their microtubule track.

Casein Kinase 2 Reverses Tail-Independent Inactivation of Kinesin-1

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Xu, Jing

2013-03-01

Kinesin-1 is a plus-end microtubule-based motor, and defects in kinesin-based transport are linked to diseases including neurodegeneration. Kinesin can auto-inhibit via a head-tail interaction, but is believed to be active otherwise. Here we report a tail-independent inactivation of kinesin, reversible by the disease-relevant signalling protein, casein kinase 2 (CK2). The majority of initially active kinesin (native or tail-less) loses its ability to interact with microtubules in vitro, and CK2 reverses this inactivation (approximately fourfold) without altering kinesin's single motor properties. This activation pathway does not require motor phosphorylation, and is independent of head-tail auto-inhibition. In cultured mammalian cells, reducing CK2 expression, but not its kinase activity, decreases the force required to stall lipid droplet transport, consistent with a decreased number of active kinesin motors. Our results (Nat. Commun., 3:754, 2012) provide the first direct evidence of a protein kinase upregulating kinesin-based transport, and suggest a novel pathway for regulating the activity of cargo-bound kinesin. Work supported by NIGMS grants GM64624 to SPG, GM74830-06A1 to LH, GM76516 to LB, NS048501 to SJK, and AHA grant 825278F to JX.

The kinesin-3 family motor KLP-4 regulates anterograde trafficking of GLR-1 glutamate receptors in the ventral nerve cord of Caenorhabditis elegans.

Science.gov (United States)

Monteiro, Michael I; Ahlawat, Shikha; Kowalski, Jennifer R; Malkin, Emily; Koushika, Sandhya P; Juo, Peter

2012-09-01

The transport of glutamate receptors from the cell body to synapses is essential during neuronal development and may contribute to the regulation of synaptic strength in the mature nervous system. We previously showed that cyclin-dependent kinase-5 (CDK-5) positively regulates the abundance of GLR-1 glutamate receptors at synapses in the ventral nerve cord (VNC) of Caenorhabditis elegans. Here we identify a kinesin-3 family motor klp-4/KIF13 in a cdk-5 suppressor screen for genes that regulate GLR-1 trafficking. klp-4 mutants have decreased abundance of GLR-1 in the VNC. Genetic analysis of klp-4 and the clathrin adaptin unc-11/AP180 suggests that klp-4 functions before endocytosis in the ventral cord. Time-lapse microscopy indicates that klp-4 mutants exhibit decreased anterograde flux of GLR-1. Genetic analysis of cdk-5 and klp-4 suggests that they function in the same pathway to regulate GLR-1 in the VNC. Interestingly, GLR-1 accumulates in cell bodies of cdk-5 but not klp-4 mutants. However, GLR-1 does accumulate in klp-4-mutant cell bodies if receptor degradation in the multivesicular body/lysosome pathway is blocked. This study identifies kinesin KLP-4 as a novel regulator of anterograde glutamate receptor trafficking and reveals a cellular control mechanism by which receptor cargo is targeted for degradation in the absence of its motor.

Identification of cytoskeleton-associated proteins essential for lysosomal stability and survival of human cancer cells

DEFF Research Database (Denmark)

Groth-Pedersen, Line; Aits, Sonja; Corcelle-Termeau, Elisabeth

2012-01-01

Microtubule-disturbing drugs inhibit lysosomal trafficking and induce lysosomal membrane permeabilization followed by cathepsin-dependent cell death. To identify specific trafficking-related proteins that control cell survival and lysosomal stability, we screened a molecular motor siRNA library...... in human MCF7 breast cancer cells. SiRNAs targeting four kinesins (KIF11/Eg5, KIF20A, KIF21A, KIF25), myosin 1G (MYO1G), myosin heavy chain 1 (MYH1) and tropomyosin 2 (TPM2) were identified as effective inducers of non-apoptotic cell death. The cell death induced by KIF11, KIF21A, KIF25, MYH1 or TPM2 si......), increased dextran accumulation (KIF20A), or reduced autophagic flux (MYO1G, MYH1). Importantly, all seven siRNAs also killed human cervix cancer (HeLa) and osteosarcoma (U-2-OS) cells and sensitized cancer cells to other lysosome-destabilizing treatments, i.e. photo-oxidation, siramesine, etoposide...

Motoring through: the role of kinesin superfamily proteins in female meiosis.

Science.gov (United States)

Camlin, Nicole J; McLaughlin, Eileen A; Holt, Janet E

2017-07-01

The kinesin motor protein family consists of 14 distinct subclasses and 45 kinesin proteins in humans. A large number of these proteins, or their orthologues, have been shown to possess essential function(s) in both the mitotic and the meiotic cell cycle. Kinesins have important roles in chromosome separation, microtubule dynamics, spindle formation, cytokinesis and cell cycle progression. This article contains a review of the literature with respect to the role of kinesin motor proteins in female meiosis in model species. Throughout, we discuss the function of each class of kinesin proteins during oocyte meiosis, and where such data are not available their role in mitosis is considered. Finally, the review highlights the potential clinical importance of this family of proteins for human oocyte quality. To examine the role of kinesin motor proteins in oocyte meiosis. A search was performed on the Pubmed database for journal articles published between January 1970 and February 2017. Search terms included 'oocyte kinesin' and 'meiosis kinesin' in addition to individual kinesin names with the terms oocyte or meiosis. Within human cells 45 kinesin motor proteins have been discovered, with the role of only 13 of these proteins, or their orthologues, investigated in female meiosis. Furthermore, of these kinesins only half have been examined in mammalian oocytes, despite alterations occurring in gene transcripts or protein expression with maternal ageing, cryopreservation or behavioral conditions, such as binge drinking, for many of them. Kinesin motor proteins have distinct and important roles throughout oocyte meiosis in many non-mammalian model species. However, the functions these proteins have in mammalian meiosis, particularly in humans, are less clear owing to lack of research. This review brings to light the need for more experimental investigation of kinesin motor proteins, particularly those associated with maternal ageing, cryopreservation or exposure to

Chlorpyrifos, chlorpyrifos-oxon, and diisopropylfluorophosphate inhibit kinesin-dependent microtubule motility

International Nuclear Information System (INIS)

Gearhart, Debra A.; Sickles, Dale W.; Buccafusco, Jerry J.; Prendergast, Mark A.; Terry, Alvin V.

2007-01-01

Diisopropylfluorophosphate, originally developed as a chemical warfare agent, is structurally similar to nerve agents, and chlorpyrifos has extensive worldwide use as an agricultural pesticide. While inhibition of cholinesterases underlies the acute toxicity of these organophosphates, we previously reported impaired axonal transport in the sciatic nerves from rats treated chronically with subthreshold doses of chlorpyrifos. Those data indicate that chlorpyrifos (and/or its active metabolite, chlorpyrifos-oxon) might directly affect the function of kinesin and/or microtubules-the principal proteins that mediate anterograde axonal transport. The current report describes in vitro assays to assess the concentration-dependent effects of chlorpyrifos (0-10 μM), chlorpyrifos-oxon (0-10 μM), and diisopropylfluorophosphate (0-0.59 nM) on kinesin-dependent microtubule motility. Preincubating bovine brain microtubules with the organophosphates did not alter kinesin-mediated microtubule motility. In contrast, preincubation of bovine brain kinesin with diisopropylfluorophosphate, chlorpyrifos, or chlorpyrifos-oxon produced a concentration-dependent increase in the number of locomoting microtubules that detached from the kinesin-coated glass cover slip. Our data suggest that the organophosphates-chlorpyrifos-oxon, chlorpyrifos, and diisopropylfluorophosphate-directly affect kinesin, thereby disrupting kinesin-dependent transport on microtubules. Kinesin-dependent movement of vesicles, organelles, and other cellular components along microtubules is fundamental to the organization of all eukaryotic cells, especially in neurons where organelles and proteins synthesized in the cell body must move down long axons to pre-synaptic sites in nerve terminals. We postulate that disruption of kinesin-dependent intracellular transport could account for some of the long-term effects of organophosphates on the peripheral and central nervous system

Small Molecule Screen for Candidate Antimalarials Targeting Plasmodium Kinesin-5*

Science.gov (United States)

Liu, Liqiong; Richard, Jessica; Kim, Sunyoung; Wojcik, Edward J.

2014-01-01

Plasmodium falciparum and vivax are responsible for the majority of malaria infections worldwide, resulting in over a million deaths annually. Malaria parasites now show measured resistance to all currently utilized drugs. Novel antimalarial drugs are urgently needed. The Plasmodium Kinesin-5 mechanoenzyme is a suitable “next generation” target. Discovered via small molecule screen experiments, the human Kinesin-5 has multiple allosteric sites that are “druggable.” One site in particular, unique in its sequence divergence across all homologs in the superfamily and even within the same family, exhibits exquisite drug specificity. We propose that Plasmodium Kinesin-5 shares this allosteric site and likewise can be targeted to uncover inhibitors with high specificity. To test this idea, we performed a screen for inhibitors selective for Plasmodium Kinesin-5 ATPase activity in parallel with human Kinesin-5. Our screen of nearly 2000 compounds successfully identified compounds that selectively inhibit both P. vivax and falciparum Kinesin-5 motor domains but, as anticipated, do not impact human Kinesin-5 activity. Of note is a candidate drug that did not biochemically compete with the ATP substrate for the conserved active site or disrupt the microtubule-binding site. Together, our experiments identified MMV666693 as a selective allosteric inhibitor of Plasmodium Kinesin-5; this is the first identified protein target for the Medicines of Malaria Venture validated collection of parasite proliferation inhibitors. This work demonstrates that chemical screens against human kinesins are adaptable to homologs in disease organisms and, as such, extendable to strategies to combat infectious disease. PMID:24737313

Changes in microtubule overlap length regulate kinesin-14-driven microtubule sliding

Czech Academy of Sciences Publication Activity Database

Braun, Marcus; Lánský, Zdeněk; Szuba, A.; Schwarz, F. W.; Mitra, A.; Gao, M.; Luedecke, A.; ten Wolde, P.R.; Diez, S.

2017-01-01

Roč. 13, č. 12 (2017), s. 1245-1252 ISSN 1552-4450 R&D Projects: GA ČR(CZ) GA15-17488S; GA ČR(CZ) GA17-12496Y; GA ČR(CZ) GJ17-12496Y; GA MŠk(CZ) ED1.1.00/02.0109 Institutional support: RVO:86652036 Keywords : SPINDLE ELONGATION * MITOTIC SPINDLE * KINESIN-5 CIN8 * CROSS-LINKERS Subject RIV: EB - Genetics ; Molecular Biology OBOR OECD: Biophysics Impact factor: 15.066, year: 2016

The homodimeric GBS1074 from Streptococcus agalactiae

International Nuclear Information System (INIS)

Shukla, Anshuman; Pallen, Mark; Anthony, Mark; White, Scott A.

2010-01-01

The homodimeric nature of the ESAT-6 homologue GBS1074 and the potential for fibre-like assemblies are revealed by the 2 Å resolution crystal structure. ESAT-6 is a well characterized secreted protein from Mycobacterium tuberculosis and represents the archetype of the WXG100 family of proteins. Genes encoding ESAT-6 homologues have been identified in the genome of the human pathogen Streptococcus agalactiae; one of these genes, esxA, has been cloned and the recombinant protein has been crystallized. In contrast to M. tuberculosis ESAT-6, the crystal structure of GBS1074 reveals a homodimeric structure similar to homologous structures from Staphylococcus aureus and Helicobacter pylori. Intriguingly, GBS1074 forms elongated fibre-like assemblies in the crystal structure

Highly loaded behavior of kinesins increases the robustness of transport under high resisting loads.

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Woochul Nam

2015-03-01

Full Text Available Kinesins are nano-sized biological motors which walk by repeating a mechanochemical cycle. A single kinesin molecule is able to transport its cargo about 1 μm in the absence of external loads. However, kinesins perform much longer range transport in cells by working collectively. This long range of transport by a team of kinesins is surprising because the motion of the cargo in cells can be hindered by other particles. To reveal how the kinesins are able to accomplish their tasks of transport in harsh intracellular circumstances, stochastic studies on the kinesin motion are performed by considering the binding and unbinding of kinesins to microtubules and their dependence on the force acting on kinesin molecules. The unbinding probabilities corresponding to each mechanochemical state of kinesin are modeled. The statistical characterization of the instants and locations of binding are captured by computing the probability of unbound kinesin being at given locations. It is predicted that a group of kinesins has a more efficient transport than a single kinesin from the perspective of velocity and run length. Particularly, when large loads are applied, the leading kinesin remains bound to the microtubule for long time which increases the chances of the other kinesins to bind to the microtubule. To predict effects of this behavior of the leading kinesin under large loads on the collective transport, the motion of the cargo is studied when the cargo confronts obstacles. The result suggests that the behavior of kinesins under large loads prevents the early termination of the transport which can be caused by the interference with the static or moving obstacles.

Localization of the kinesin adaptor proteins trafficking kinesin proteins 1 and 2 in primary cultures of hippocampal pyramidal and cortical neurons.

Science.gov (United States)

Loss, Omar; Stephenson, F Anne

2015-07-01

Neuronal function requires regulated anterograde and retrograde trafficking of mitochondria along microtubules by using the molecular motors kinesin and dynein. Previous work has established that trafficking kinesin proteins (TRAKs),TRAK1 and TRAK2, are kinesin adaptor proteins that link mitochondria to kinesin motor proteins via an acceptor protein in the mitochondrial outer membrane, etc. the Rho GTPase Miro. Recent studies have shown that TRAK1 preferentially controls mitochondrial transport in axons of hippocampal neurons by virtue of its binding to both kinesin and dynein motor proteins, whereas TRAK2 controls mitochondrial transport in dendrites resulting from its binding to dynein. This study further investigates the subcellular localization of TRAK1 and TRAK2 in primary cultures of hippocampal and cortical neurons by using both commercial antibodies and anti-TRAK1 and anti-TRAK2 antibodies raised in our own laboratory (in-house). Whereas TRAK1 was prevalently localized in axons of hippocampal and cortical neurons, TRAK2 was more prevalent in dendrites of hippocampal neurons. In cortical neurons, TRAK2 was equally distributed between axons and dendrites. Some qualitative differences were observed between commercial and in-house-generated antibody immunostaining. © 2015 Wiley Periodicals, Inc.

The Molecular Motor KIF1A Transports the TrkA Neurotrophin Receptor and Is Essential for Sensory Neuron Survival and Function.

Science.gov (United States)

Tanaka, Yosuke; Niwa, Shinsuke; Dong, Ming; Farkhondeh, Atena; Wang, Li; Zhou, Ruyun; Hirokawa, Nobutaka

2016-06-15

KIF1A is a major axonal transport motor protein, but its functional significance remains elusive. Here we show that KIF1A-haploinsufficient mice developed sensory neuropathy. We found progressive loss of TrkA(+) sensory neurons in Kif1a(+/-) dorsal root ganglia (DRGs). Moreover, axonal transport of TrkA was significantly disrupted in Kif1a(+/-) neurons. Live imaging and immunoprecipitation assays revealed that KIF1A bound to TrkA-containing vesicles through the adaptor GTP-Rab3, suggesting that TrkA is a cargo of the KIF1A motor. Physiological measurements revealed a weaker capsaicin response in Kif1a(+/-) DRG neurons. Moreover, these neurons were hyposensitive to nerve growth factor, which could explain the reduced neuronal survival and the functional deficiency of the pain receptor TRPV1. Because phosphatidylinositol 3-kinase (PI3K) signaling significantly rescued these phenotypes and also increased Kif1a mRNA, we propose that KIF1A is essential for the survival and function of sensory neurons because of the TrkA transport and its synergistic support of the NGF/TrkA/PI3K signaling pathway. Copyright © 2016 Elsevier Inc. All rights reserved.

Loading direction regulates the affinity of ADP for kinesin.

Science.gov (United States)

Uemura, Sotaro; Ishiwata, Shin'ichi

2003-04-01

Kinesin is an ATP-driven molecular motor that moves processively along a microtubule. Processivity has been explained as a mechanism that involves alternating single- and double-headed binding of kinesin to microtubules coupled to the ATPase cycle of the motor. The internal load imposed between the two bound heads has been proposed to be a key factor regulating the ATPase cycle in each head. Here we show that external load imposed along the direction of motility on a single kinesin molecule enhances the binding affinity of ADP for kinesin, whereas an external load imposed against the direction of motility decreases it. This coupling between loading direction and enzymatic activity is in accord with the idea that the internal load plays a key role in the unidirectional and cooperative movement of processive motors.

Microtubule Destabilizer KIF2A Undergoes Distinct Site-Specific Phosphorylation Cascades that Differentially Affect Neuronal Morphogenesis

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Tadayuki Ogawa

2015-09-01

Full Text Available Neurons exhibit dynamic structural changes in response to extracellular stimuli. Microtubules (MTs provide rapid and dramatic cytoskeletal changes within the structural framework. However, the molecular mechanisms and signaling networks underlying MT dynamics remain unknown. Here, we have applied a comprehensive and quantitative phospho-analysis of the MT destabilizer KIF2A to elucidate the regulatory mechanisms of MT dynamics within neurons in response to extracellular signals. Interestingly, we identified two different sets of KIF2A phosphorylation profiles that accelerate (A-type and brake (B-type the MT depolymerization activity of KIF2A. Brain-derived neurotrophic factor (BDNF stimulates PAK1 and CDK5 kinases, which decrease the MT depolymerizing activity of KIF2A through B-type phosphorylation, resulting in enhanced outgrowth of neural processes. In contrast, lysophosphatidic acid (LPA induces ROCK2 kinase, which suppresses neurite outgrowth from round cells via A-type phosphorylation. We propose that these two mutually exclusive forms of KIF2A phosphorylation differentially regulate neuronal morphogenesis during development.

Novel Insights Into the Phenotypical Spectrum of KIF11-Associated Retinopathy, Including a New Form of Retinal Ciliopathy.

Science.gov (United States)

Birtel, Johannes; Gliem, Martin; Mangold, Elisabeth; Tebbe, Lars; Spier, Isabel; Müller, Philipp L; Holz, Frank G; Neuhaus, Christine; Wolfrum, Uwe; Bolz, Hanno J; Charbel Issa, Peter

2017-08-01

This study sought to characterize the ophthalmic and extraocular phenotype in patients with known and novel KIF11 mutations. Four patients (3, 5, 36, and 38 years of age, on father-daughter constellation) from three unrelated families were characterized by retinal examination including multimodal retinal imaging, investigation for syndromic disease manifestations, and targeted next-generation sequencing. The subcellular localization of Kif11 in the retina was analyzed by light and electron microcopy. There was considerable interindividual and intrafamilial phenotypic heterogeneity of KIF11-related retinopathy. Two patients presented with a progressive retinal dystrophy, one with chorioretinal dysplasia and one with familial exudative vitreoretinopathy (FEVR) in one eye and thinning of the photoreceptor layer in the fellow eye. Obvious syndromic disease manifestations were present only in the youngest patient, but minor signs (e.g. reduced head circumference) were present in the three other individuals. Immunohistochemistry results demonstrated Kif11 localization in the inner segment and ciliary compartments of photoreceptor cells and in the retinal pigment epithelium. Progressive retinal degeneration in KIF11-related retinopathy indicates a role for KIF11 not only in ocular development but also in maintaining retinal morphology and function. The remarkable variability of the ocular phenotype suggests four different types of retinopathy which may overlap. KIF11 should be considered in the screening of patients with retinal dystrophies because other syndromic manifestations may be subtle. Evaluation of head circumference may be considered as a potential shortcut to the genetic diagnosis. The localization of Kif11 in photoreceptor cells indicates a retinal ciliopathy.

Fe-S cluster coordination of the chromokinesin KIF4A alters its sub-cellular localization during mitosis.

Science.gov (United States)

Ben-Shimon, Lilach; Paul, Viktoria D; David-Kadoch, Galit; Volpe, Marina; Stümpfig, Martin; Bill, Eckhard; Mühlenhoff, Ulrich; Lill, Roland; Ben-Aroya, Shay

2018-05-30

Fe-S clusters act as co-factors of proteins with diverse functions, e.g. in DNA repair. Down-regulation of the cytosolic iron-sulfur protein assembly (CIA) machinery promotes genomic instability by the inactivation of multiple DNA repair pathways. Furthermore, CIA deficiencies are associated with so far unexplained mitotic defects. Here, we show that CIA2B and MMS19, constituents of the CIA targeting complex involved in facilitating Fe-S cluster insertion into cytosolic and nuclear target proteins, co-localize with components of the mitotic machinery. Down-regulation of CIA2B and MMS19 impairs the mitotic cycle. We identify the chromokinesin KIF4A as a mitotic component involved in these effects. KIF4A binds a Fe-S cluster in vitro through its conserved cysteine-rich domain. We demonstrate in vivo that this domain is required for the mitosis-related KIF4A localization and for the mitotic defects associated with KIF4A knockout. KIF4A is the first identified mitotic component carrying such a post-translational modification. These findings suggest that the lack of Fe-S clusters in KIF4A upon down-regulation of the CIA targeting complex contributes to the mitotic defects. © 2018. Published by The Company of Biologists Ltd.

Potential involvement of kinesin-1 in the regulation of subcellular localization of Girdin

Energy Technology Data Exchange (ETDEWEB)

Muramatsu, Aya [Department of Pathology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550 (Japan); Enomoto, Atsushi, E-mail: enomoto@iar.nagoya-u.ac.jp [Department of Pathology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550 (Japan); Kato, Takuya; Weng, Liang [Department of Pathology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550 (Japan); Kuroda, Keisuke [Department of Cell Pharmacology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550 (Japan); Asai, Naoya; Asai, Masato; Mii, Shinji [Department of Pathology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550 (Japan); Takahashi, Masahide, E-mail: mtakaha@med.nagoya-u.ac.jp [Department of Pathology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550 (Japan)

2015-08-07

Girdin is an actin-binding protein that has multiple functions in postnatal neural development and cancer progression. We previously showed that Girdin is a regulator of migration for neuroblasts born from neural stem cells in the subventricular zone (SVZ) and the dentate gyrus of the hippocampus in the postnatal brain. Despite a growing list of Girdin-interacting proteins, the mechanism of Girdin-mediated migration has not been fully elucidated. Girdin interacts with Disrupted-In-Schizophrenia 1 and partitioning-defective 3, both of which have been shown to interact with the kinesin microtubule motor proteins. Based on this, we have identified that Girdin also interacts with kinesin-1, a member of neuronal kinesin proteins. Although a direct interaction of Girdin and kinesin-1 has not been determined, it is of interest to find that Girdin loss-of-function mutant mice with the mutation of a basic amino acid residue-rich region (Basic mut mice) exhibit limited interaction with kinesin-1. Furthermore, expression of a kinesin-1 mutant with motor defects, leads to Girdin mislocalization. Finally, consistent with previous studies on the role of kinesin proteins in trafficking a cell–cell adhesion molecule N-cadherin, Basic mut mice showed an aberrant expression pattern of N-cadherin in migrating SVZ neuroblasts. These findings suggest a potential role of Girdin/kinesin-1 interaction in the regulation of neuroblast migration in the postnatal brain. - Highlights: • Girdin is a regulator of migration for neuroblasts in the postnatal brain. • Girdin interacts with kinesin-1, a member of neuronal kinesin proteins. • Girdin mutant mice showed an aberrant expression of N-cadherin in neuroblasts.

Heterogeneity in kinesin function

NARCIS (Netherlands)

Reddy, Babu J N; Tripathy, Suvranta; Vershinin, Michael; Tanenbaum, Marvin E; Xu, Jing; Mattson-Hoss, Michelle; Arabi, Karim; Chapman, Dail; Doolin, Tory; Hyeon, Changbong; Gross, Steven P

2017-01-01

The kinesin family proteins are often studied as prototypical molecular motors; a deeper understanding of them can illuminate regulation of intracellular transport. It is typically assumed that they function identically. Here we find that this assumption of homogeneous function appears incorrect:

Investigation of KIF6 Trp719Arg in a case-control study of myocardial infarction: a Costa Rican population.

Directory of Open Access Journals (Sweden)

Lance A Bare

2010-09-01

Full Text Available The 719Arg allele of KIF6 (rs20455 was associated with coronary events in Caucasian participants of five prospective studies. We investigated whether this KIF6 variant was associated with non-fatal myocardial infarction (MI in a case-control study of an admixed population from the Central Valley of Costa Rica. Genotypes of the KIF6 variant were determined for 4,134 men and women. Cases (1,987 had survived a first MI; controls (2,147 had no history of MI and were matched to cases by age, sex, and area of residence. We tested the association between the KIF6 719Arg allele and non-fatal MI by conditional logistic regression and adjusted for admixture of founder populations.Compared with the reference Trp/Trp homozygotes, KIF6 719Arg carriers were not at significantly higher risk for non-fatal MI in this study after adjustment for traditional risk factors or admixture (OR= 1.12; 95%CI, 0.98-1.28. Heterozygotes of the KIF6 Trp719Arg variant were at increased risk of non-fatal MI: the adjusted odds ratio was 1.16 (95% confidence interval, 1.01-1.34, but this association would not be significant after a multiple testing correction.We found that carriers of the KIF6 719Arg allele were not at increased risk of non-fatal MI in a case-control study of Costa Ricans living in the Central Valley of Costa Rica.

FRET measurements of kinesin neck orientation reveal a structural basis for processivity and asymmetry.

Science.gov (United States)

Martin, Douglas S; Fathi, Reza; Mitchison, Timothy J; Gelles, Jeff

2010-03-23

As the smallest and simplest motor enzymes, kinesins have served as the prototype for understanding the relationship between protein structure and mechanochemical function of enzymes in this class. Conventional kinesin (kinesin-1) is a motor enzyme that transports cargo toward the plus end of microtubules by a processive, asymmetric hand-over-hand mechanism. The coiled-coil neck domain, which connects the two kinesin motor domains, contributes to kinesin processivity (the ability to take many steps in a row) and is proposed to be a key determinant of the asymmetry in the kinesin mechanism. While previous studies have defined the orientation and position of microtubule-bound kinesin motor domains, the disposition of the neck coiled-coil remains uncertain. We determined the neck coiled-coil orientation using a multidonor fluorescence resonance energy transfer (FRET) technique to measure distances between microtubules and bound kinesin molecules. Microtubules were labeled with a new fluorescent taxol donor, TAMRA-X-taxol, and kinesin derivatives with an acceptor fluorophore attached at positions on the motor and neck coiled-coil domains were used to reconstruct the positions and orientations of the domains. FRET measurements to positions on the motor domain were largely consistent with the domain orientation determined in previous studies, validating the technique. Measurements to positions on the neck coiled-coil were inconsistent with a radial orientation and instead demonstrated that the neck coiled-coil is parallel to the microtubule surface. The measured orientation provides a structural explanation for how neck surface residues enhance processivity and suggests a simple hypothesis for the origin of kinesin step asymmetry and "limping."

Kinesin-8 effects on mitotic microtubule dynamics contribute to spindle function in fission yeast

Science.gov (United States)

Gergely, Zachary R.; Crapo, Ammon; Hough, Loren E.; McIntosh, J. Richard; Betterton, Meredith D.

2016-01-01

Kinesin-8 motor proteins destabilize microtubules. Their absence during cell division is associated with disorganized mitotic chromosome movements and chromosome loss. Despite recent work studying effects of kinesin-8s on microtubule dynamics, it remains unclear whether the kinesin-8 mitotic phenotypes are consequences of their effect on microtubule dynamics, their well-established motor activity, or additional, unknown functions. To better understand the role of kinesin-8 proteins in mitosis, we studied the effects of deletion of the fission yeast kinesin-8 proteins Klp5 and Klp6 on chromosome movements and spindle length dynamics. Aberrant microtubule-driven kinetochore pushing movements and tripolar mitotic spindles occurred in cells lacking Klp5 but not Klp6. Kinesin-8–deletion strains showed large fluctuations in metaphase spindle length, suggesting a disruption of spindle length stabilization. Comparison of our results from light microscopy with a mathematical model suggests that kinesin-8–induced effects on microtubule dynamics, kinetochore attachment stability, and sliding force in the spindle can explain the aberrant chromosome movements and spindle length fluctuations seen. PMID:27146110

Coupling of kinesin ATP turnover to translocation and microtubule regulation: one engine, many machines.

Science.gov (United States)

Friel, Claire T; Howard, Jonathon

2012-12-01

The cycle of ATP turnover is integral to the action of motor proteins. Here we discuss how variation in this cycle leads to variation of function observed amongst members of the kinesin superfamily of microtubule associated motor proteins. Variation in the ATP turnover cycle among superfamily members can tune the characteristic kinesin motor to one of the range of microtubule-based functions performed by kinesins. The speed at which ATP is hydrolysed affects the speed of translocation. The ratio of rate constants of ATP turnover in relation to association and dissociation from the microtubule influence the processivity of translocation. Variation in the rate-limiting step of the cycle can reverse the way in which the motor domain interacts with the microtubule producing non-motile kinesins. Because the ATP turnover cycle is not fully understood for the majority of kinesins, much work remains to show how the kinesin engine functions in such a wide variety of molecular machines.

A Kinesin-Related Protein Required for the Mitotic Spindle Assembly

Science.gov (United States)

1999-05-01

A. Pereira, P. Pesavento , Y. Yannoni, A.C. Spralding, and L.S.B. Goldstein. 1993. The kinesin-like protein KLP61F is essential for mitosis in...1169. 30. Heck MM, Pereira A, Pesavento P, Yannoni Y, Spradling AC, Goldstein LS: The kinesin-like protein KLP61F is essential for mitosis in

KIF3A and IL-4 are disease-specific biomarkers for psoriatic arthritis susceptibility

Science.gov (United States)

Ragazzo, Michele; Manzo, Laura; Costanza, Gaetana; Bowes, John; Hüffmeier, Ulrike; Potenza, Saverio; Sangiuolo, Federica; Reis, André; Barton, Anne; Novelli, Giuseppe; Orlandi, Augusto; Giardina, Emiliano

2017-01-01

To date, the genes associated with Psoriatic Arthritis (PsA) are principally involved in inflammation, immune response and epidermal differentiation, without any information about the relationship between disease and bone metabolism genes. Our work was focused on 5q31 locus, which contains several genetic variants significantly associated with PsA. The study involved 1526 subjects (500 PsA, 426 PsV, 600 controls). The region was evaluated by selecting and genotyping the SNPs of interest by Real Time PCR and direct sequencing. The results were subjected to biostatistic and bioinformatic analysis. The case-control study highlighted a significant association between KIF3A/IL-4 and PsA, but not with PsV (Psoriasis Vulgaris) patients. In addition, the haplotype analysis revealed two haplotypes significantly associated with PsA susceptibility. The Linkage Disequilibrium (LD) study showed the presence of a specific block in high LD within 132,692,628-132,737,638 bp of 5q31, giving additional evidence of specific association of the 5q31 region in PsA patients. Moreover, KIF3A expression was assessed by immunohistochemistry assays which showed a marked and significant difference of KIF3A expression between pathological and normal tissues. Our analysis described KIF3A and IL-4 as novel susceptibility genes for PsA, suggesting a clear implication of bone metabolism genes in the disease etiopathogenesis. PMID:29221136

Initial conformation of kinesin's neck linker

International Nuclear Information System (INIS)

Geng Yi-Zhao; Yan Shi-Wei; Ji Qing; Liu Shu-Xia

2014-01-01

How ATP binding initiates the docking process of kinesin's neck linker is a key question in understanding kinesin mechanisms. By exploiting a molecular dynamics method, we investigate the initial conformation of kinesin's neck linker in its docking process. We find that, in the initial conformation, the neck linker has interactions with β0 and forms a ‘cover-neck bundle’ structure with β0. From this initial structure, the formation of extra turns and the docking of the cover-neck bundle structure can be achieved. The motor head provides a forward force on the initial cover-neck bundle structure through ATP-induced rotation. This force, together with the hydrophobic interaction of ILE327 with the hydrophobic pocket on the motor head, drives the formation of the extra turn and initiates the neck linker docking process. Based on these findings, a pathway from ATP binding-induced motor head rotation to neck linker docking is proposed. (interdisciplinary physics and related areas of science and technology)

Bidirectional motility of the fission yeast kinesin-5, Cut7

Energy Technology Data Exchange (ETDEWEB)

Edamatsu, Masaki, E-mail: cedam@mail.ecc.u-tokyo.ac.jp

2014-03-28

Highlights: • Motile properties of Cut7 (fission yeast kinesin-5) were studied for the first time. • Half-length Cut7 moved toward plus-end direction of microtubule. • Full-length Cut7 moved toward minus-end direction of microtubule. • N- and C-terminal microtubule binding sites did not switch the motile direction. - Abstract: Kinesin-5 is a homotetrameric motor with its motor domain at the N-terminus. Kinesin-5 crosslinks microtubules and functions in separating spindle poles during mitosis. In this study, the motile properties of Cut7, fission yeast kinesin-5, were examined for the first time. In in vitro motility assays, full-length Cut7 moved toward minus-end of microtubules, but the N-terminal half of Cut7 moved toward the opposite direction. Furthermore, additional truncated constructs lacking the N-terminal or C-terminal regions, but still contained the motor domain, did not switch the motile direction. These indicated that Cut7 was a bidirectional motor, and microtubule binding regions at the N-terminus and C-terminus were not involved in its directionality.

SPG10 is a rare cause of spastic paraplegia in European families.

NARCIS (Netherlands)

Schule, R.; Kremer, H.P.H.; Kassubek, J.; Auer-Grumbach, M.; Kostic, V.; Klopstock, T.; Klimpe, S.; Otto, S.; Boesch, S.; Warrenburg, B.P.C. van de; Schols, L.

2008-01-01

BACKGROUND: SPG10 is an autosomal dominant form of hereditary spastic paraplegia (HSP), which is caused by mutations in the neural kinesin heavy chain KIF5A gene, the neuronal motor of fast anterograde axonal transport. Only four mutations have been identified to date. OBJECTIVE: To determine the

SPG10 is a rare cause of spastic paraplegia in European families

NARCIS (Netherlands)

Schuele, R.; Kremer, B. P. H.; Kassubek, J.; Auer-Grumbach, M.; Kostic, V.; Klopstock, T.; Klimpe, S.; Otto, S.; Boesch, S.; van de Warrenburg, B. P.; Schoels, L.

Background: SPG10 is an autosomal dominant form of hereditary spastic paraplegia (HSP), which is caused by mutations in the neural kinesin heavy chain KIF5A gene, the neuronal motor of fast anterograde axonal transport. Only four mutations have been identified to date. Objective: To determine the

Production of β-xylosidase from Trichoderma asperellum KIF125 and its application in efficient hydrolysis of pretreated rice straw with fungal cellulase.

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Inoue, Hiroyuki; Kitao, Chiaki; Yano, Shinichi; Sawayama, Shigeki

2016-11-01

On-site cellulase and hemicellulase production is a promising way to reduce enzyme cost in the commercialization of the lignocellulose-to-ethanol process. A hemicellulase-producing fungal strain suitable for on-site enzyme production was selected from cultures prepared using wet disc-milling rice straw (WDM-RS) and identified as Trichoderma asperellum KIF125. KIF125 hemicellulase showed uniquely high abundance of β-xylosidase in the xylanolytic enzyme system compared to other fungal hemicellulase preparations. Supplementation of Talaromyces cellulolyticus cellulase with KIF125 hemicellulase was more effective than that with the hemicellulases from other fungal sources in reducing the total enzyme loading for the improvement of xylose yield in the hydrolysis of ball-milling RS, due to its high β-xylosidase dominance. β-Xylosidase in KIF125 hemicellulase was purified and classified as a glycosyl hydrolase family 3 enzyme with relatively high specificity for xylobiose. The production of KIF125 β-xylosidase in the fermentor was estimated as 118 U/g-WDM-RS (2350 U/L culture) at 48 h. These results demonstrate that KIF125 is promising as a practical hemicellulase source to combine with on-site cellulase production using T. cellulolyticus.

Heterotrimeric Kinesin II Is the Microtubule Motor Protein Responsible for Pigment Dispersion in Xenopus Melanophores

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Tuma, M. Carolina; Zill, Andrew; Le Bot, Nathalie; Vernos, Isabelle; Gelfand, Vladimir

1998-01-01

Melanophores move pigment organelles (melanosomes) from the cell center to the periphery and vice-versa. These bidirectional movements require cytoplasmic microtubules and microfilaments and depend on the function of microtubule motors and a myosin. Earlier we found that melanosomes purified from Xenopus melanophores contain the plus end microtubule motor kinesin II, indicating that it may be involved in dispersion (Rogers, S.L., I.S. Tint, P.C. Fanapour, and V.I. Gelfand. 1997. Proc. Natl. Acad. Sci. USA. 94: 3720–3725). Here, we generated a dominant-negative construct encoding green fluorescent protein fused to the stalk-tail region of Xenopus kinesin-like protein 3 (Xklp3), the 95-kD motor subunit of Xenopus kinesin II, and introduced it into melanophores. Overexpression of the fusion protein inhibited pigment dispersion but had no effect on aggregation. To control for the specificity of this effect, we studied the kinesin-dependent movement of lysosomes. Neither dispersion of lysosomes in acidic conditions nor their clustering under alkaline conditions was affected by the mutant Xklp3. Furthermore, microinjection of melanophores with SUK4, a function-blocking kinesin antibody, inhibited dispersion of lysosomes but had no effect on melanosome transport. We conclude that melanosome dispersion is powered by kinesin II and not by conventional kinesin. This paper demonstrates that kinesin II moves membrane-bound organelles. PMID:9852150

Disruption of kif3a results in defective osteoblastic differentiation in dental mesenchymal stem/precursor cells via the Wnt signaling pathway.

Science.gov (United States)

Jiang, Sicong; Chen, Guoqing; Feng, Lian; Jiang, Zongting; Yu, Mei; Bao, Jinku; Tian, Weidong

2016-09-01

The anterograde intraflagellar transport motor protein, kif3a, regulates the integrity of primary cilia and various cellular functions, however, the role of kif3a in dental mesenchymal stem/precursor cell differentiation remains to be fully elucidated. In the present study, the expression of kif3a was knocked down in human dental follicle cells (hDFCs) and human dental pulp cells (hDPCs) using short hairpin RNA. The results of subsequent immunofluorescence revealed that knocking down kif3a resulted in the loss of primary cilia, which led to impairment of substantial mineralization and expression of the differentiation‑associated markers, including alkaline phosphatase, Runt‑related transcription factor 2, dentin matrix protein 1 and dentin sialophosphoprotein in the hDFCs and hDPCs. The results of reverse transcription‑quantitative polymerase chain reaction and western blot analyses showed that the expression levels of Wnt3a‑mediated active β‑catenin and lymphoid enhancer‑binding factor 1 were attenuated, whereas the expression of phosphorylated glycogen synthase kinase 3β was enhanced, in the kif3a‑knockdown cells. In addition, exogenous Wnt3a partially rescued osteoblastic differentiation in the hDFCs and hDPCs. These results demonstrated that inhibition of kif3a in the hDFCs and hDPCs disrupted primary cilia formation and/or function, and indicated that kif3a is important in the differentiation of hDFCs and hDPCs through the Wnt pathway. These findings not only enhance current understanding of tooth development and diseases of tooth mineralization, but also indicate possible strategies to regulate mineralization during tooth repair and regeneration.

Tumour Suppressor Adenomatous Polyposis Coli (APC) localisation is regulated by both Kinesin-1 and Kinesin-2

NARCIS (Netherlands)

Ruane, Peter T; Gumy, Laura F; Bola, Becky; Anderson, Beverley; Wozniak, Marcin J; Hoogenraad, Casper C; Allan, Victoria J

2016-01-01

Microtubules and their associated proteins (MAPs) underpin the polarity of specialised cells. Adenomatous polyposis coli (APC) is one such MAP with a multifunctional agenda that requires precise intracellular localisations. Although APC has been found to associate with kinesin-2 subfamily members,

Mechanical splitting of microtubules into protofilament bundles by surface-bound kinesin-1.

Science.gov (United States)

VanDelinder, Virginia; Adams, Peter G; Bachand, George D

2016-12-21

The fundamental biophysics of gliding microtubule (MT) motility by surface-tethered kinesin-1 motor proteins has been widely studied, as well as applied to capture and transport analytes in bioanalytical microdevices. In these systems, phenomena such as molecular wear and fracture into shorter MTs have been reported due the mechanical forces applied on the MT during transport. In the present work, we show that MTs can be split longitudinally into protofilament bundles (PFBs) by the work performed by surface-bound kinesin motors. We examine the properties of these PFBs using several techniques (e.g., fluorescence microscopy, SEM, AFM), and show that the PFBs continue to be mobile on the surface and display very high curvature compared to MT. Further, higher surface density of kinesin motors and shorter kinesin-surface tethers promote PFB formation, whereas modifying MT with GMPCPP or higher paclitaxel concentrations did not affect PFB formation.

KIF1A, an axonal transporter of synaptic vesicles, is mutated in hereditary sensory and autonomic neuropathy type 2.

Science.gov (United States)

Rivière, Jean-Baptiste; Ramalingam, Siriram; Lavastre, Valérie; Shekarabi, Masoud; Holbert, Sébastien; Lafontaine, Julie; Srour, Myriam; Merner, Nancy; Rochefort, Daniel; Hince, Pascale; Gaudet, Rébecca; Mes-Masson, Anne-Marie; Baets, Jonathan; Houlden, Henry; Brais, Bernard; Nicholson, Garth A; Van Esch, Hilde; Nafissi, Shahriar; De Jonghe, Peter; Reilly, Mary M; Timmerman, Vincent; Dion, Patrick A; Rouleau, Guy A

2011-08-12

Hereditary sensory and autonomic neuropathy type II (HSANII) is a rare autosomal-recessive disorder characterized by peripheral nerve degeneration resulting in a severe distal sensory loss. Although mutations in FAM134B and the HSN2 exon of WNK1 were associated with HSANII, the etiology of a substantial number of cases remains unexplained. In addition, the functions of WNK1/HSN2 and FAM134B and their role in the peripheral nervous system remain poorly understood. Using a yeast two-hybrid screen, we found that KIF1A, an axonal transporter of synaptic vesicles, interacts with the domain encoded by the HSN2 exon. In parallel to this screen, we performed genome-wide homozygosity mapping in a consanguineous Afghan family affected by HSANII and identified a unique region of homozygosity located on chromosome 2q37.3 and spanning the KIF1A gene locus. Sequencing of KIF1A in this family revealed a truncating mutation segregating with the disease phenotype. Subsequent sequencing of KIF1A in a series of 112 unrelated patients with features belonging to the clinical spectrum of ulcero-mutilating sensory neuropathies revealed truncating mutations in three additional families, thus indicating that mutations in KIF1A are a rare cause of HSANII. Similarly to WNK1 mutations, pathogenic mutations in KIF1A were almost exclusively restricted to an alternatively spliced exon. This study provides additional insights into the molecular pathogenesis of HSANII and highlights the potential biological relevance of alternative splicing in the peripheral sensory nervous system. Copyright © 2011 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

Evaluation of Activity and Combination Strategies with the Microtubule-Targeting Drug Sagopilone in Breast Cancer Cell Lines

International Nuclear Information System (INIS)

Eschenbrenner, Julia; Winsel, Sebastian; Hammer, Stefanie; Sommer, Anette; Mittelstaedt, Kevin; Drosch, Michael; Klar, Ulrich; Sachse, Christoph; Hannus, Michael; Seidel, Monika; Weiss, Bertram; Merz, Claudia; Siemeister, Gerhard; Hoffmann, Jens

2011-01-01

Sagopilone, a fully synthetic epothilone, is a microtubule-stabilizing agent optimized for high in vitro and in vivo activity against a broad range of tumor models, including those resistant to paclitaxel and other systemic treatments. Sagopilone development is accompanied by translational research studies to evaluate the molecular mode of action, to recognize mechanisms leading to resistance, to identify predictive response biomarkers, and to establish a rationale for combination with different therapies. Here, we profiled sagopilone activity in breast cancer cell lines. To analyze the mechanisms of mitotic arrest and apoptosis and to identify additional targets and biomarkers, an siRNA-based RNAi drug modifier screen interrogating 300 genes was performed in four cancer cell lines. Defects of the spindle assembly checkpoint (SAC) were identified to cause resistance against sagopilone-induced mitotic arrest and apoptosis. Potential biomarkers for resistance could therefore be functional defects like polymorphisms or mutations in the SAC, particularly in the central SAC kinase BUB1B. Moreover, chromosomal heterogeneity and polyploidy are also potential biomarkers of sagopilone resistance since they imply an increased tolerance for aberrant mitosis. RNAi screening further demonstrated that the sagopilone-induced mitotic arrest can be enhanced by concomitant inhibition of mitotic kinesins, thus suggesting a potential combination therapy of sagopilone with a KIF2C (MCAK) kinesin inhibitor. However, the combination of sagopilone and inhibition of the prophase kinesin KIF11 (EG5) is antagonistic, indicating that the kinesin inhibitor has to be highly specific to bring about the required therapeutic benefit.

Evaluation of Activity and Combination Strategies with the Microtubule-Targeting Drug Sagopilone in Breast Cancer Cell Lines

Energy Technology Data Exchange (ETDEWEB)

Eschenbrenner, Julia [Global Drug Discovery, Therapeutic Research Group Oncology, Bayer Healthcare Pharmaceuticals, Berlin (Germany); Institute for Biotechnology, Technical University Berlin, Berlin (Germany); Winsel, Sebastian [Global Drug Discovery, Therapeutic Research Group Oncology, Bayer Healthcare Pharmaceuticals, Berlin (Germany); Institute for Chemistry and Biochemistry, Free University Berlin, Berlin (Germany); Medical Biotechnology, VTT Technical Research Centre of Finland, Turku (Finland); Hammer, Stefanie [Global Drug Discovery, Therapeutic Research Group Oncology, Bayer Healthcare Pharmaceuticals, Berlin (Germany); Sommer, Anette [Global Drug Discovery, Target Discovery, Bayer Healthcare Pharmaceuticals, Berlin (Germany); Mittelstaedt, Kevin [Global Drug Discovery, Therapeutic Research Group Oncology, Bayer Healthcare Pharmaceuticals, Berlin (Germany); Institute for Chemistry and Biochemistry, Free University Berlin, Berlin (Germany); Department of Medicine, The University of Melbourne, Melbourne, VIC (Australia); Drosch, Michael [Global Drug Discovery, Therapeutic Research Group Oncology, Bayer Healthcare Pharmaceuticals, Berlin (Germany); Center of Human Genetics, University of Bremen, Bremen (Germany); Klar, Ulrich [Global Drug Discovery, Therapeutic Research Group Oncology, Bayer Healthcare Pharmaceuticals, Berlin (Germany); Sachse, Christoph; Hannus, Michael; Seidel, Monika [Cenix BioScience GmbH, Dresden (Germany); Weiss, Bertram; Merz, Claudia [Global Drug Discovery, Target Discovery, Bayer Healthcare Pharmaceuticals, Berlin (Germany); Siemeister, Gerhard [Global Drug Discovery, Therapeutic Research Group Oncology, Bayer Healthcare Pharmaceuticals, Berlin (Germany); Hoffmann, Jens, E-mail: jens.hoffmann@epo-berlin.com [Global Drug Discovery, Therapeutic Research Group Oncology, Bayer Healthcare Pharmaceuticals, Berlin (Germany); Experimentelle Pharmakologie und Onkologie Berlin-Buch GmbH, Berlin (Germany)

2011-11-16

Sagopilone, a fully synthetic epothilone, is a microtubule-stabilizing agent optimized for high in vitro and in vivo activity against a broad range of tumor models, including those resistant to paclitaxel and other systemic treatments. Sagopilone development is accompanied by translational research studies to evaluate the molecular mode of action, to recognize mechanisms leading to resistance, to identify predictive response biomarkers, and to establish a rationale for combination with different therapies. Here, we profiled sagopilone activity in breast cancer cell lines. To analyze the mechanisms of mitotic arrest and apoptosis and to identify additional targets and biomarkers, an siRNA-based RNAi drug modifier screen interrogating 300 genes was performed in four cancer cell lines. Defects of the spindle assembly checkpoint (SAC) were identified to cause resistance against sagopilone-induced mitotic arrest and apoptosis. Potential biomarkers for resistance could therefore be functional defects like polymorphisms or mutations in the SAC, particularly in the central SAC kinase BUB1B. Moreover, chromosomal heterogeneity and polyploidy are also potential biomarkers of sagopilone resistance since they imply an increased tolerance for aberrant mitosis. RNAi screening further demonstrated that the sagopilone-induced mitotic arrest can be enhanced by concomitant inhibition of mitotic kinesins, thus suggesting a potential combination therapy of sagopilone with a KIF2C (MCAK) kinesin inhibitor. However, the combination of sagopilone and inhibition of the prophase kinesin KIF11 (EG5) is antagonistic, indicating that the kinesin inhibitor has to be highly specific to bring about the required therapeutic benefit.

De novo mutations in the motor domain of KIF1A cause cognitive impairment, spastic paraparesis, axonal neuropathy, and cerebellar atrophy

NARCIS (Netherlands)

Lee, Jae Ran; Srour, Myriam; Kim, Doyoun; Hamdan, Fadi F.; Lim, So Hee; Brunel-Guitton, Catherine; Décarie, Jean Claude; Rossignol, Elsa; Mitchell, Grant A.; Schreiber, Allison; Moran, Rocio; Van Haren, Keith; Richardson, Randal; Nicolai, Joost; Oberndorff, Karin M E J; Wagner, Justin D.; Boycott, Kym M.; Rahikkala, Elisa; Junna, Nella; Tyynismaa, Henna; Cuppen, Inge; Verbeek, Nienke E.; Stumpel, Connie T R M; Willemsen, Michel A.; de Munnik, Sonja A.; Rouleau, Guy A.; Kim, Eunjoon; Kamsteeg, Erik Jan; Kleefstra, Tjitske; Michaud, Jacques L.

2015-01-01

KIF1A is a neuron-specific motor protein that plays important roles in cargo transport along neurites. Recessive mutations in KIF1A were previously described in families with spastic paraparesis or sensory and autonomic neuropathy type-2. Here, we report 11 heterozygous de novo missense mutations

Ribosomal protein NtRPL17 interacts with kinesin-12 family protein NtKRP and functions in the regulation of embryo/seed size and radicle growth.

Science.gov (United States)

Tian, Shujuan; Wu, Jingjing; Liu, Yuan; Huang, Xiaorong; Li, Fen; Wang, Zhaodan; Sun, Meng-Xiang

2017-11-28

We previously reported that a novel motor protein belonging to the kinesin-12 family, NtKRP, displays critical roles in regulating embryo and seed size establishment. However, it remains unknown exactly how NtKRP contributes to this developmental process. Here, we report that a 60S ribosomal protein NtRPL17 directly interacts with NtKRP. The phenotypes of NtRPL17 RNAi lines show notable embryo and seed size reduction. Structural observations of the NtRPL17-silenced embryos/seeds reveal that the embryo size reduction is due to a decrease in cell number. In these embryos, cell division cycle progression is delayed at the G2/M transition. These phenotypes are similar to that in NtKRP-silenced embryos/seeds, indicating that NtKRP and NtRPL17 function as partners in the same regulatory pathway during seed development and specifically regulate cell cycle progression to control embryo/seed size. This work reveals that NtRPL17, as a widely distributed ribosomal protein, plays a critical role in seed development and provides a new clue in the regulation of seed size. Confirmation of the interaction between NtKRP and NtRPL17 and their co-function in the control of the cell cycle also suggests that the mechanism might be conserved in both plants and animals. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

Effects of Obstacles on the Dynamics of Kinesins, Including Velocity and Run Length, Predicted by a Model of Two Dimensional Motion.

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Woochul Nam

Full Text Available Kinesins are molecular motors which walk along microtubules by moving their heads to different binding sites. The motion of kinesin is realized by a conformational change in the structure of the kinesin molecule and by a diffusion of one of its two heads. In this study, a novel model is developed to account for the 2D diffusion of kinesin heads to several neighboring binding sites (near the surface of microtubules. To determine the direction of the next step of a kinesin molecule, this model considers the extension in the neck linkers of kinesin and the dynamic behavior of the coiled-coil structure of the kinesin neck. Also, the mechanical interference between kinesins and obstacles anchored on the microtubules is characterized. The model predicts that both the kinesin velocity and run length (i.e., the walking distance before detaching from the microtubule are reduced by static obstacles. The run length is decreased more significantly by static obstacles than the velocity. Moreover, our model is able to predict the motion of kinesin when other (several motors also move along the same microtubule. Furthermore, it suggests that the effect of mechanical interaction/interference between motors is much weaker than the effect of static obstacles. Our newly developed model can be used to address unanswered questions regarding degraded transport caused by the presence of excessive tau proteins on microtubules.

Kinesin-dependent mechanism for controlling triglyceride secretion from the liver.

Science.gov (United States)

Rai, Priyanka; Kumar, Mukesh; Sharma, Geetika; Barak, Pradeep; Das, Saumitra; Kamat, Siddhesh S; Mallik, Roop

2017-12-05

Despite massive fluctuations in its internal triglyceride content, the liver secretes triglyceride under tight homeostatic control. This buffering function is most visible after fasting, when liver triglyceride increases manyfold but circulating serum triglyceride barely fluctuates. How the liver controls triglyceride secretion is unknown, but is fundamentally important for lipid and energy homeostasis in animals. Here we find an unexpected cellular and molecular mechanism behind such control. We show that kinesin motors are recruited to triglyceride-rich lipid droplets (LDs) in the liver by the GTPase ARF1, which is a key activator of lipolysis. This recruitment is activated by an insulin-dependent pathway and therefore responds to fed/fasted states of the animal. In fed state, ARF1 and kinesin appear on LDs, consequently transporting LDs to the periphery of hepatocytes where the smooth endoplasmic reticulum (sER) is present. Because the lipases that catabolize LDs in hepatocytes reside on the sER, LDs can now be catabolized efficiently to provide triglyceride for lipoprotein assembly and secretion from the sER. Upon fasting, insulin is lowered to remove ARF1 and kinesin from LDs, thus down-regulating LD transport and sER-LD contacts. This tempers triglyceride availabiity for very low density lipoprotein assembly and allows homeostatic control of serum triglyceride in a fasted state. We further show that kinesin knockdown inhibits hepatitis-C virus replication in hepatocytes, likely because translated viral proteins are unable to transfer from the ER to LDs. Copyright © 2017 the Author(s). Published by PNAS.

Late-onset spastic paraplegia type 10 (SPG10) family presenting with bulbar symptoms and fasciculations mimicking amyotrophic lateral sclerosis.

Science.gov (United States)

Kaji, Seiji; Kawarai, Toshitaka; Miyamoto, Ryosuke; Nodera, Hiroyuki; Pedace, Lucia; Orlacchio, Antonio; Izumi, Yuishin; Takahashi, Ryosuke; Kaji, Ryuji

2016-05-15

Pathogenic mutations in the KIF5A-SPG10 gene, encoding the kinesin HC5A, can be associated with autosomal dominant hereditary spastic paraplegia (ADHSP). It accounts for about 10% of the complicated forms of ADHSP. Peripheral neuropathy, distal upper limb amyotrophy, and cognitive decline are the most common additional clinical features. We examined a 66-year-old Japanese woman manifesting gait disturbance and spastic dysarthria for 6years with positive family history. She showed evidence of upper and lower motor neuron involvement and fasciculations, thus mimicking amyotrophic lateral sclerosis (ALS). Genetic analysis revealed a heterozygous variant in KIF5A (c.484C>T, p.Arg162Trp) in 2 symptomatic members. The mutation was also identified in 4 asymptomatic members, including 2 elderly members aged over 78years. Electromyography in the 2 symptomatic members revealed evidence of lower motor neuron involvement and fasciculation potentials in distal muscles. This report describes the first known Asian family with a KIF5A mutation and broadens the clinical and electrophysiological spectrum associated with KIF5A-SPG10 mutations. Given that our cases showed pseudobulbar palsy, fasciculation and altered penetrance, KIF5A-SPG10 might well be considered as a differential diagnosis of sporadic ALS. Copyright © 2016 Elsevier B.V. All rights reserved.

Monte Carlo analysis of neck linker extension in kinesin molecular motors.

Directory of Open Access Journals (Sweden)

Matthew L Kutys

2010-11-01

Full Text Available Kinesin stepping is thought to involve both concerted conformational changes and diffusive movement, but the relative roles played by these two processes are not clear. The neck linker docking model is widely accepted in the field, but the remainder of the step--diffusion of the tethered head to the next binding site--is often assumed to occur rapidly with little mechanical resistance. Here, we investigate the effect of tethering by the neck linker on the diffusive movement of the kinesin head, and focus on the predicted behavior of motors with naturally or artificially extended neck linker domains. The kinesin chemomechanical cycle was modeled using a discrete-state Markov chain to describe chemical transitions. Brownian dynamics were used to model the tethered diffusion of the free head, incorporating resistive forces from the neck linker and a position-dependent microtubule binding rate. The Brownian dynamics and chemomechanical cycle were coupled to model processive runs consisting of many 8 nm steps. Three mechanical models of the neck linker were investigated: Constant Stiffness (a simple spring, Increasing Stiffness (analogous to a Worm-Like Chain, and Reflecting (negligible stiffness up to a limiting contour length. Motor velocities and run lengths from simulated paths were compared to experimental results from Kinesin-1 and a mutant containing an extended neck linker domain. When tethered by an increasingly stiff spring, the head is predicted to spend an unrealistically short amount of time within the binding zone, and extending the neck is predicted to increase both the velocity and processivity, contrary to experiments. These results suggest that the Worm-Like Chain is not an adequate model for the flexible neck linker domain. The model can be reconciled with experimental data if the neck linker is either much more compliant or much stiffer than generally assumed, or if weak kinesin-microtubule interactions stabilize the diffusing

AP-1/KIF13A Blocking Peptides Impair Melanosome Maturation and Melanin Synthesis

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Cécile Campagne

2018-02-01

Full Text Available Melanocytes are specialized cells that generate unique organelles called melanosomes in which melanin is synthesized and stored. Melanosome biogenesis and melanocyte pigmentation require the transport and delivery of melanin synthesizing enzymes, such as tyrosinase and related proteins (e.g., TYRP1, from endosomes to maturing melanosomes. Among the proteins controlling endosome-melanosome transport, AP-1 together with KIF13A coordinates the endosomal sorting and trafficking of TYRP1 to melanosomes. We identify here β1-adaptin AP-1 subunit-derived peptides of 5 amino acids that block the interaction of KIF13A with AP-1 in cells. Incubating these peptides with human MNT-1 cells or 3D-reconstructed pigmented epidermis decreases pigmentation by impacting the maturation of melanosomes in fully pigmented organelles. This study highlights that peptides targeting the intracellular trafficking of melanocytes are candidate molecules to tune pigmentation in health and disease.

Vaccinia protein F12 has structural similarity to kinesin light chain and contains a motor binding motif required for virion export.

Directory of Open Access Journals (Sweden)

Gareth W Morgan

2010-02-01

Full Text Available Vaccinia virus (VACV uses microtubules for export of virions to the cell surface and this process requires the viral protein F12. Here we show that F12 has structural similarity to kinesin light chain (KLC, a subunit of the kinesin-1 motor that binds cargo. F12 and KLC share similar size, pI, hydropathy and cargo-binding tetratricopeptide repeats (TPRs. Moreover, molecular modeling of F12 TPRs upon the crystal structure of KLC2 TPRs showed a striking conservation of structure. We also identified multiple TPRs in VACV proteins E2 and A36. Data presented demonstrate that F12 is critical for recruitment of kinesin-1 to virions and that a conserved tryptophan and aspartic acid (WD motif, which is conserved in the kinesin-1-binding sequence (KBS of the neuronal protein calsyntenin/alcadein and several other cellular kinesin-1 binding proteins, is essential for kinesin-1 recruitment and virion transport. In contrast, mutation of WD motifs in protein A36 revealed they were not required for kinesin-1 recruitment or IEV transport. This report of a viral KLC-like protein containing a KBS that is conserved in several cellular proteins advances our understanding of how VACV recruits the kinesin motor to virions, and exemplifies how viruses use molecular mimicry of cellular components to their advantage.

Reversible control of kinesin activity and microtubule gliding speeds by switching the doping states of a conducting polymer support

Energy Technology Data Exchange (ETDEWEB)

Martin, Brett D [US Naval Research Laboratory, Code 6930, Washington, DC 20375 (United States); Velea, Luminita M [US Naval Research Laboratory, Code 6930, Washington, DC 20375 (United States); Soto, Carissa M [US Naval Research Laboratory, Code 6930, Washington, DC 20375 (United States); Whitaker, Craig M [US Naval Academy, Department of Chemistry, Annapolis, MD 21402 (United States); Gaber, Bruce P [US Naval Research Laboratory, Code 6930, Washington, DC 20375 (United States); Ratna, Banahalli [US Naval Research Laboratory, Code 6930, Washington, DC 20375 (United States)

2007-02-07

We describe a method for reversibly controlling the ATPase activity of streptavidin-linked kinesin by changing the doping states of a conducting polymer support. When the polymer (poly(CH{sub 2}OH-EDOT)) was electrochemically switched from its dedoped (semiconducting) state to its doped (conducting) state, the ATPase activity of the adsorbed kinesin complex decreased by 35% with a concomitant decrease in the gliding speeds of kinesin-driven microtubules. When the polymer was switched back to its original dedoped state, nearly identical increases were observed in the kinesin ATPase activity and microtubule speeds. Use of a fluorescent ATP substrate analogue showed that the total amount of kinesin adsorbed on the poly(CH{sub 2}OH-EDOT) surface remained constant as the doping state of the polymer was switched. The microtubules exhibited nearly identical speed differences on the doped and dedoped surfaces for both chemical and electrochemical doping methods. Michaelis-Menten modelling suggests that the doped surface acts as an 'uncompetitive inhibitor' of kinesin. This work represents an investigation into the phenomenon of an electrically switchable surface exerting a moderating effect on the activity of an adsorbed protein that does not contain a bound, electroactive metal ion.

Kinesin-3 and dynein cooperate in long-range retrograde endosome motility along a nonuniform microtubule array

NARCIS (Netherlands)

Schuster, M.; Kilaru, S.; Fink, G.; Collemare, J.A.R.; Roger, Y.; Steinberg, G.

2011-01-01

The polarity of microtubules (MTs) determines the motors for intracellular motility, with kinesins moving to plus ends and dynein to minus ends. In elongated cells of Ustilago maydis, dynein is thought to move early endosomes (EEs) toward the septum (retrograde), whereas kinesin-3 transports them to

Detectable states, cycle fluxes, and motility scaling of molecular motor kinesin: An integrative kinetic graph theory analysis

Science.gov (United States)

Ren, Jie

2017-12-01

The process by which a kinesin motor couples its ATPase activity with concerted mechanical hand-over-hand steps is a foremost topic of molecular motor physics. Two major routes toward elucidating kinesin mechanisms are the motility performance characterization of velocity and run length, and single-molecular state detection experiments. However, these two sets of experimental approaches are largely uncoupled to date. Here, we introduce an integrative motility state analysis based on a theorized kinetic graph theory for kinesin, which, on one hand, is validated by a wealth of accumulated motility data, and, on the other hand, allows for rigorous quantification of state occurrences and chemomechanical cycling probabilities. An interesting linear scaling for kinesin motility performance across species is discussed as well. An integrative kinetic graph theory analysis provides a powerful tool to bridge motility and state characterization experiments, so as to forge a unified effort for the elucidation of the working mechanisms of molecular motors.

In vivo collection of rare proteins using kinesin-based "nano-harvesters".

Energy Technology Data Exchange (ETDEWEB)

Bachand, Marlene; Bachand, George David; Greene, Adrienne Celeste; Carroll-Portillo, Amanda

2008-11-01

In this project, we have developed a novel platform for capturing, transport, and separating target analytes using the work harnessed from biomolecular transport systems. Nanoharvesters were constructed by co-organizing kinesin motor proteins and antibodies on a nanocrystal quantum dot (nQD) scaffold. Attachment of kinesin and antibodies to the nQD was achieved through biotin-streptavidin non-covalent bonds. Assembly of the nanoharvesters was characterized using a modified enzyme-linked immunosorbent assay (ELISA) that confirmed attachment of both proteins. Nanoharvesters selective against tumor necrosis factor-{alpha} (TNF-{alpha}) and nuclear transcription factor-{kappa}B (NF-{kappa}B) were capable of detecting target antigens at <100 ng/mL in ELISAs. A motility-based assay was subsequently developed using an antibody-sandwich approach in which the target antigen (TNF-{alpha}) formed a sandwich with the red-emitting nanoharvester and green-emitting detection nQD. In this format, successful sandwich formation resulted in a yellow emission associated with surface-bound microtubules. Step-wise analysis of sandwich formation suggested that the motility function of the kinesin motors was not adversely affected by either antigen capture or the subsequent binding of the detection nQDs. TNF-{alpha} was detected as low as {approx}1.5 ng/mL TNF-{alpha}, with 5.2% of the nanoharvesters successfully capturing the target analyte and detection nQDs. Overall, these results demonstrate the ability to capture target protein analytes in vitro using the kinesin-based nanoharvesters in nanofluidic environments. This system has direct relevance for lab-on-a-chip applications where pressure-driven or electrokinetic movement of fluids is impractical, and offers potential application for in vivo capture of rare proteins within the cytoplasmic domain of live cells.

Mapping the structural and dynamical features of kinesin motor domains.

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Guido Scarabelli

Full Text Available Kinesin motor proteins drive intracellular transport by coupling ATP hydrolysis to conformational changes that mediate directed movement along microtubules. Characterizing these distinct conformations and their interconversion mechanism is essential to determining an atomic-level model of kinesin action. Here we report a comprehensive principal component analysis of 114 experimental structures along with the results of conventional and accelerated molecular dynamics simulations that together map the structural dynamics of the kinesin motor domain. All experimental structures were found to reside in one of three distinct conformational clusters (ATP-like, ADP-like and Eg5 inhibitor-bound. These groups differ in the orientation of key functional elements, most notably the microtubule binding α4-α5, loop8 subdomain and α2b-β4-β6-β7 motor domain tip. Group membership was found not to correlate with the nature of the bound nucleotide in a given structure. However, groupings were coincident with distinct neck-linker orientations. Accelerated molecular dynamics simulations of ATP, ADP and nucleotide free Eg5 indicate that all three nucleotide states could sample the major crystallographically observed conformations. Differences in the dynamic coupling of distal sites were also evident. In multiple ATP bound simulations, the neck-linker, loop8 and the α4-α5 subdomain display correlated motions that are absent in ADP bound simulations. Further dissection of these couplings provides evidence for a network of dynamic communication between the active site, microtubule-binding interface and neck-linker via loop7 and loop13. Additional simulations indicate that the mutations G325A and G326A in loop13 reduce the flexibility of these regions and disrupt their couplings. Our combined results indicate that the reported ATP and ADP-like conformations of kinesin are intrinsically accessible regardless of nucleotide state and support a model where neck

Deletion of the Tail Domain of the Kinesin-5 Cin8 Affects Its Directionality*

Science.gov (United States)

Düselder, André; Fridman, Vladimir; Thiede, Christina; Wiesbaum, Alice; Goldstein, Alina; Klopfenstein, Dieter R.; Zaitseva, Olga; Janson, Marcel E.; Gheber, Larisa; Schmidt, Christoph F.

2015-01-01

The bipolar kinesin-5 motors are one of the major players that govern mitotic spindle dynamics. Their bipolar structure enables them to cross-link and slide apart antiparallel microtubules (MTs) emanating from the opposing spindle poles. The budding yeast kinesin-5 Cin8 was shown to switch from fast minus-end- to slow plus-end-directed motility upon binding between antiparallel MTs. This unexpected finding revealed a new dimension of cellular control of transport, the mechanism of which is unknown. Here we have examined the role of the C-terminal tail domain of Cin8 in regulating directionality. We first constructed a stable dimeric Cin8/kinesin-1 chimera (Cin8Kin), consisting of head and neck linker of Cin8 fused to the stalk of kinesin-1. As a single dimeric motor, Cin8Kin switched frequently between plus and minus directionality along single MTs, demonstrating that the Cin8 head domains are inherently bidirectional, but control over directionality was lost. We next examined the activity of a tetrameric Cin8 lacking only the tail domains (Cin8Δtail). In contrast to wild-type Cin8, the motility of single molecules of Cin8Δtail in high ionic strength was slow and bidirectional, with almost no directionality switches. Cin8Δtail showed only a weak ability to cross-link MTs in vitro. In vivo, Cin8Δtail exhibited bias toward the plus-end of the MTs and was unable to support viability of cells as the sole kinesin-5 motor. We conclude that the tail of Cin8 is not necessary for bidirectional processive motion, but is controlling the switch between plus- and minus-end-directed motility. PMID:25991727

Depletion of kinesin 5B affects lysosomal distribution and stability and induces peri-nuclear accumulation of autophagosomes in cancer cells

DEFF Research Database (Denmark)

Cardoso, Carla M P; Groth-Pedersen, Line; Høyer-Hansen, Maria

2009-01-01

BACKGROUND: Enhanced lysosomal trafficking is associated with metastatic cancer. In an attempt to discover cancer relevant lysosomal motor proteins, we compared the lysosomal proteomes from parental MCF-7 breast cancer cells with those from highly invasive MCF-7 cells that express an active form...... in HeLa cervix carcinoma cells as analyzed by subcellular fractionation. The depletion of KIF5B triggered peripheral aggregations of lysosomes followed by lysosomal destabilization, and cell death in HeLa cells. Lysosomal exocytosis in response to plasma membrane damage as well as fluid phase...... cells. In KIF5B-depleted cells the autophagosomes formed and accumulated in the close proximity to the Golgi apparatus, whereas in the control cells they appeared uniformly distributed in the cytoplasm. CONCLUSIONS/SIGNIFICANCE: Our data identify KIF5B as a cancer relevant lysosomal motor protein...

Dauer pheromone and G-protein signaling modulate the coordination of intraflagellar transport kinesin motor proteins in C. elegans

NARCIS (Netherlands)

J.A. Burghoorn (Jan); M.P.J. Dekkers (Martijn); S. Rademakers (Suzanne); A.A.W. de Jong (Ton); R. Willemsen (Rob); P. Swoboda (Peter); J. McCafferty (Gert)

2010-01-01

textabstractCilia length and function are dynamically regulated by modulation of intraflagellar transport (IFT). The cilia of C. elegans amphid channel neurons provide an excellent model to study this process, since they use two different kinesins for anterograde transport: kinesin-II and OSM-3

Activities and specificities of homodimeric TALENs in Saccharomyces cerevisiae

KAUST Repository

Aouida, Mustapha; Piatek, Marek J.; Bangarusamy, Dhinoth Kumar; Mahfouz, Magdy M.

2013-01-01

The development of highly efficient genome engineering reagents is of paramount importance to launch the next wave of biotechnology. TAL effectors have been developed as an adaptable DNA binding scaffold that can be engineered to bind to any user-defined sequence. Thus, TAL-based DNA binding modules have been used to generate chimeric proteins for a variety of targeted genome modifications across eukaryotic species. For example, TAL effectors fused to the catalytic domain of FokI endonuclease (TALENs) were used to generate site-specific double strand breaks (DSBs), the repair of which can be harnessed to dictate user-desired, genome-editing outcomes. To cleave DNA, FokI endonuclease must dimerize which can be achieved using a pair of TALENs that bind to the DNA targeted in a tail-to-tail orientation with proper spacing allowing the dimer formation. Because TALENs binding to DNA are dependent on their repeat sequences and nucleotides binding specificities, homodimers and heterodimers binding can be formed. In the present study, we used several TALEN monomers with increased repeats binding degeneracy to allow homodimer formation at increased number of genomic loci. We assessed their binding specificities and genome modification activities. Our results indicate that homodimeric TALENs could be used to modify the yeast genome in a site-specific manner and their binding to the promoter regions might modulate the expression of target genes. Taken together, our data indicate that homodimeric TALENs could be used to achieve different engineering possibilities of biotechnological applications and that their transcriptional modulations need to be considered when analyzing their phenotypic effects. © 2013 Springer-Verlag.

Activities and specificities of homodimeric TALENs in Saccharomyces cerevisiae

KAUST Repository

Aouida, Mustapha

2013-10-01

The development of highly efficient genome engineering reagents is of paramount importance to launch the next wave of biotechnology. TAL effectors have been developed as an adaptable DNA binding scaffold that can be engineered to bind to any user-defined sequence. Thus, TAL-based DNA binding modules have been used to generate chimeric proteins for a variety of targeted genome modifications across eukaryotic species. For example, TAL effectors fused to the catalytic domain of FokI endonuclease (TALENs) were used to generate site-specific double strand breaks (DSBs), the repair of which can be harnessed to dictate user-desired, genome-editing outcomes. To cleave DNA, FokI endonuclease must dimerize which can be achieved using a pair of TALENs that bind to the DNA targeted in a tail-to-tail orientation with proper spacing allowing the dimer formation. Because TALENs binding to DNA are dependent on their repeat sequences and nucleotides binding specificities, homodimers and heterodimers binding can be formed. In the present study, we used several TALEN monomers with increased repeats binding degeneracy to allow homodimer formation at increased number of genomic loci. We assessed their binding specificities and genome modification activities. Our results indicate that homodimeric TALENs could be used to modify the yeast genome in a site-specific manner and their binding to the promoter regions might modulate the expression of target genes. Taken together, our data indicate that homodimeric TALENs could be used to achieve different engineering possibilities of biotechnological applications and that their transcriptional modulations need to be considered when analyzing their phenotypic effects. © 2013 Springer-Verlag.

Compound heterozygous loss-of-function mutations in KIF20A are associated with a novel lethal congenital cardiomyopathy in two siblings.

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Jacoba J Louw

2018-01-01

Full Text Available Congenital or neonatal cardiomyopathies are commonly associated with a poor prognosis and have multiple etiologies. In two siblings, a male and female, we identified an undescribed type of lethal congenital restrictive cardiomyopathy affecting the right ventricle. We hypothesized a novel autosomal recessive condition. To identify the cause, we performed genetic, in vitro and in vivo studies. Genome-wide SNP typing and parametric linkage analysis was done in a recessive model to identify candidate regions. Exome sequencing analysis was done in unaffected and affected siblings. In the linkage regions, we selected candidate genes that harbor two rare variants with predicted functional effects in the patients and for which the unaffected sibling is either heterozygous or homozygous reference. We identified two compound heterozygous variants in KIF20A; a maternal missense variant (c.544C>T: p.R182W and a paternal frameshift mutation (c.1905delT: p.S635Tfs*15. Functional studies confirmed that the R182W mutation creates an ATPase defective form of KIF20A which is not able to support efficient transport of Aurora B as part of the chromosomal passenger complex. Due to this, Aurora B remains trapped on chromatin in dividing cells and fails to translocate to the spindle midzone during cytokinesis. Translational blocking of KIF20A in a zebrafish model resulted in a cardiomyopathy phenotype. We identified a novel autosomal recessive congenital restrictive cardiomyopathy, caused by a near complete loss-of-function of KIF20A. This finding further illustrates the relationship of cytokinesis and congenital cardiomyopathy.

Relationship between Lipid Phenotypes, Overweight, Lipid Lowering Drug Response and KIF6 and HMG-CoA Genotypes in a Subset of the Brisighella Heart Study Population

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Sabrina Angelini

2017-12-01

Full Text Available The existence of genetic traits might explain the susceptibility to develop hypercholesterolemia and the inter-individual differences in statin response. This study was performed to evaluate whether individuals’ polymorphisms in HMG-CoA and KIF6 genes are independently associated with hypercholesterolemia, other lipid-associated traits, and statin response in unselected individuals enrolled in the Brisighella heart study (Survey 2012. A total of 1622 individuals, of which 183 under statin medication, were genotyped for a total of five polymorphisms (KIF6 rs20455, rs9471077, rs9462535; HMG-CoA rs3761740, rs3846662. The relationships between the five loci and clinical characteristics were analyzed. The principal basic parameters calculated on 12 h fasting blood included total cholesterol (TC, High Density Lipoprotein Cholesterol (HDL-C, Low-Density Lipoprotein Cholesterol (LDL-C, and triglycerides (TG. Hypercholesterolemia was defined as a TC >200 mg/dL or use of lipid-lowering medication. 965 individuals were characterized by hypercholesterolemia; these subjects were significantly older (p < 0.001, with body mass index (BMI and waist circumference significantly higher (p < 0.001 compared to the others. HMG-CoA rs3846662 GG genotype was significantly over-represented in the hypercholesterolemic group (p = 0.030. HMG-CoA rs3846662 genotype was associated with the level of TC and LDL-C. Furthermore, in the same subset of untreated subjects, we observed a significant correlation between the KIF6 rs20455 and HDL-C. KIF6 variants were associated with a significantly lower (rs20455 or higher (rs9471077 and rs9462535 risk of obesity, in males only. No association between responsiveness to statins and the polymorphisms under investigation were observed. Our results showed associations between HMG-CoA rs3846662 and KIF6 rs20455 and lipid phenotypes, which may have an influence on dyslipidemia-related events. Moreover, this represents the first study

Asymmetric switching in a homodimeric ABC transporter: a simulation study.

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Jussi Aittoniemi

2010-04-01

Full Text Available ABC transporters are a large family of membrane proteins involved in a variety of cellular processes, including multidrug and tumor resistance and ion channel regulation. Advances in the structural and functional understanding of ABC transporters have revealed that hydrolysis at the two canonical nucleotide-binding sites (NBSs is co-operative and non-simultaneous. A conserved core architecture of bacterial and eukaryotic ABC exporters has been established, as exemplified by the crystal structure of the homodimeric multidrug exporter Sav1866. Currently, it is unclear how sequential ATP hydrolysis arises in a symmetric homodimeric transporter, since it implies at least transient asymmetry at the NBSs. We show by molecular dynamics simulation that the initially symmetric structure of Sav1866 readily undergoes asymmetric transitions at its NBSs in a pre-hydrolytic nucleotide configuration. MgATP-binding residues and a network of charged residues at the dimer interface are shown to form a sequence of putative molecular switches that allow ATP hydrolysis only at one NBS. We extend our findings to eukaryotic ABC exporters which often consist of two non-identical half-transporters, frequently with degeneracy substitutions at one of their two NBSs. Interestingly, many residues involved in asymmetric conformational switching in Sav1866 are substituted in degenerate eukaryotic NBS. This finding strengthens recent suggestions that the interplay of a consensus and a degenerate NBS in eukaroytic ABC proteins pre-determines the sequence of hydrolysis at the two NBSs.

Protein friction limits diffusive and directed movements of kinesin motors on microtubules.

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Bormuth, Volker; Varga, Vladimir; Howard, Jonathon; Schäffer, Erik

2009-08-14

Friction limits the operation of macroscopic engines and is critical to the performance of micromechanical devices. We report measurements of friction in a biological nanomachine. Using optical tweezers, we characterized the frictional drag force of individual kinesin-8 motor proteins interacting with their microtubule tracks. At low speeds and with no energy source, the frictional drag was related to the diffusion coefficient by the Einstein relation. At higher speeds, the frictional drag force increased nonlinearly, consistent with the motor jumping 8 nanometers between adjacent tubulin dimers along the microtubule, and was asymmetric, reflecting the structural polarity of the microtubule. We argue that these frictional forces arise from breaking bonds between the motor domains and the microtubule, and they limit the speed and efficiency of kinesin.

Crystal structure of the Candida albicans Kar3 kinesin motor domain fused to maltose-binding protein

International Nuclear Information System (INIS)

Delorme, Caroline; Joshi, Monika; Allingham, John S.

2012-01-01

Highlights: ► The Candida albicans Kar3 motor domain structure was solved as a maltose-binding protein fusion. ► The electrostatic surface and part of the ATPase pocket of the motor domain differs markedly from other kinesins. ► The MBP–Kar3 interface highlights a new site for intramolecular or intermolecular interactions. -- Abstract: In the human fungal pathogen Candida albicans, the Kinesin-14 motor protein Kar3 (CaKar3) is critical for normal mitotic division, nuclear fusion during mating, and morphogenic transition from the commensal yeast form to the virulent hyphal form. As a first step towards detailed characterization of this motor of potential medical significance, we have crystallized and determined the X-ray structure of the motor domain of CaKar3 as a maltose-binding protein (MBP) fusion. The structure shows strong conservation of overall motor domain topology to other Kar3 kinesins, but with some prominent differences in one of the motifs that compose the nucleotide-binding pocket and the surface charge distribution. The MBP and Kar3 modules are arranged such that MBP interacts with the Kar3 motor domain core at the same site where the neck linker of conventional kinesins docks during the “ATP state” of the mechanochemical cycle. This site differs from the Kar3 neck–core interface in the recent structure of the ScKar3Vik1 heterodimer. The position of MBP is also completely distinct from the Vik1 subunit in this complex. This may suggest that the site of MBP interaction on the CaKar3 motor domain provides an interface for the neck, or perhaps a partner subunit, at an intermediate state of its motile cycle that has not yet been observed for Kinesin-14 motors.

Crystal structure of the Candida albicans Kar3 kinesin motor domain fused to maltose-binding protein

Energy Technology Data Exchange (ETDEWEB)

Delorme, Caroline; Joshi, Monika [Department of Biomedical and Molecular Sciences, Queen' s University, Kingston, ON, Canada K7L 3N6 (Canada); Allingham, John S., E-mail: allinghj@queensu.ca [Department of Biomedical and Molecular Sciences, Queen' s University, Kingston, ON, Canada K7L 3N6 (Canada)

2012-11-30

Highlights: Black-Right-Pointing-Pointer The Candida albicans Kar3 motor domain structure was solved as a maltose-binding protein fusion. Black-Right-Pointing-Pointer The electrostatic surface and part of the ATPase pocket of the motor domain differs markedly from other kinesins. Black-Right-Pointing-Pointer The MBP-Kar3 interface highlights a new site for intramolecular or intermolecular interactions. -- Abstract: In the human fungal pathogen Candida albicans, the Kinesin-14 motor protein Kar3 (CaKar3) is critical for normal mitotic division, nuclear fusion during mating, and morphogenic transition from the commensal yeast form to the virulent hyphal form. As a first step towards detailed characterization of this motor of potential medical significance, we have crystallized and determined the X-ray structure of the motor domain of CaKar3 as a maltose-binding protein (MBP) fusion. The structure shows strong conservation of overall motor domain topology to other Kar3 kinesins, but with some prominent differences in one of the motifs that compose the nucleotide-binding pocket and the surface charge distribution. The MBP and Kar3 modules are arranged such that MBP interacts with the Kar3 motor domain core at the same site where the neck linker of conventional kinesins docks during the 'ATP state' of the mechanochemical cycle. This site differs from the Kar3 neck-core interface in the recent structure of the ScKar3Vik1 heterodimer. The position of MBP is also completely distinct from the Vik1 subunit in this complex. This may suggest that the site of MBP interaction on the CaKar3 motor domain provides an interface for the neck, or perhaps a partner subunit, at an intermediate state of its motile cycle that has not yet been observed for Kinesin-14 motors.

Parsing the roles of neck-linker docking and tethered head diffusion in the stepping dynamics of kinesin.

Science.gov (United States)

Zhang, Zhechun; Goldtzvik, Yonathan; Thirumalai, D

2017-11-14

Kinesin walks processively on microtubules (MTs) in an asymmetric hand-over-hand manner consuming one ATP molecule per 16-nm step. The individual contributions due to docking of the approximately 13-residue neck linker to the leading head (deemed to be the power stroke) and diffusion of the trailing head (TH) that contributes in propelling the motor by 16 nm have not been quantified. We use molecular simulations by creating a coarse-grained model of the MT-kinesin complex, which reproduces the measured stall force as well as the force required to dislodge the motor head from the MT, to show that nearly three-quarters of the step occurs by bidirectional stochastic motion of the TH. However, docking of the neck linker to the leading head constrains the extent of diffusion and minimizes the probability that kinesin takes side steps, implying that both the events are necessary in the motility of kinesin and for the maintenance of processivity. Surprisingly, we find that during a single step, the TH stochastically hops multiple times between the geometrically accessible neighboring sites on the MT before forming a stable interaction with the target binding site with correct orientation between the motor head and the [Formula: see text] tubulin dimer.

Regulation of mitotic spindle formation by the RhoA guanine nucleotide exchange factor ARHGEF10

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Satoh Takaya

2009-07-01

Full Text Available Abstract Background The Dbl family guanine nucleotide exchange factor ARHGEF10 was originally identified as the product of the gene associated with slowed nerve-conduction velocities of peripheral nerves. However, the function of ARHGEF10 in mammalian cells is totally unknown at a molecular level. ARHGEF10 contains no distinctive functional domains except for tandem Dbl homology-pleckstrin homology and putative transmembrane domains. Results Here we show that RhoA is a substrate for ARHGEF10. In both G1/S and M phases, ARHGEF10 was localized in the centrosome in adenocarcinoma HeLa cells. Furthermore, RNA interference-based knockdown of ARHGEF10 resulted in multipolar spindle formation in M phase. Each spindle pole seems to contain a centrosome consisting of two centrioles and the pericentriolar material. Downregulation of RhoA elicited similar phenotypes, and aberrant mitotic spindle formation following ARHGEF10 knockdown was rescued by ectopic expression of constitutively activated RhoA. Multinucleated cells were not increased upon ARHGEF10 knockdown in contrast to treatment with Y-27632, a specific pharmacological inhibitor for the RhoA effector kinase ROCK, which induced not only multipolar spindle formation, but also multinucleation. Therefore, unregulated centrosome duplication rather than aberration in cytokinesis may be responsible for ARHGEF10 knockdown-dependent multipolar spindle formation. We further isolated the kinesin-like motor protein KIF3B as a binding partner of ARHGEF10. Knockdown of KIF3B again caused multipolar spindle phenotypes. The supernumerary centrosome phenotype was also observed in S phase-arrested osteosarcoma U2OS cells when the expression of ARHGEF10, RhoA or KIF3B was abrogated by RNA interference. Conclusion Collectively, our results suggest that a novel RhoA-dependent signaling pathway under the control of ARHGEF10 has a pivotal role in the regulation of the cell division cycle. This pathway is not involved in

Crystal structure of human importin-α1 (Rch1, revealing a potential autoinhibition mode involving homodimerization.

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Hideyuki Miyatake

Full Text Available In this study, we determined the crystal structure of N-terminal importin-β-binding domain (IBB-truncated human importin-α1 (ΔIBB-h-importin-α1 at 2.63 Å resolution. The crystal structure of ΔIBB-h-importin-α1 reveals a novel closed homodimer. The homodimer exists in an autoinhibited state in which both the major and minor nuclear localization signal (NLS binding sites are completely buried in the homodimerization interface, an arrangement that restricts NLS binding. Analytical ultracentrifugation studies revealed that ΔIBB-h-importin-α1 is in equilibrium between monomers and dimers and that NLS peptides shifted the equilibrium toward the monomer side. This finding suggests that the NLS binding sites are also involved in the dimer interface in solution. These results show that when the IBB domain dissociates from the internal NLS binding sites, e.g., by binding to importin-β, homodimerization possibly occurs as an autoinhibition state.

Kinesin-1 and mitochondrial motility control by discrimination of structurally equivalent but distinct subdomains in Ran-GTP-binding domains of Ran-binding protein 2.

Science.gov (United States)

Patil, Hemangi; Cho, Kyoung-in; Lee, James; Yang, Yi; Orry, Andrew; Ferreira, Paulo A

2013-03-27

The pleckstrin homology (PH) domain is a versatile fold that mediates a variety of protein-protein and protein-phosphatidylinositol lipid interactions. The Ran-binding protein 2 (RanBP2) contains four interspersed Ran GTPase-binding domains (RBD(n = 1-4)) with close structural homology to the PH domain of Bruton's tyrosine kinase. The RBD2, kinesin-binding domain (KBD) and RBD3 comprise a tripartite domain (R2KR3) of RanBP2 that causes the unfolding, microtubule binding and biphasic activation of kinesin-1, a crucial anterograde motor of mitochondrial motility. However, the interplay between Ran GTPase and R2KR3 of RanBP2 in kinesin-1 activation and mitochondrial motility is elusive. We use structure-function, biochemical, kinetic and cell-based assays with time-lapse live-cell microscopy of over 260,000 mitochondrial-motility-related events to find mutually exclusive subdomains in RBD2 and RBD3 towards Ran GTPase binding, kinesin-1 activation and mitochondrial motility regulation. The RBD2 and RBD3 exhibit Ran-GTP-independent, subdomain and stereochemical-dependent discrimination on the biphasic kinetics of kinesin-1 activation or regulation of mitochondrial motility. Further, KBD alone and R2KR3 stimulate and suppress, respectively, multiple biophysical parameters of mitochondrial motility. The regulation of the bidirectional transport of mitochondria by either KBD or R2KR3 is highly coordinated, because their kinetic effects are accompanied always by changes in mitochondrial motile events of either transport polarity. These studies uncover novel roles in Ran GTPase-independent subdomains of RBD2 and RBD3, and KBD of RanBP2, that confer antagonizing and multi-modal mechanisms of kinesin-1 activation and regulation of mitochondrial motility. These findings open new venues towards the pharmacological harnessing of cooperative and competitive mechanisms regulating kinesins, RanBP2 or mitochondrial motility in disparate human disorders.

Cytoplasmic streaming in Drosophila oocytes varies with kinesin activity and correlates with the microtubule cytoskeleton architecture.

Science.gov (United States)

Ganguly, Sujoy; Williams, Lucy S; Palacios, Isabel M; Goldstein, Raymond E

2012-09-18

Cells can localize molecules asymmetrically through the combined action of cytoplasmic streaming, which circulates their fluid contents, and specific anchoring mechanisms. Streaming also contributes to the distribution of nutrients and organelles such as chloroplasts in plants, the asymmetric position of the meiotic spindle in mammalian embryos, and the developmental potential of the zygote, yet little is known quantitatively about the relationship between streaming and the motor activity which drives it. Here we use Particle Image Velocimetry to quantify the statistical properties of Kinesin-dependent streaming during mid-oogenesis in Drosophila. We find that streaming can be used to detect subtle changes in Kinesin activity and that the flows reflect the architecture of the microtubule cytoskeleton. Furthermore, based on characterization of the rheology of the cytoplasm in vivo, we establish estimates of the number of Kinesins required to drive the observed streaming. Using this in vivo data as the basis of a model for transport, we suggest that the disordered character of transport at mid-oogenesis, as revealed by streaming, is an important component of the localization dynamics of the body plan determinant oskar mRNA.

A Mechanism for Cytoplasmic Streaming: Kinesin-Driven Alignment of Microtubules and Fast Fluid Flows.

Science.gov (United States)

Monteith, Corey E; Brunner, Matthew E; Djagaeva, Inna; Bielecki, Anthony M; Deutsch, Joshua M; Saxton, William M

2016-05-10

The transport of cytoplasmic components can be profoundly affected by hydrodynamics. Cytoplasmic streaming in Drosophila oocytes offers a striking example. Forces on fluid from kinesin-1 are initially directed by a disordered meshwork of microtubules, generating minor slow cytoplasmic flows. Subsequently, to mix incoming nurse cell cytoplasm with ooplasm, a subcortical layer of microtubules forms parallel arrays that support long-range, fast flows. To analyze the streaming mechanism, we combined observations of microtubule and organelle motions with detailed mathematical modeling. In the fast state, microtubules tethered to the cortex form a thin subcortical layer and undergo correlated sinusoidal bending. Organelles moving in flows along the arrays show velocities that are slow near the cortex and fast on the inward side of the subcortical microtubule layer. Starting with fundamental physical principles suggested by qualitative hypotheses, and with published values for microtubule stiffness, kinesin velocity, and cytoplasmic viscosity, we developed a quantitative coupled hydrodynamic model for streaming. The fully detailed mathematical model and its simulations identify key variables that can shift the system between disordered (slow) and ordered (fast) states. Measurements of array curvature, wave period, and the effects of diminished kinesin velocity on flow rates, as well as prior observations on f-actin perturbation, support the model. This establishes a concrete mechanistic framework for the ooplasmic streaming process. The self-organizing fast phase is a result of viscous drag on kinesin-driven cargoes that mediates equal and opposite forces on cytoplasmic fluid and on microtubules whose minus ends are tethered to the cortex. Fluid moves toward plus ends and microtubules are forced backward toward their minus ends, resulting in buckling. Under certain conditions, the buckling microtubules self-organize into parallel bending arrays, guiding varying directions

Synaptic Control of Secretory Trafficking in Dendrites

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Cyril Hanus

2014-06-01

Full Text Available Localized signaling in neuronal dendrites requires tight spatial control of membrane composition. Upon initial synthesis, nascent secretory cargo in dendrites exits the endoplasmic reticulum (ER from local zones of ER complexity that are spatially coupled to post-ER compartments. Although newly synthesized membrane proteins can be processed locally, the mechanisms that control the spatial range of secretory cargo transport in dendritic segments are unknown. Here, we monitored the dynamics of nascent membrane proteins in dendritic post-ER compartments under regimes of low or increased neuronal activity. In response to activity blockade, post-ER carriers are highly mobile and are transported over long distances. Conversely, increasing synaptic activity dramatically restricts the spatial scale of post-ER trafficking along dendrites. This activity-induced confinement of secretory cargo requires site-specific phosphorylation of the kinesin motor KIF17 by Ca2+/calmodulin-dependent protein kinases (CaMK. Thus, the length scales of early secretory trafficking in dendrites are tuned by activity-dependent regulation of microtubule-dependent transport.

Distribution of tubulin, kinesin, and dynein in light- and dark-adapted octopus retinas.

Science.gov (United States)

Martinez, J M; Elfarissi, H; De Velasco, B; Ochoa, G H; Miller, A M; Clark, Y M; Matsumoto, B; Robles, L J

2000-01-01

Cephalopod retinas exhibit several responses to light and dark adaptation, including rhabdom size changes, photopigment movements, and pigment granule migration. Light- and dark-directed rearrangements of microfilament and microtubule cytoskeletal transport pathways could drive these changes. Recently, we localized actin-binding proteins in light-/dark-adapted octopus rhabdoms and suggested that actin cytoskeletal rearrangements bring about the formation and degradation of rhabdomere microvilli subsets. To determine if the microtubule cytoskeleton and associated motor proteins control the other light/dark changes, we used immunoblotting and immunocytochemical procedures to map the distribution of tubulin, kinesin, and dynein in dorsal and ventral halves of light- and dark-adapted octopus retinas. Immunoblots detected alpha- and beta-tubulin, dynein intermediate chain, and kinesin heavy chain in extracts of whole retinas. Epifluorescence and confocal microscopy showed that the tubulin proteins were distributed throughout the retina with more immunoreactivity in retinas exposed to light. Kinesin localization was heavy in the pigment layer of light- and dark-adapted ventral retinas but was less prominent in the dorsal region. Dynein distribution also varied in dorsal and ventral retinas with more immunoreactivity in light- and dark-adapted ventral retinas and confocal microscopy emphasized the granular nature of this labeling. We suggest that light may regulate the distribution of microtubule cytoskeletal proteins in the octopus retina and that position, dorsal versus ventral, also influences the distribution of motor proteins. The microtubule cytoskeleton is most likely involved in pigment granule migration in the light and dark and with the movement of transport vesicles from the photoreceptor inner segments to the rhabdoms.

Microtubule–microtubule sliding by kinesin-1 is essential for normal cytoplasmic streaming in Drosophila oocytes

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Lu, Wen; Winding, Michael; Lakonishok, Margot; Wildonger, Jill

2016-01-01

Cytoplasmic streaming in Drosophila oocytes is a microtubule-based bulk cytoplasmic movement. Streaming efficiently circulates and localizes mRNAs and proteins deposited by the nurse cells across the oocyte. This movement is driven by kinesin-1, a major microtubule motor. Recently, we have shown that kinesin-1 heavy chain (KHC) can transport one microtubule on another microtubule, thus driving microtubule–microtubule sliding in multiple cell types. To study the role of microtubule sliding in oocyte cytoplasmic streaming, we used a Khc mutant that is deficient in microtubule sliding but able to transport a majority of cargoes. We demonstrated that streaming is reduced by genomic replacement of wild-type Khc with this sliding-deficient mutant. Streaming can be fully rescued by wild-type KHC and partially rescued by a chimeric motor that cannot move organelles but is active in microtubule sliding. Consistent with these data, we identified two populations of microtubules in fast-streaming oocytes: a network of stable microtubules anchored to the actin cortex and free cytoplasmic microtubules that moved in the ooplasm. We further demonstrated that the reduced streaming in sliding-deficient oocytes resulted in posterior determination defects. Together, we propose that kinesin-1 slides free cytoplasmic microtubules against cortically immobilized microtubules, generating forces that contribute to cytoplasmic streaming and are essential for the refinement of posterior determinants. PMID:27512034

Mechanisms controlling the temporal degradation of Nek2A and Kif18A by the APC/C-Cdc20 complex

DEFF Research Database (Denmark)

Sedgwick, G.G.; Hayward, D.G.; Nilsson, J.

2013-01-01

The Anaphase Promoting Complex/Cyclosome (APC/C) in complex with its co-activator Cdc20 is responsible for targeting proteins for ubiquitin-mediated degradation during mitosis. The activity of APC/C-Cdc20 is inhibited during prometaphase by the Spindle Assembly Checkpoint (SAC) yet certain...... substrates escape this inhibition. Nek2A degradation during prometaphase depends on direct binding of Nek2A to the APC/C via a C-terminal MR dipeptide but whether this motif alone is sufficient is not clear. Here, we identify Kif18A as a novel APC/C-Cdc20 substrate and show that Kif18A degradation depends...... by the APC/C. Nek2A and the mitotic checkpoint complex (MCC) have an overlap in APC/C subunit requirements for binding and we propose that Nek2A binds with high affinity to apo-APC/C and is degraded by the pool of Cdc20 that avoids inhibition by the SAC....

Genetic and Informatic Analyses Implicate Kif12 as a Candidate Gene within the Mpkd2 Locus That Modulates Renal Cystic Disease Severity in the Cys1cpk Mouse.

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Michal Mrug

Full Text Available We have previously mapped the interval on Chromosome 4 for a major polycystic kidney disease modifier (Mpkd of the B6(Cg-Cys1cpk/J mouse model of recessive polycystic kidney disease (PKD. Informatic analyses predicted that this interval contains at least three individual renal cystic disease severity-modulating loci (Mpkd1-3. In the current study, we provide further validation of these predicted effects using a congenic mouse line carrying the entire CAST/EiJ (CAST-derived Mpkd1-3 interval on the C57BL/6J background. We have also generated a derivative congenic line with a refined CAST-derived Mpkd1-2 interval and demonstrated its dominantly-acting disease-modulating effects (e.g., 4.2-fold increase in total cyst area; p<0.001. The relative strength of these effects allowed the use of recombinants from these crosses to fine map the Mpkd2 effects to a <14 Mbp interval that contains 92 RefSeq sequences. One of them corresponds to the previously described positional Mpkd2 candidate gene, Kif12. Among the positional Mpkd2 candidates, only expression of Kif12 correlates strongly with the expression pattern of Cys1 across multiple anatomical nephron structures and developmental time points. Also, we demonstrate that Kif12 encodes a primary cilium-associated protein. Together, these data provide genetic and informatic validation of the predicted renal cystic disease-modulating effects of Mpkd1-3 loci and implicate Kif12 as the candidate locus for Mpkd2.

Meta-analyses of KIF6 Trp719Arg in coronary heart disease and statin therapeutic effect.

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Ping Peng

Full Text Available The goal of our study is to assess the contribution of KIF6 Trp719Arg to both the risk of CHD and the efficacy of statin therapy in CHD patients.Meta-analysis of 8 prospective studies among 77,400 Caucasians provides evidence that 719Arg increases the risk of CHD (P<0.001, HR = 1.27, 95% CI = 1.15-1.41. However, another meta-analysis of 7 case-control studies among 65,200 individuals fails to find a significant relationship between Trp719Arg and the risk of CHD (P = 0.642, OR = 1.02, 95% CI = 0.95-1.08. This suggests that the contribution of Trp719Arg to CHD varies in different ethnic groups. Additional meta-analysis also shows that statin therapy only benefit the vascular patients carry 719Arg allele (P<0.001, relative ratio (RR = 0.60, 95% CI = 0.54-0.67. To examine the role of this genetic variant in CHD risk in Han Chinese, we have conducted a case-control study with 289 CHD cases, 193 non-CHD controls, and 329 unrelated healthy volunteers as healthy controls. On post hoc analysis, significant allele frequency difference of 719Arg is observed between female CHD cases and female total controls under the dominant model (P = 0.04, χ(2 = 4.228, df = 1, odd ratio (OR = 1.979, 95% confidence interval (CI = 1.023-3.828. Similar trends are observed for post hoc analysis between female CHD cases and female healthy controls (dominant model: P = 0.04, χ(2 = 4.231, df = 1, OR = 2.015, 95% CI = 1.024-3.964. Non-genetic CHD risk factors are not controlled in these analyses.Our meta-analysis demonstrates the role of Trp719Arg of KIF6 gene in the risk of CHD in Caucasians. The meta-analysis also suggests the role of this variant in statin therapeutic response in vascular diseases. Our case-control study suggests that Trp719Arg of KIF6 gene is associated with CHD in female Han Chinese through a post hoc analysis.

The contribution of a 9p21.3 variant, a KIF6 variant, and C-reactive protein to predicting risk of myocardial infarction in a prospective study

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Tracy Russell P

2011-03-01

Full Text Available Abstract Background Genetic risk factors might improve prediction of coronary events. Several variants at chromosome 9p21.3 have been widely reported to be associated with coronary heart disease (CHD in prospective and case-control studies. A variant of KIF6 (719Arg has also been reported to be associated with increased risk of CHD in large prospective studies, but not in case-control studies. We asked whether the addition of genetic information (the 9p21.3 or KIF6 variants or a well-established non-genetic risk factor (C-reactive protein [CRP] can improve risk prediction by the Framingham Risk Score (FRS in the Cardiovascular Health Study (CHS--a prospective observational study of risk factors for cardiovascular disease among > 5,000 participants aged 65 or older. Methods Improvement of risk prediction was assessed by change in the area under the receiver-operator characteristic curve (AUC and by net reclassification improvement (NRI. Results Among white participants the FRS was improved by addition of KIF6 719Arg carrier status among men as assessed by the AUC (from 0.581 to 0.596, P = 0.03 but not by NRI (NRI = 0.027, P = 0.32. Adding both CRP and 719Arg carrier status to the FRS improved risk prediction by the AUC (0.608, P = 0.02 and NRI (0.093, P = 0.008 in men, but not women (P ≥ 0.24. Conclusions While none of these risk markers individually or in combination improved risk prediction among women, a combination of KIF6 719Arg carrier status and CRP levels modestly improved risk prediction among white men; although this improvement is not significant after multiple-testing correction. These observations should be investigated in other prospective studies.

A unique set of SH3-SH3 interactions controls IB1 homodimerization

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Kristensen, Ole; Guenat, Sylvie; Dar, Imran

2006-01-01

Islet-brain 1 (IB1 or JIP-1) is a scaffold protein that interacts with components of the c-Jun N-terminal kinase (JNK) signal-transduction pathway. IB1 is expressed at high levels in neurons and in pancreatic beta-cells, where it controls expression of several insulin-secretory components...... reduces IB1-dependent basal JNK activity in 293T cells. Impaired dimerization also results in a reduction in glucose transporter type 2 expression and in glucose-dependent insulin secretion in pancreatic beta-cells. Taken together, these results indicate that IB1 homodimerization through its SH3 domain...

Kinesin and Dynein Mechanics: Measurement Methods and Research Applications.

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Abraham, Zachary; Hawley, Emma; Hayosh, Daniel; Webster-Wood, Victoria A; Akkus, Ozan

2018-02-01

Motor proteins play critical roles in the normal function of cells and proper development of organisms. Among motor proteins, failings in the normal function of two types of proteins, kinesin and dynein, have been shown to lead many pathologies, including neurodegenerative diseases and cancers. As such, it is critical to researchers to understand the underlying mechanics and behaviors of these proteins, not only to shed light on how failures may lead to disease, but also to guide research toward novel treatment and nano-engineering solutions. To this end, many experimental techniques have been developed to measure the force and motility capabilities of these proteins. This review will (a) discuss such techniques, specifically microscopy, atomic force microscopy (AFM), optical trapping, and magnetic tweezers, and (b) the resulting nanomechanical properties of motor protein functions such as stalling force, velocity, and dependence on adenosine triphosophate (ATP) concentrations will be comparatively discussed. Additionally, this review will highlight the clinical importance of these proteins. Furthermore, as the understanding of the structure and function of motor proteins improves, novel applications are emerging in the field. Specifically, researchers have begun to modify the structure of existing proteins, thereby engineering novel elements to alter and improve native motor protein function, or even allow the motor proteins to perform entirely new tasks as parts of nanomachines. Kinesin and dynein are vital elements for the proper function of cells. While many exciting experiments have shed light on their function, mechanics, and applications, additional research is needed to completely understand their behavior.

Lemur tyrosine kinase-2 signalling regulates kinesin-1 light chain-2 phosphorylation and binding of Smad2 cargo.

LENUS (Irish Health Repository)

Manser, C

2012-05-31

A recent genome-wide association study identified the gene encoding lemur tyrosine kinase-2 (LMTK2) as a susceptibility gene for prostate cancer. The identified genetic alteration is within intron 9, but the mechanisms by which LMTK2 may impact upon prostate cancer are not clear because the functions of LMTK2 are poorly understood. Here, we show that LMTK2 regulates a known pathway that controls phosphorylation of kinesin-1 light chain-2 (KLC2) by glycogen synthase kinase-3β (GSK3β). KLC2 phosphorylation by GSK3β induces the release of cargo from KLC2. LMTK2 signals via protein phosphatase-1C (PP1C) to increase inhibitory phosphorylation of GSK3β on serine-9 that reduces KLC2 phosphorylation and promotes binding of the known KLC2 cargo Smad2. Smad2 signals to the nucleus in response to transforming growth factor-β (TGFβ) receptor stimulation and transport of Smad2 by kinesin-1 is required for this signalling. We show that small interfering RNA loss of LMTK2 not only reduces binding of Smad2 to KLC2, but also inhibits TGFβ-induced Smad2 signalling. Thus, LMTK2 may regulate the activity of kinesin-1 motor function and Smad2 signalling.

A mutation in KIF7 is responsible for the autosomal recessive syndrome of macrocephaly, multiple epiphyseal dysplasia and distinctive facial appearance

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Ali Bassam R

2012-05-01

Full Text Available Abstract Background We previously reported the existence of a unique autosomal recessive syndrome consisting of macrocephaly, multiple epiphyseal dysplasia and distinctive facial appearance mapping to chromosome 15q26. Methods In this manuscript, we have used whole exome sequencing on two affected members of a consanguineous family with this condition and carried out detailed bioinformatics analysis to elucidate the causative mutation. Results Our analysis resulted in the identification of a homozygous p.N1060S missense mutation in a highly conserved residue in KIF7, a regulator of Hedgehog signaling that has been recently found to be causing Joubert syndrome, fetal hydrolethalus and acrocallosal syndromes. The phenotype in our patients partially overlaps with the phenotypes associated with those syndromes but they also exhibit some distinctive features including multiple epiphyseal dysplasia. Conclusions We report the first missense homozygous disease-causing mutation in KIF7 and expand the clinical spectrum associated with mutations in this gene to include multiple epiphyseal dysplasia. The missense nature of the mutation might account for the unique presentation in our patients.

ATPase Cycle of the Nonmotile Kinesin NOD Allows Microtubule End Tracking and Drives Chromosome Movement

Energy Technology Data Exchange (ETDEWEB)

Cochran, J.; Sindelar, C; Mulko, N; Collins, K; Kong, S; Hawley, R; Kull, F

2009-01-01

Segregation of nonexchange chromosomes during Drosophila melanogaster meiosis requires the proper function of NOD, a nonmotile kinesin-10. We have determined the X-ray crystal structure of the NOD catalytic domain in the ADP- and AMPPNP-bound states. These structures reveal an alternate conformation of the microtubule binding region as well as a nucleotide-sensitive relay of hydrogen bonds at the active site. Additionally, a cryo-electron microscopy reconstruction of the nucleotide-free microtubule-NOD complex shows an atypical binding orientation. Thermodynamic studies show that NOD binds tightly to microtubules in the nucleotide-free state, yet other nucleotide states, including AMPPNP, are weakened. Our pre-steady-state kinetic analysis demonstrates that NOD interaction with microtubules occurs slowly with weak activation of ADP product release. Upon rapid substrate binding, NOD detaches from the microtubule prior to the rate-limiting step of ATP hydrolysis, which is also atypical for a kinesin. We propose a model for NOD's microtubule plus-end tracking that drives chromosome movement.

The Association between KIF6 Single Nucleotide Polymorphism rs20455 and Serum Lipids in Filipino-American Women

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Irma B. Ancheta

2014-01-01

Full Text Available The Trp719Arg allele of KIF6 rs20455, a putative risk factor for CHD especially in those with elevated low-density lipoprotein cholesterol (LDL-C, was investigated in Filipino-American women (FAW, n=235 participating in health screenings in four cities. The rs20455 genotype of each subject was determined by a multiplex assay using a Luminex-OLA procedure. The risk allele Trp719Arg was present in 77% of the subjects. The genotype distribution was 23% Trp/Trp, 51% Arg/Trp, and 26% Arg/Arg. Genotype did not predict the presence of CHD risk factors. Moreover, LDL-C, HDL-C, and triglycerides mean values did not vary as a function of genotype. However, those with the Arg/Arg genotype on statin medication exhibited a significantly higher mean triglycerides level (P<0.01. Approximately 60% of participants regardless of genotype exhibited LDL-C levels ≥100 mg/dL but were not taking medication. Approximately 43% of those with the Trp719Arg risk allele on statins exhibited elevated LDL-C levels. Our study suggests that the Trp719Arg allele of KIF 6 rs20455 is common among Filipino-American women; thus, even with borderline LDL-C levels would benefit from statin treatment. Secondly, many participants did not exhibit guideline recommended LDL-C levels including many who were on statin drugs.

Engineering of a novel Ca2+-regulated kinesin molecular motor using a calmodulin dimer linker

International Nuclear Information System (INIS)

Shishido, Hideki; Maruta, Shinsaku

2012-01-01

Highlights: ► Engineered kinesin–M13 and calmodulin involving single cysteine were prepared. ► CaM mutant was cross-linked to dimer by bifunctional thiol reactive reagent. ► Kinesin–M13 was dimerized via CaM dimer in the presence of calcium. ► Function of the engineered kinesin was regulated by a Ca 2+ -calmodulin dimer linker. -- Abstract: The kinesin–microtubule system holds great promise as a molecular shuttle device within biochips. However, one current barrier is that such shuttles do not have “on–off” control of their movement. Here we report the development of a novel molecular motor powered by an accelerator and brake system, using a kinesin monomer and a calmodulin (CaM) dimer. The kinesin monomer, K355, was fused with a CaM target peptide (M13 peptide) at the C-terminal part of the neck region (K355–M13). We also prepared CaM dimers using CaM mutants (Q3C), (R86C), or (A147C) and crosslinkers that react with cysteine residues. Following induction of K355–M13 dimerization with CaM dimers, we measured K355–M13 motility and found that it can be reversibly regulated in a Ca 2+ -dependent manner. We also found that velocities of K355–M13 varied depending on the type and crosslink position of the CaM dimer used; crosslink length also had a moderate effect on motility. These results suggest Ca 2+ -dependent dimerization of K355–M13 could be used as a novel molecular shuttle, equipped with an accelerator and brake system, for biochip applications.

IRF5, PTPN22, CD28, IL2RA, KIF5A, BLK and TNFAIP3 genes polymorphisms and lupus susceptibility in a cohort from the Egypt Delta; relation to other ethnic groups.

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Elghzaly, Ashraf A; Metwally, Shereen S; El-Chennawi, Farha A; Elgayaar, Maha A; Mosaad, Youssef M; El-Toraby, Ehab E; Hegab, Mohsen M; Ibrahim, Saleh M

2015-07-01

To replicate a single nucleotide polymorphism (SNP) of known genes for lupus (IRF5 rs10488631, PTPN22 rs2476601, BLK rs2736340 and TNFAIP3 rs5029939) and other autoimmune diseases (CD28 rs1980422, IL2RA rs2104286 and KIF5A rs1678542) on a newly studied Egyptian cohort to investigate the genetic disparity with different studied ethnic groups in relation to lupus susceptibility. 170 Egyptian patients from Egypt Delta with SLE and 241 matched healthy controls were genotyped by Taqman real time PCR for the selected SNPs. The results revealed significant association with IRF5 (p<0.0001) and PTPN22 (p=0.008) and insignificant association with KIF5A, CD28, IL2RA, BLK and TNFAIP3 genes. This study may provide an additional evidence for the association between IRF5 and PTPN22 and lupus susceptibility and may exclude it for CD28, IL2RA, and KIF5A. Copyright © 2015 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.

Expression analyses of the genes harbored by the type 2 diabetes and pediatric BMI associated locus on 10q23

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Zhao Jianhua

2012-09-01

Full Text Available Abstract Background There is evidence that one of the key type 2 diabetes (T2D loci identified by GWAS exerts its influence early on in life through its impact on pediatric BMI. This locus on 10q23 harbors three genes, encoding hematopoietically expressed homeobox (HHEX, insulin-degrading enzyme (IDE and kinesin family member 11 (KIF11, respectively. Methods We analyzed the impact of adipogeneis on the mRNA and protein expression levels of these genes in the human adipocyte Simpson-Golabi-Behmel syndrome (SGBS cell line in order to investigate which could be the culprit gene(s in this region of linkage disequilibrium. Results Following activation of differentiation with a PPARγ ligand, we observed ~20% decrease in IDE, ~40% decrease in HHEX and in excess of 80% decrease in KIF11 mRNA levels when comparing the adipocyte and pre-adipocyte states. We also observed decreases in KIF11 and IDE protein levels, but conversely we observed a dramatic increase in HHEX protein levels. Subsequent time course experiments revealed some marked changes in expression as early as three hours after activation of differentiation. Conclusion Our data suggest that the expression of all three genes at this locus are impacted during SGBS adipogenesis and provides insights in to the possible mechanisms of how the genes at this 10q23 locus could influence both adipocyte differentiation and susceptibility to T2D through insulin resistance.

Dynamic Changes in Sarcoplasmic Reticulum Structure in Ventricular Myocytes

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Amanda L. Vega

2011-01-01

sarcoplasmic reticulum (SR and the sarcolemma where Ca2+ release is activated. Here, we tested the hypothesis that the SR is a structurally inert organelle in ventricular myocytes. Our data suggest that rather than being static, the SR undergoes frequent dynamic structural changes. SR boutons expressing functional ryanodine receptors moved throughout the cell, approaching or moving away from the sarcolemma of ventricular myocytes. These changes in SR structure occurred in the absence of changes in [Ca2+] during EC coupling. Microtubules and the molecular motors dynein and kinesin 1(Kif5b were important regulators of SR motility. These findings support a model in which the SR is a motile organelle capable of molecular motor protein-driven structural changes.

The kinesin AtPSS1 promotes synapsis and is required for proper crossover distribution in meiosis.

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Yann Duroc

2014-10-01

Full Text Available Meiotic crossovers (COs shape genetic diversity by mixing homologous chromosomes at each generation. CO distribution is a highly regulated process. CO assurance forces the occurrence of at least one obligatory CO per chromosome pair, CO homeostasis smoothes out the number of COs when faced with variation in precursor number and CO interference keeps multiple COs away from each other along a chromosome. In several organisms, it has been shown that cytoskeleton forces are transduced to the meiotic nucleus via KASH- and SUN-domain proteins, to promote chromosome synapsis and recombination. Here we show that the Arabidopsis kinesin AtPSS1 plays a major role in chromosome synapsis and regulation of CO distribution. In Atpss1 meiotic cells, chromosome axes and DNA double strand breaks (DSBs appear to form normally but only a variable portion of the genome synapses and is competent for CO formation. Some chromosomes fail to form the obligatory CO, while there is an increased CO density in competent regions. However, the total number of COs per cell is unaffected. We further show that the kinesin motor domain of AtPSS1 is required for its meiotic function, and that AtPSS1 interacts directly with WIP1 and WIP2, two KASH-domain proteins. Finally, meiocytes missing AtPSS1 and/or SUN proteins show similar meiotic defects suggesting that AtPSS1 and SUNs act in the same pathway. This suggests that forces produced by the AtPSS1 kinesin and transduced by WIPs/SUNs, are required to authorize complete synapsis and regulate maturation of recombination intermediates into COs. We suggest that a form of homeostasis applies, which maintains the total number of COs per cell even if only a part of the genome is competent for CO formation.

Small-angle X-ray scattering analysis reveals the ATP-bound monomeric state of the ATPase domain from the homodimeric MutL endonuclease, a GHKL phosphotransferase superfamily protein.

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Iino, Hitoshi; Hikima, Takaaki; Nishida, Yuya; Yamamoto, Masaki; Kuramitsu, Seiki; Fukui, Kenji

2015-05-01

DNA mismatch repair is an excision system that removes mismatched bases chiefly generated by replication errors. In this system, MutL endonucleases direct the excision reaction to the error-containing strand of the duplex by specifically incising the newly synthesized strand. Both bacterial homodimeric and eukaryotic heterodimeric MutL proteins belong to the GHKL ATPase/kinase superfamily that comprises the N-terminal ATPase and C-terminal dimerization regions. Generally, the GHKL proteins show large ATPase cycle-dependent conformational changes, including dimerization-coupled ATP binding of the N-terminal domain. Interestingly, the ATPase domain of human PMS2, a subunit of the MutL heterodimer, binds ATP without dimerization. The monomeric ATP-bound state of the domain has been thought to be characteristic of heterodimeric GHKL proteins. In this study, we characterized the ATP-bound state of the ATPase domain from the Aquifex aeolicus MutL endonuclease, which is a homodimeric GHKL protein unlike the eukaryotic MutL. Gel filtration, dynamic light scattering, and small-angle X-ray scattering analyses clearly showed that the domain binds ATP in a monomeric form despite its homodimeric nature. This indicates that the uncoupling of dimerization and ATP binding is a common feature among bacterial and eukaryotic MutL endonucleases, which we suggest is closely related to the molecular mechanisms underlying mismatch repair.

A novel mutation in KIF5A in a Malian family with spastic paraplegia and sensory loss.

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Guinto, Cheick O; Diarra, Salimata; Diallo, Salimata; Cissé, Lassana; Coulibaly, Thomas; Diallo, Seybou H; Taméga, Abdoulaye; Chen, Ke-Lian; Schindler, Alice B; Bagayoko, Koumba; Simaga, Assiatou; Blackstone, Craig; Fischbeck, Kenneth H; Landouré, Guida

2017-04-01

Hereditary spastic paraplegias (HSPs) are well-characterized disorders but rarely reported in Africa. We evaluated a Malian family in which three individuals had HSP and distal muscle atrophy and sensory loss. HSP panel testing identified a novel heterozygous missense mutation in KIF5A (c.1086G>C, p.Lys362Asn) that segregated with the disease (SPG10). Lys362 is highly conserved across species and Lys362Asn is predicted to be damaging. This study shows that HSPs are present in sub-Saharan Africa, although likely underdiagnosed. Increasing efficiency and decreasing costs of DNA sequencing will make it more feasible to diagnose HSPs in developing countries.

A smart dust biosensor powered by kinesin motors.

Science.gov (United States)

Fischer, Thorsten; Agarwal, Ashutosh; Hess, Henry

2009-03-01

Biosensors can be miniaturized by either injecting smaller volumes into micro- and nanofluidic devices or immersing increasingly sophisticated particles known as 'smart dust' into the sample. The term 'smart dust' originally referred to cubic-millimetre wireless semiconducting sensor devices that could invisibly monitor the environment in buildings and public spaces, but later it also came to include functional micrometre-sized porous silicon particles used to monitor yet smaller environments. The principal challenge in designing smart dust biosensors is integrating transport functions with energy supply into the device. Here, we report a hybrid microdevice that is powered by ATP and relies on antibody-functionalized microtubules and kinesin motors to transport the target analyte into a detection region. The transport step replaces the wash step in traditional double-antibody sandwich assays. Owing to their small size and autonomous function, we envision that large numbers of such smart dust biosensors could be inserted into organisms or distributed into the environment for remote sensing.

ATP-binding motifs play key roles in Krp1p, kinesin-related protein 1, function for bi-polar growth control in fission yeast

International Nuclear Information System (INIS)

Rhee, Dong Keun; Cho, Bon A; Kim, Hyong Bai

2005-01-01

Kinesin is a microtubule-based motor protein with various functions related to the cell growth and division. It has been reported that Krp1p, kinesin-related protein 1, which belongs to the kinesin heavy chain superfamily, localizes on microtubules and may play an important role in cytokinesis. However, the function of Krp1p has not been fully elucidated. In this study, we overexpressed an intact form and three different mutant forms of Krp1p in fission yeast constructed by site-directed mutagenesis in two ATP-binding motifs or by truncation of the leucine zipper-like motif (LZiP). We observed hyper-extended microtubules and the aberrant nuclear shape in Krp1p-overexpressed fission yeast. As a functional consequence, a point mutation of ATP-binding domain 1 (G89E) in Krp1p reversed the effect of Krp1p overexpression in fission yeast, whereas the specific mutation in ATP-binding domain 2 (G238E) resulted in the altered cell polarity. Additionally, truncation of the leucine zipper-like domain (LZiP) at the C-terminal of Krp1p showed a normal nuclear division. Taken together, we suggest that krp1p is involved in regulation of cell-polarized growth through ATP-binding motifs in fission yeast

Experimental protocols for and studies of the effects of surface passivation and water isotopes on the gliding speed of microtubules propelled by kinesin-1

Science.gov (United States)

Maloney, Roger Andrew

This dissertation explores how the kinesin-1 and microtubule system is affected by surface passivation and water isotopes. Surface passivation was found to affect the gliding speed that microtubules exhibit in the gliding motility assay and the lengths of microtubules supported by the passivation. It was also found that gliding speeds of microtubules are very sensitive to temperature changes. Studies changing the water isotope were a first attempt to investigate if changing the solvent changed the osmotic pressure of the solution kinesin and microtubules were in. No osmotic pressure changes were observed, however, the experiments using different isotopes of water did illuminate the possibility that kinesin may be sensitive to viscosity changes in the solvent. This experiment also suggests further experiments that can be specifically designed to probe osmotic pressure changes. This thesis was also the first thesis ever, to the best of the author's knowledge, to be done in a completely open format. All information and notebook entries that are related to it, as well as the thesis itself, can be found on the website OpenWetWare. The thesis can also be found there including all the different versions that went into its editing. The philosophy and process of making data open and accessible to every one is also discussed.

The E. coli monothiol glutaredoxin GrxD forms homodimeric and heterodimeric FeS cluster containing complexes.

Science.gov (United States)

Yeung, N; Gold, B; Liu, N L; Prathapam, R; Sterling, H J; Willams, E R; Butland, G

2011-10-18

Monothiol glutaredoxins (mono-Grx) represent a highly evolutionarily conserved class of proteins present in organisms ranging from prokaryotes to humans. Mono-Grxs have been implicated in iron sulfur (FeS) cluster biosynthesis as potential scaffold proteins and in iron homeostasis via an FeS-containing complex with Fra2p (homologue of E. coli BolA) in yeast and are linked to signal transduction in mammalian systems. However, the function of the mono-Grx in prokaryotes and the nature of an interaction with BolA-like proteins have not been established. Recent genome-wide screens for E. coli genetic interactions reported the synthetic lethality (combination of mutations leading to cell death; mutation of only one of these genes does not) of a grxD mutation when combined with strains defective in FeS cluster biosynthesis (isc operon) functions [Butland, G., et al. (2008) Nature Methods 5, 789-795]. These data connected the only E. coli mono-Grx, GrxD to a potential role in FeS cluster biosynthesis. We investigated GrxD to uncover the molecular basis of this synthetic lethality and observed that GrxD can form FeS-bound homodimeric and BolA containing heterodimeric complexes. These complexes display substantially different spectroscopic and functional properties, including the ability to act as scaffold proteins for intact FeS cluster transfer to the model [2Fe-2S] acceptor protein E. coli apo-ferredoxin (Fdx), with the homodimer being significantly more efficient. In this work, we functionally dissect the potential cellular roles of GrxD as a component of both homodimeric and heterodimeric complexes to ultimately uncover if either of these complexes performs functions linked to FeS cluster biosynthesis. © 2011 American Chemical Society

Dynamic microtubule organization and mitochondrial transport are regulated by distinct Kinesin-1 pathways

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Anna Melkov

2015-12-01

Full Text Available The microtubule (MT plus-end motor kinesin heavy chain (Khc is well known for its role in long distance cargo transport. Recent evidence showed that Khc is also required for the organization of the cellular MT network by mediating MT sliding. We found that mutations in Khc and the gene of its adaptor protein, kinesin light chain (Klc resulted in identical bristle morphology defects, with the upper part of the bristle being thinner and flatter than normal and failing to taper towards the bristle tip. We demonstrate that bristle mitochondria transport requires Khc but not Klc as a competing force to dynein heavy chain (Dhc. Surprisingly, we demonstrate for the first time that Dhc is the primary motor for both anterograde and retrograde fast mitochondria transport. We found that the upper part of Khc and Klc mutant bristles lacked stable MTs. When following dynamic MT polymerization via the use of GFP-tagged end-binding protein 1 (EB1, it was noted that at Khc and Klc mutant bristle tips, dynamic MTs significantly deviated from the bristle parallel growth axis, relative to wild-type bristles. We also observed that GFP-EB1 failed to concentrate as a focus at the tip of Khc and Klc mutant bristles. We propose that the failure of bristle tapering is due to defects in directing dynamic MTs at the growing tip. Thus, we reveal a new function for Khc and Klc in directing dynamic MTs during polarized cell growth. Moreover, we also demonstrate a novel mode of coordination in mitochondrial transport between Khc and Dhc.

The kinesin spindle protein inhibitor filanesib enhances the activity of pomalidomide and dexamethasone in multiple myeloma

OpenAIRE

Hernández-García, Susana; San-Segundo, Laura; González-Méndez, Lorena; Corchete, Luis A; Misiewicz-Krzeminska, Irena; Martín-Sánchez, Montserrat; López-Iglesias, Ana-Alicia; Algarín, Esperanza Macarena; Mogollón, Pedro; Díaz-Tejedor, Andrea; Paíno, Teresa; Tunquist, Brian; Mateos, María-Victoria; Gutiérrez, Norma C; Díaz-Rodriguez, Elena

2017-01-01

[EN]Kinesin spindle protein inhibition is known to be an effective therapeutic approach in several malignancies. Filanesib (ARRY-520), an inhibitor of this protein, has demonstrated activity in heavily pre-treated multiple myeloma patients. The aim of the work herein was to investigate the activity of filanesib in combination with pomalidomide plus dexamethasone backbone, and the mechanisms underlying the potential synergistic effect. The ability of filanesib to enhance the activity of pomali...

Mutation of Rice BC12/GDD1, Which Encodes a Kinesin-Like Protein That Binds to a GA Biosynthesis Gene Promoter, Leads to Dwarfism with Impaired Cell Elongation[W][OA

Science.gov (United States)

Li, Juan; Jiang, Jiafu; Qian, Qian; Xu, Yunyuan; Zhang, Cui; Xiao, Jun; Du, Cheng; Luo, Wei; Zou, Guoxing; Chen, Mingluan; Huang, Yunqing; Feng, Yuqi; Cheng, Zhukuan; Yuan, Ming; Chong, Kang

2011-01-01

The kinesins are a family of microtubule-based motor proteins that move directionally along microtubules and are involved in many crucial cellular processes, including cell elongation in plants. Less is known about kinesins directly regulating gene transcription to affect cellular physiological processes. Here, we describe a rice (Oryza sativa) mutant, gibberellin-deficient dwarf1 (gdd1), that has a phenotype of greatly reduced length of root, stems, spikes, and seeds. This reduced length is due to decreased cell elongation and can be rescued by exogenous gibberellic acid (GA3) treatment. GDD1 was cloned by a map-based approach, was expressed constitutively, and was found to encode the kinesin-like protein BRITTLE CULM12 (BC12). Microtubule cosedimentation assays revealed that BC12/GDD1 bound to microtubules in an ATP-dependent manner. Whole-genome microarray analysis revealed the expression of ent-kaurene oxidase (KO2), which encodes an enzyme involved in GA biosynthesis, was downregulated in gdd1. Electrophoretic mobility shift and chromatin immunoprecipitation assays revealed that GDD1 bound to the element ACCAACTTGAA in the KO2 promoter. In addition, GDD1 was shown to have transactivation activity. The level of endogenous GAs was reduced in gdd1, and the reorganization of cortical microtubules was altered. Therefore, BC12/GDD1, a kinesin-like protein with transcription regulation activity, mediates cell elongation by regulating the GA biosynthesis pathway in rice. PMID:21325138

Crystallization and X-ray diffraction analysis of the CH domain of the cotton kinesin GhKCH2

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Qin, Xinghua [China Agricultural University, No. 2 Yuanmingyuanxilu, Haidian District, Beijing 100094, People’s Republic of (China); The Fourth Military Medical University, No. 169 Changlexi Road, Xincheng District, Xi’an 710032, People’s Republic of (China); Chen, Ziwei; Li, Ping; Liu, Guoqin, E-mail: liu@cau.edu.cn [China Agricultural University, No. 2 Yuanmingyuanxilu, Haidian District, Beijing 100094, People’s Republic of (China)

2016-02-19

The cloning, expression, purification and crystallization of the CH domain of the plant-specific kinesin GhKCH2 is reported. GhKCH2 belongs to a group of plant-specific kinesins (KCHs) containing an actin-binding calponin homology (CH) domain in the N-terminus. Previous studies revealed that the GhKCH2 CH domain (GhKCH2-CH) had a higher affinity for F-actin (K{sub d} = 0.42 ± 0.02 µM) than most other CH-domain-containing proteins. To understand the underlying mechanism, prokaryotically expressed GhKCH2-CH (amino acids 30–166) was purified and crystallized. Crystals were grown by the sitting-drop vapour-diffusion method using 0.1 M Tris–HCl pH 7.0, 20%(w/v) PEG 8000 as a precipitant. The crystals diffracted to a resolution of 2.5 Å and belonged to space group P2{sub 1}, with unit-cell parameters a = 41.57, b = 81.92, c = 83.00 Å, α = 90.00, β = 97.31, γ = 90.00°. Four molecules were found in the asymmetric unit with a Matthews coefficient of 2.22 Å{sup 3} Da{sup −1}, corresponding to a solvent content of 44.8%.

The Kinesin Adaptor Calsyntenin-1 Organizes Microtubule Polarity and Regulates Dynamics during Sensory Axon Arbor Development

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Mary C. Halloran

2017-04-01

Full Text Available Axon growth and branching, and development of neuronal polarity are critically dependent on proper organization and dynamics of the microtubule (MT cytoskeleton. MTs must organize with correct polarity for delivery of diverse cargos to appropriate subcellular locations, yet the molecular mechanisms regulating MT polarity remain poorly understood. Moreover, how an actively branching axon reorganizes MTs to direct their plus ends distally at branch points is unknown. We used high-speed, in vivo imaging of polymerizing MT plus ends to characterize MT dynamics in developing sensory axon arbors in zebrafish embryos. We find that axonal MTs are highly dynamic throughout development, and that the peripheral and central axons of sensory neurons show differences in MT behaviors. Furthermore, we show that Calsyntenin-1 (Clstn-1, a kinesin adaptor required for sensory axon branching, also regulates MT polarity in developing axon arbors. In wild type neurons the vast majority of MTs are directed in the correct plus-end-distal orientation from early stages of development. Loss of Clstn-1 causes an increase in MTs polymerizing in the retrograde direction. These misoriented MTs most often are found near growth cones and branch points, suggesting Clstn-1 is particularly important for organizing MT polarity at these locations. Together, our results suggest that Clstn-1, in addition to regulating kinesin-mediated cargo transport, also organizes the underlying MT highway during axon arbor development.

A Trypanosoma brucei kinesin heavy chain promotes parasite growth by triggering host arginase activity.

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Géraldine De Muylder

2013-10-01

Full Text Available In order to promote infection, the blood-borne parasite Trypanosoma brucei releases factors that upregulate arginase expression and activity in myeloid cells.By screening a cDNA library of T. brucei with an antibody neutralizing the arginase-inducing activity of parasite released factors, we identified a Kinesin Heavy Chain isoform, termed TbKHC1, as responsible for this effect. Following interaction with mouse myeloid cells, natural or recombinant TbKHC1 triggered SIGN-R1 receptor-dependent induction of IL-10 production, resulting in arginase-1 activation concomitant with reduction of nitric oxide (NO synthase activity. This TbKHC1 activity was IL-4Rα-independent and did not mirror M2 activation of myeloid cells. As compared to wild-type T. brucei, infection by TbKHC1 KO parasites was characterized by strongly reduced parasitaemia and prolonged host survival time. By treating infected mice with ornithine or with NO synthase inhibitor, we observed that during the first wave of parasitaemia the parasite growth-promoting effect of TbKHC1-mediated arginase activation resulted more from increased polyamine production than from reduction of NO synthesis. In late stage infection, TbKHC1-mediated reduction of NO synthesis appeared to contribute to liver damage linked to shortening of host survival time.A kinesin heavy chain released by T. brucei induces IL-10 and arginase-1 through SIGN-R1 signaling in myeloid cells, which promotes early trypanosome growth and favors parasite settlement in the host. Moreover, in the late stage of infection, the inhibition of NO synthesis by TbKHC1 contributes to liver pathogenicity.

Activity of the kinesin spindle protein inhibitor ispinesib (SB-715992) in models of breast cancer

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Purcell, James W; Davis, Jefferson; Reddy, Mamatha; Martin, Shamra; Samayoa, Kimberly; Vo, Hung; Thomsen, Karen; Bean, Peter; Kuo, Wen Lin; Ziyad, Safiyyah; Billig, Jessica; Feiler, Heidi S; Gray, Joe W; Wood, Kenneth W; Cases, Sylvaine

2009-06-10

Ispinesib (SB-715992) is a potent inhibitor of kinesin spindle protein (KSP), a kinesin motor protein essential for the formation of a bipolar mitotic spindle and cell cycle progression through mitosis. Clinical studies of ispinesib have demonstrated a 9% response rate in patients with locally advanced or metastatic breast cancer, and a favorable safety profile without significant neurotoxicities, gastrointestinal toxicities or hair loss. To better understand the potential of ispinesib in the treatment of breast cancer we explored the activity of ispinesib alone and in combination several therapies approved for the treatment of breast cancer. We measured the ispinesib sensitivity and pharmacodynamic response of breast cancer cell lines representative of various subtypes in vitro and as xenografts in vivo, and tested the ability of ispinesib to enhance the anti-tumor activity of approved therapies. In vitro, ispinesib displayed broad anti-proliferative activity against a panel of 53 breast cell-lines. In vivo, ispinesib produced regressions in each of five breast cancer models, and tumor free survivors in three of these models. The effects of ispinesib treatment on pharmacodynamic markers of mitosis and apoptosis were examined in vitro and in vivo, revealing a greater increase in both mitotic and apoptotic markers in the MDA-MB-468 model than in the less sensitive BT-474 model. In vivo, ispinesib enhanced the anti-tumor activity of trastuzumab, lapatinib, doxorubicin, and capecitabine, and exhibited activity comparable to paclitaxel and ixabepilone. These findings support further clinical exploration of KSP inhibitors for the treatment of breast cancer.

Structural plasticity of the N-terminal capping helix of the TPR domain of kinesin light chain.

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The Quyen Nguyen

Full Text Available Kinesin1 plays a major role in neuronal transport by recruiting many different cargos through its kinesin light chain (KLC. Various structurally unrelated cargos interact with the conserved tetratricopeptide repeat (TPR domain of KLC. The N-terminal capping helix of the TPR domain exhibits an atypical sequence and structural features that may contribute to the versatility of the TPR domain to bind different cargos. We determined crystal structures of the TPR domain of both KLC1 and KLC2 encompassing the N-terminal capping helix and show that this helix exhibits two distinct and defined orientations relative to the rest of the TPR domain. Such a difference in orientation gives rise, at the N-terminal part of the groove, to the formation of one hydrophobic pocket, as well as to electrostatic variations at the groove surface. We present a comprehensive structural analysis of available KLC1/2-TPR domain structures that highlights that ligand binding into the groove can be specific of one or the other N-terminal capping helix orientations. Further, structural analysis reveals that the N-terminal capping helix is always involved in crystal packing contacts, especially in a TPR1:TPR1' contact which highlights its propensity to be a protein-protein interaction site. Together, these results underline that the structural plasticity of the N-terminal capping helix might represent a structural determinant for TPR domain structural versatility in cargo binding.

BORC/kinesin-1 ensemble drives polarized transport of lysosomes into the axon.

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Farías, Ginny G; Guardia, Carlos M; De Pace, Raffaella; Britt, Dylan J; Bonifacino, Juan S

2017-04-04

The ability of lysosomes to move within the cytoplasm is important for many cellular functions. This ability is particularly critical in neurons, which comprise vast, highly differentiated domains such as the axon and dendrites. The mechanisms that control lysosome movement in these domains, however, remain poorly understood. Here we show that an ensemble of BORC, Arl8, SKIP, and kinesin-1, previously shown to mediate centrifugal transport of lysosomes in nonneuronal cells, specifically drives lysosome transport into the axon, and not the dendrites, in cultured rat hippocampal neurons. This transport is essential for maintenance of axonal growth-cone dynamics and autophagosome turnover. Our findings illustrate how a general mechanism for lysosome dispersal in nonneuronal cells is adapted to drive polarized transport in neurons, and emphasize the importance of this mechanism for critical axonal processes.

BORC/kinesin-1 ensemble drives polarized transport of lysosomes into the axon

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Farías, Ginny G.; Guardia, Carlos M.; De Pace, Raffaella; Britt, Dylan J.; Bonifacino, Juan S.

2017-01-01

The ability of lysosomes to move within the cytoplasm is important for many cellular functions. This ability is particularly critical in neurons, which comprise vast, highly differentiated domains such as the axon and dendrites. The mechanisms that control lysosome movement in these domains, however, remain poorly understood. Here we show that an ensemble of BORC, Arl8, SKIP, and kinesin-1, previously shown to mediate centrifugal transport of lysosomes in nonneuronal cells, specifically drives lysosome transport into the axon, and not the dendrites, in cultured rat hippocampal neurons. This transport is essential for maintenance of axonal growth-cone dynamics and autophagosome turnover. Our findings illustrate how a general mechanism for lysosome dispersal in nonneuronal cells is adapted to drive polarized transport in neurons, and emphasize the importance of this mechanism for critical axonal processes. PMID:28320970

Self-homodimerization of an actinoporin by disulfide bridging reveals implications for their structure and pore formation.

Science.gov (United States)

Valle, Aisel; Pérez-Socas, Luis Benito; Canet, Liem; Hervis, Yadira de la Patria; de Armas-Guitart, German; Martins-de-Sa, Diogo; Lima, Jônatas Cunha Barbosa; Souza, Adolfo Carlos Barros; Barbosa, João Alexandre Ribeiro Gonçalves; de Freitas, Sonia Maria; Pazos, Isabel Fabiola

2018-04-26

The Trp111 to Cys mutant of sticholysin I, an actinoporin from Stichodactyla helianthus sea anemone, forms a homodimer via a disulfide bridge. The purified dimer is 193 times less hemolytic than the monomer. Ultracentrifugation, dynamic light scattering and size-exclusion chromatography demonstrate that monomers and dimers are the only independent oligomeric states encountered. Indeed, circular dichroism and fluorescence spectroscopies showed that Trp/Tyr residues participate in homodimerization and that the dimer is less thermostable than the monomer. A homodimer three-dimensional model was constructed and indicates that Trp147/Tyr137 are at the homodimer interface. Spectroscopy results validated the 3D-model and assigned 85° to the disulfide bridge dihedral angle responsible for dimerization. The homodimer model suggests that alterations in the membrane/carbohydrate-binding sites in one of the monomers, as result of dimerization, could explain the decrease in the homodimer ability to form pores.

KIF16B is a candidate gene for a novel autosomal-recessive intellectual disability syndrome.

Science.gov (United States)

Alsahli, Saud; Arold, Stefan T; Alfares, Ahmed; Alhaddad, Bader; Al Balwi, Mohammed; Kamsteeg, Erik-Jan; Al-Twaijri, Waleed; Alfadhel, Majid

2018-05-07

Intellectual disability (ID) and global developmental delay are closely related; the latter is reserved for children under the age of 5 years as it is challenging to reliably assess clinical severity in this population. ID is a common condition, with up to 1%-3% of the population being affected and leading to a huge social and economic impact. ID is attributed to genetic abnormalities most of the time; however, the exact role of genetic involvement in ID is yet to be determined. Whole exome sequencing (WES) has gained popularity in the workup for ID, and multiple studies have been published examining the diagnostic yield in identification of the disease-causing variant (16%-55%), with the genetic involvement increasing as intelligence quotient decreases. WES has also accelerated novel disease gene discovery in this field. We identified a novel biallelic variant in the KIF16B gene (NM_024704.4:c.3611T > G) in two brothers that may be the cause of their phenotype. © 2018 Wiley Periodicals, Inc.

Arabidopsis CDS blastp result: AK287457 [KOME

Lifescience Database Archive (English)

Full Text Available AK287457 J043019L17 At4g14330.1 68417.m02207 phragmoplast-associated kinesin-relate...d protein 2 (PAKRP2) identical to cDNA phragmoplast-associated kinesin-related protein 2 (PAKRP2) GI:16973450 3e-24 ...

The actin-binding protein capulet genetically interacts with the microtubule motor kinesin to maintain neuronal dendrite homeostasis.

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Paul M B Medina

Full Text Available BACKGROUND: Neurons require precise cytoskeletal regulation within neurites, containing microtubule tracks for cargo transport in axons and dendrites or within synapses containing organized actin. Due to the unique architecture and specialized function of neurons, neurons are particularly susceptible to perturbation of the cytoskeleton. Numerous actin-binding proteins help maintain proper cytoskeletal regulation. METHODOLOGY/PRINCIPAL FINDINGS: From a Drosophila forward genetic screen, we identified a mutation in capulet--encoding a conserved actin-binding protein--that causes abnormal aggregates of actin within dendrites. Through interaction studies, we demonstrate that simultaneous genetic inactivation of capulet and kinesin heavy chain, a microtubule motor protein, produces elongate cofilin-actin rods within dendrites but not axons. These rods resemble actin-rich structures induced in both mammalian neurodegenerative and Drosophila Alzheimer's models, but have not previously been identified by loss of function mutations in vivo. We further demonstrate that mitochondria, which are transported by Kinesin, have impaired distribution along dendrites in a capulet mutant. While Capulet and Cofilin may biochemically cooperate in certain circumstances, in neuronal dendrites they genetically antagonize each other. CONCLUSIONS/SIGNIFICANCE: The present study is the first molecularly defined loss of function demonstration of actin-cofilin rods in vivo. This study suggests that simultaneous, seemingly minor perturbations in neuronal dendrites can synergize producing severe abnormalities affecting actin, microtubules and mitochondria/energy availability in dendrites. Additionally, as >90% of Alzheimer's and Parkinson's cases are sporadic this study suggests mechanisms by which multiple mutations together may contribute to neurodegeneration instead of reliance on single mutations to produce disease.

The Association between KIF6 Single Nucleotide Polymorphism rs20455 and Serum Lipids in Filipino-American Women.

Science.gov (United States)

Ancheta, Irma B; Battie, Cynthia A; Richard, Dan; Ancheta, Christine V; Borja-Hart, Nancy; Volgman, Annabelle S; Conley, Yvette

2014-01-01

The Trp719Arg allele of KIF6 rs20455, a putative risk factor for CHD especially in those with elevated low-density lipoprotein cholesterol (LDL-C), was investigated in Filipino-American women (FAW, n = 235) participating in health screenings in four cities. The rs20455 genotype of each subject was determined by a multiplex assay using a Luminex-OLA procedure. The risk allele Trp719Arg was present in 77% of the subjects. The genotype distribution was 23% Trp/Trp, 51% Arg/Trp, and 26% Arg/Arg. Genotype did not predict the presence of CHD risk factors. Moreover, LDL-C, HDL-C, and triglycerides mean values did not vary as a function of genotype. However, those with the Arg/Arg genotype on statin medication exhibited a significantly higher mean triglycerides level (P Filipino-American women; thus, even with borderline LDL-C levels would benefit from statin treatment. Secondly, many participants did not exhibit guideline recommended LDL-C levels including many who were on statin drugs.

DISC1 Protein Regulates γ-Aminobutyric Acid, Type A (GABAA) Receptor Trafficking and Inhibitory Synaptic Transmission in Cortical Neurons.

Science.gov (United States)

Wei, Jing; Graziane, Nicholas M; Gu, Zhenglin; Yan, Zhen

2015-11-13

Association studies have suggested that Disrupted-in-Schizophrenia 1 (DISC1) confers a genetic risk at the level of endophenotypes that underlies many major mental disorders. Despite the progress in understanding the significance of DISC1 at neural development, the mechanisms underlying DISC1 regulation of synaptic functions remain elusive. Because alterations in the cortical GABA system have been strongly linked to the pathophysiology of schizophrenia, one potential target of DISC1 that is critically involved in the regulation of cognition and emotion is the GABAA receptor (GABAAR). We found that cellular knockdown of DISC1 significantly reduced GABAAR-mediated synaptic and whole-cell current, whereas overexpression of wild-type DISC1, but not the C-terminal-truncated DISC1 (a schizophrenia-related mutant), significantly increased GABAAR currents in pyramidal neurons of the prefrontal cortex. These effects were accompanied by DISC1-induced changes in surface GABAAR expression. Moreover, the regulation of GABAARs by DISC1 knockdown or overexpression depends on the microtubule motor protein kinesin 1 (KIF5). Our results suggest that DISC1 exerts an important effect on GABAergic inhibitory transmission by regulating KIF5/microtubule-based GABAAR trafficking in the cortex. The knowledge gained from this study would shed light on how DISC1 and the GABA system are linked mechanistically and how their interactions are critical for maintaining a normal mental state. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

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.

Loss of function of Saccharomyces cerevisiae kinesin-related CIN8 and KIP1 is suppressed by KAR3 motor domain mutations.

Science.gov (United States)

Hoyt, M A; He, L; Totis, L; Saunders, W S

1993-09-01

The kinesin-related products of the CIN8 and KIP1 genes of Saccharomyces cerevisiae redundantly perform an essential function in mitosis. The action of either gene-product is required for an outwardly directed force that acts upon the spindle poles. We have selected mutations that suppress the temperature-sensitivity of a cin8-temperature-sensitive kip1-delta strain. The extragenic suppressors analyzed were all found to be alleles of the KAR3 gene. KAR3 encodes a distinct kinesin-related protein whose action antagonizes Cin8p/Kip1p function. All seven alleles analyzed were altered within the region of KAR3 that encodes the putative force-generating (or "motor") domain. These mutations also suppressed the inviability associated with the cin8-delta kip1-delta genotype, a property not shared by a deletion of KAR3. Other properties of the suppressing alleles revealed that they were not null for function. Six of the seven were unaffected for the essential karyogamy and meiosis properties of KAR3 and the seventh was dominant for the suppressing trait. Our findings suggest that despite an antagonistic relationship between Cin8p/Kip1p and Kar3p, aspects of their mitotic roles may be similar.

Crystal Structures of the Tetratricopeptide Repeat Domains of Kinesin Light Chains: Insight into Cargo Recognition Mechanisms

Energy Technology Data Exchange (ETDEWEB)

Zhu, Haizhong; Lee, Han Youl; Tong, Yufeng; Hong, Bum-Soo; Kim, Kyung-Phil; Shen, Yang; Lim, Kyung Jik; Mackenzie, Farrell; Tempel, Wolfram; Park, Hee-Won (SGC-Toronto); (PPCS); (Toronto)

2012-10-23

Kinesin-1 transports various cargos along the axon by interacting with the cargos through its light chain subunit. Kinesin light chains (KLC) utilize its tetratricopeptide repeat (TPR) domain to interact with over 10 different cargos. Despite a high sequence identity between their TPR domains (87%), KLC1 and KLC2 isoforms exhibit differential binding properties towards some cargos. We determined the structures of human KLC1 and KLC2 tetratricopeptide repeat (TPR) domains using X-ray crystallography and investigated the different mechanisms by which KLCs interact with their cargos. Using isothermal titration calorimetry, we attributed the specific interaction between KLC1 and JNK-interacting protein 1 (JIP1) cargo to residue N343 in the fourth TRP repeat. Structurally, the N343 residue is adjacent to other asparagines and lysines, creating a positively charged polar patch within the groove of the TPR domain. Whereas, KLC2 with the corresponding residue S328 did not interact with JIP1. Based on these finding, we propose that N343 of KLC1 can form 'a carboxylate clamp' with its neighboring asparagine to interact with JIP1, similar to that of HSP70/HSP90 organizing protein-1's (HOP1) interaction with heat shock proteins. For the binding of cargos shared by KLC1 and KLC2, we propose a different site located within the groove but not involving N343. We further propose a third binding site on KLC1 which involves a stretch of polar residues along the inter-TPR loops that may form a network of hydrogen bonds to JIP3 and JIP4. Together, these results provide structural insights into possible mechanisms of interaction between KLC TPR domains and various cargo proteins.

The Association between KIF6 Single Nucleotide Polymorphism rs20455 and Serum Lipids in Filipino-American Women

Science.gov (United States)

Ancheta, Irma B.; Battie, Cynthia A.; Ancheta, Christine V.; Volgman, Annabelle S.; Conley, Yvette

2014-01-01

The Trp719Arg allele of KIF6 rs20455, a putative risk factor for CHD especially in those with elevated low-density lipoprotein cholesterol (LDL-C), was investigated in Filipino-American women (FAW, n = 235) participating in health screenings in four cities. The rs20455 genotype of each subject was determined by a multiplex assay using a Luminex-OLA procedure. The risk allele Trp719Arg was present in 77% of the subjects. The genotype distribution was 23% Trp/Trp, 51% Arg/Trp, and 26% Arg/Arg. Genotype did not predict the presence of CHD risk factors. Moreover, LDL-C, HDL-C, and triglycerides mean values did not vary as a function of genotype. However, those with the Arg/Arg genotype on statin medication exhibited a significantly higher mean triglycerides level (P Filipino-American women; thus, even with borderline LDL-C levels would benefit from statin treatment. Secondly, many participants did not exhibit guideline recommended LDL-C levels including many who were on statin drugs. PMID:24587901

Role of kinesin heavy chain in Crumbs localization along the rhabdomere elongation in Drosophila photoreceptor.

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Garrett P League

Full Text Available BACKGROUND: Crumbs (Crb, a cell polarity gene, has been shown to provide a positional cue for the extension of the apical membrane domain, adherens junction (AJ, and rhabdomere along the growing proximal-distal axis during Drosophila photoreceptor morphogenesis. In developing Drosophila photoreceptors, a stabilized microtubule structure was discovered and its presence was linked to polarity protein localization. It was therefore hypothesized that the microtubules may provide trafficking routes for the polarity proteins during photoreceptor morphogenesis. This study has examined whether Kinesin heavy chain (Khc, a subunit of the microtubule-based motor Kinesin-1, is essential in polarity protein localization in developing photoreceptors. METHODOLOGY/PRINCIPAL FINDINGS: Because a genetic interaction was found between crb and khc, Crb localization was examined in the developing photoreceptors of khc mutants. khc was dispensable during early eye differentiation and development. However, khc mutant photoreceptors showed a range of abnormalities in the apical membrane domain depending on the position along the proximal-distal axis in pupal photoreceptors. The khc mutant showed a progressive mislocalization in the apical domain along the distal-proximal axis during rhabdomere elongation. The khc mutation also led to a similar progressive defect in the stabilized microtubule structures, strongly suggesting that Khc is essential for microtubule structure and Crb localization during distal to proximal rhabdomere elongation in pupal morphogenesis. This role of Khc in apical domain control was further supported by khc's gain-of-function phenotype. Khc overexpression in photoreceptors caused disruption of the apical membrane domain and the stabilized microtubules in the developing photoreceptors. CONCLUSIONS/SIGNIFICANCE: In summary, we examined the role of khc in the regulation of the apical Crb domain in developing photoreceptors. Since the rhabdomeres in

VISTA - computational tools for comparative genomics

Energy Technology Data Exchange (ETDEWEB)

Frazer, Kelly A.; Pachter, Lior; Poliakov, Alexander; Rubin,Edward M.; Dubchak, Inna

2004-01-01

Comparison of DNA sequences from different species is a fundamental method for identifying functional elements in genomes. Here we describe the VISTA family of tools created to assist biologists in carrying out this task. Our first VISTA server at http://www-gsd.lbl.gov/VISTA/ was launched in the summer of 2000 and was designed to align long genomic sequences and visualize these alignments with associated functional annotations. Currently the VISTA site includes multiple comparative genomics tools and provides users with rich capabilities to browse pre-computed whole-genome alignments of large vertebrate genomes and other groups of organisms with VISTA Browser, submit their own sequences of interest to several VISTA servers for various types of comparative analysis, and obtain detailed comparative analysis results for a set of cardiovascular genes. We illustrate capabilities of the VISTA site by the analysis of a 180 kilobase (kb) interval on human chromosome 5 that encodes for the kinesin family member3A (KIF3A) protein.

Positioning of AMPA Receptor-Containing Endosomes Regulates Synapse Architecture

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Marta Esteves da Silva

2015-11-01

Full Text Available Lateral diffusion in the membrane and endosomal trafficking both contribute to the addition and removal of AMPA receptors (AMPARs at postsynaptic sites. However, the spatial coordination between these mechanisms has remained unclear, because little is known about the dynamics of AMPAR-containing endosomes. In addition, how the positioning of AMPAR-containing endosomes affects synapse organization and functioning has never been directly explored. Here, we used live-cell imaging in hippocampal neuron cultures to show that intracellular AMPARs are transported in Rab11-positive recycling endosomes, which frequently enter dendritic spines and depend on the microtubule and actin cytoskeleton. By using chemically induced dimerization systems to recruit kinesin (KIF1C or myosin (MyosinV/VI motors to Rab11-positive recycling endosomes, we controlled their trafficking and found that induced removal of recycling endosomes from spines decreases surface AMPAR expression and PSD-95 clusters at synapses. Our data suggest a mechanistic link between endosome positioning and postsynaptic structure and composition.

NR2B Expression in Rat DRG Is Differentially Regulated Following Peripheral Nerve Injuries That Lead to Transient or Sustained Stimuli-Evoked Hypersensitivity.

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Norcini, Monica; Sideris, Alexandra; Adler, Samantha M; Hernandez, Lourdes A M; Zhang, Jin; Blanck, Thomas J J; Recio-Pinto, Esperanza

2016-01-01

Following injury, primary sensory neurons undergo changes that drive central sensitization and contribute to the maintenance of persistent hypersensitivity. NR2B expression in the dorsal root ganglia (DRG) has not been previously examined in neuropathic pain models. Here, we investigated if changes in NR2B expression within the DRG are associated with hypersensitivities that result from peripheral nerve injuries. This was done by comparing the NR2B expression in the DRG derived from two modalities of the spared nerve injury (SNI) model, since each variant produces different neuropathic pain phenotypes. Using the electronic von Frey to stimulate the spared and non-spared regions of the hindpaws, we demonstrated that sural-SNI animals develop sustained neuropathic pain in both regions while the tibial-SNI animals recover. NR2B expression was measured at Day 23 and Day 86 post-injury. At Day 23 and 86 post-injury, sural-SNI animals display strong hypersensitivity, whereas tibial-SNI animals display 50 and 100% recovery from post-injury-induced hypersensitivity, respectively. In tibial-SNI at Day 86, but not at Day 23 the perinuclear region of the neuronal somata displayed an increase in NR2B protein. This retention of NR2B protein within the perinuclear region, which will render them non-functional, correlates with the recovery observed in tibial-SNI. In sural-SNI at Day 86, DRG displayed an increase in NR2B mRNA which correlates with the development of sustained hypersensitivity in this model. The increase in NR2B mRNA was not associated with an increase in NR2B protein within the neuronal somata. The latter may result from a decrease in kinesin Kif17, since Kif17 mediates NR2B transport to the soma's plasma membrane. In both SNIs, microglia/macrophages showed a transient increase in NR2B protein detected at Day 23 but not at Day 86, which correlates with the initial post-injury induced hypersensitivity in both SNIs. In tibial-SNI at Day 86, but not at Day 23

Diffusion as a Ruler: Modeling Kinesin Diffusion as a Length Sensor for Intraflagellar Transport.

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Hendel, Nathan L; Thomson, Matthew; Marshall, Wallace F

2018-02-06

An important question in cell biology is whether cells are able to measure size, either whole cell size or organelle size. Perhaps cells have an internal chemical representation of size that can be used to precisely regulate growth, or perhaps size is just an accident that emerges due to constraint of nutrients. The eukaryotic flagellum is an ideal model for studying size sensing and control because its linear geometry makes it essentially one-dimensional, greatly simplifying mathematical modeling. The assembly of flagella is regulated by intraflagellar transport (IFT), in which kinesin motors carry cargo adaptors for flagellar proteins along the flagellum and then deposit them at the tip, lengthening the flagellum. The rate at which IFT motors are recruited to begin transport into the flagellum is anticorrelated with the flagellar length, implying some kind of communication between the base and the tip and possibly indicating that cells contain some mechanism for measuring flagellar length. Although it is possible to imagine many complex scenarios in which additional signaling molecules sense length and carry feedback signals to the cell body to control IFT, might the already-known components of the IFT system be sufficient to allow length dependence of IFT? Here we investigate a model in which the anterograde kinesin motors unbind after cargo delivery, diffuse back to the base, and are subsequently reused to power entry of new IFT trains into the flagellum. By mathematically modeling and simulating such a system, we are able to show that the diffusion time of the motors can in principle be sufficient to serve as a proxy for length measurement. We found that the diffusion model can not only achieve a stable steady-state length without the addition of any other signaling molecules or pathways, but also is able to produce the anticorrelation between length and IFT recruitment rate that has been observed in quantitative imaging studies. Copyright © 2017 Biophysical

Unc-51/ATG1 controls axonal and dendritic development via kinesin-mediated vesicle transport in the Drosophila brain.

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Hiroaki Mochizuki

2011-05-01

Full Text Available Members of the evolutionary conserved Ser/Thr kinase Unc-51 family are key regulatory proteins that control neural development in both vertebrates and invertebrates. Previous studies have suggested diverse functions for the Unc-51 protein, including axonal elongation, growth cone guidance, and synaptic vesicle transport.In this work, we have investigated the functional significance of Unc-51-mediated vesicle transport in the development of complex brain structures in Drosophila. We show that Unc-51 preferentially accumulates in newly elongating axons of the mushroom body, a center of olfactory learning in flies. Mutations in unc-51 cause disintegration of the core of the developing mushroom body, with mislocalization of Fasciclin II (Fas II, an IgG-family cell adhesion molecule important for axonal guidance and fasciculation. In unc-51 mutants, Fas II accumulates in the cell bodies, calyx, and the proximal peduncle. Furthermore, we show that mutations in unc-51 cause aberrant overshooting of dendrites in the mushroom body and the antennal lobe. Loss of unc-51 function leads to marked accumulation of Rab5 and Golgi components, whereas the localization of dendrite-specific proteins, such as Down syndrome cell adhesion molecule (DSCAM and No distributive disjunction (Nod, remains unaltered. Genetic analyses of kinesin light chain (Klc and unc-51 double heterozygotes suggest the importance of kinesin-mediated membrane transport for axonal and dendritic development. Moreover, our data demonstrate that loss of Klc activity causes similar axonal and dendritic defects in mushroom body neurons, recapitulating the salient feature of the developmental abnormalities caused by unc-51 mutations.Unc-51 plays pivotal roles in the axonal and dendritic development of the Drosophila brain. Unc-51-mediated membrane vesicle transport is important in targeted localization of guidance molecules and organelles that regulate elongation and compartmentalization of

Expression of a truncated ATHB17 protein in maize increases ear weight at silking.

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Elena A Rice

Full Text Available ATHB17 (AT2G01430 is an Arabidopsis gene encoding a member of the α-subclass of the homeodomain leucine zipper class II (HD-Zip II family of transcription factors. The ATHB17 monomer contains four domains common to all class II HD-Zip proteins: a putative repression domain adjacent to a homeodomain, leucine zipper, and carboxy terminal domain. However, it also possesses a unique N-terminus not present in other members of the family. In this study we demonstrate that the unique 73 amino acid N-terminus is involved in regulation of cellular localization of ATHB17. The ATHB17 protein is shown to function as a transcriptional repressor and an EAR-like motif is identified within the putative repression domain of ATHB17. Transformation of maize with an ATHB17 expression construct leads to the expression of ATHB17Δ113, a truncated protein lacking the first 113 amino acids which encodes a significant portion of the repression domain. Because ATHB17Δ113 lacks the repression domain, the protein cannot directly affect the transcription of its target genes. ATHB17Δ113 can homodimerize, form heterodimers with maize endogenous HD-Zip II proteins, and bind to target DNA sequences; thus, ATHB17Δ113 may interfere with HD-Zip II mediated transcriptional activity via a dominant negative mechanism. We provide evidence that maize HD-Zip II proteins function as transcriptional repressors and that ATHB17Δ113 relieves this HD-Zip II mediated transcriptional repression activity. Expression of ATHB17Δ113 in maize leads to increased ear size at silking and, therefore, may enhance sink potential. We hypothesize that this phenotype could be a result of modulation of endogenous HD-Zip II pathways in maize.

Expression of a truncated ATHB17 protein in maize increases ear weight at silking.

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Rice, Elena A; Khandelwal, Abha; Creelman, Robert A; Griffith, Cara; Ahrens, Jeffrey E; Taylor, J Philip; Murphy, Lesley R; Manjunath, Siva; Thompson, Rebecca L; Lingard, Matthew J; Back, Stephanie L; Larue, Huachun; Brayton, Bonnie R; Burek, Amanda J; Tiwari, Shiv; Adam, Luc; Morrell, James A; Caldo, Rico A; Huai, Qing; Kouadio, Jean-Louis K; Kuehn, Rosemarie; Sant, Anagha M; Wingbermuehle, William J; Sala, Rodrigo; Foster, Matt; Kinser, Josh D; Mohanty, Radha; Jiang, Dongming; Ziegler, Todd E; Huang, Mingya G; Kuriakose, Saritha V; Skottke, Kyle; Repetti, Peter P; Reuber, T Lynne; Ruff, Thomas G; Petracek, Marie E; Loida, Paul J

2014-01-01

ATHB17 (AT2G01430) is an Arabidopsis gene encoding a member of the α-subclass of the homeodomain leucine zipper class II (HD-Zip II) family of transcription factors. The ATHB17 monomer contains four domains common to all class II HD-Zip proteins: a putative repression domain adjacent to a homeodomain, leucine zipper, and carboxy terminal domain. However, it also possesses a unique N-terminus not present in other members of the family. In this study we demonstrate that the unique 73 amino acid N-terminus is involved in regulation of cellular localization of ATHB17. The ATHB17 protein is shown to function as a transcriptional repressor and an EAR-like motif is identified within the putative repression domain of ATHB17. Transformation of maize with an ATHB17 expression construct leads to the expression of ATHB17Δ113, a truncated protein lacking the first 113 amino acids which encodes a significant portion of the repression domain. Because ATHB17Δ113 lacks the repression domain, the protein cannot directly affect the transcription of its target genes. ATHB17Δ113 can homodimerize, form heterodimers with maize endogenous HD-Zip II proteins, and bind to target DNA sequences; thus, ATHB17Δ113 may interfere with HD-Zip II mediated transcriptional activity via a dominant negative mechanism. We provide evidence that maize HD-Zip II proteins function as transcriptional repressors and that ATHB17Δ113 relieves this HD-Zip II mediated transcriptional repression activity. Expression of ATHB17Δ113 in maize leads to increased ear size at silking and, therefore, may enhance sink potential. We hypothesize that this phenotype could be a result of modulation of endogenous HD-Zip II pathways in maize.

Expression of a Truncated ATHB17 Protein in Maize Increases Ear Weight at Silking

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Creelman, Robert A.; Griffith, Cara; Ahrens, Jeffrey E.; Taylor, J. Philip; Murphy, Lesley R.; Manjunath, Siva; Thompson, Rebecca L.; Lingard, Matthew J.; Back, Stephanie L.; Larue, Huachun; Brayton, Bonnie R.; Burek, Amanda J.; Tiwari, Shiv; Adam, Luc; Morrell, James A.; Caldo, Rico A.; Huai, Qing; Kouadio, Jean-Louis K.; Kuehn, Rosemarie; Sant, Anagha M.; Wingbermuehle, William J.; Sala, Rodrigo; Foster, Matt; Kinser, Josh D.; Mohanty, Radha; Jiang, Dongming; Ziegler, Todd E.; Huang, Mingya G.; Kuriakose, Saritha V.; Skottke, Kyle; Repetti, Peter P.; Reuber, T. Lynne; Ruff, Thomas G.; Petracek, Marie E.; Loida, Paul J.

2014-01-01

ATHB17 (AT2G01430) is an Arabidopsis gene encoding a member of the α-subclass of the homeodomain leucine zipper class II (HD-Zip II) family of transcription factors. The ATHB17 monomer contains four domains common to all class II HD-Zip proteins: a putative repression domain adjacent to a homeodomain, leucine zipper, and carboxy terminal domain. However, it also possesses a unique N-terminus not present in other members of the family. In this study we demonstrate that the unique 73 amino acid N-terminus is involved in regulation of cellular localization of ATHB17. The ATHB17 protein is shown to function as a transcriptional repressor and an EAR-like motif is identified within the putative repression domain of ATHB17. Transformation of maize with an ATHB17 expression construct leads to the expression of ATHB17Δ113, a truncated protein lacking the first 113 amino acids which encodes a significant portion of the repression domain. Because ATHB17Δ113 lacks the repression domain, the protein cannot directly affect the transcription of its target genes. ATHB17Δ113 can homodimerize, form heterodimers with maize endogenous HD-Zip II proteins, and bind to target DNA sequences; thus, ATHB17Δ113 may interfere with HD-Zip II mediated transcriptional activity via a dominant negative mechanism. We provide evidence that maize HD-Zip II proteins function as transcriptional repressors and that ATHB17Δ113 relieves this HD-Zip II mediated transcriptional repression activity. Expression of ATHB17Δ113 in maize leads to increased ear size at silking and, therefore, may enhance sink potential. We hypothesize that this phenotype could be a result of modulation of endogenous HD-Zip II pathways in maize. PMID:24736658

Delivery of kinesin spindle protein targeting siRNA in solid lipid nanoparticles to cellular models of tumor vasculature

International Nuclear Information System (INIS)

Ying, Bo; Campbell, Robert B.

2014-01-01

Highlights: • siRNA-lipid nanoparticles are solid particles not lipid bilayers with aqueous core. • High, but not low, PEG content can prevent nanoparticle encapsulation of siRNA. • PEG reduces cellular toxicity of cationic nanoparticles in vitro. • PEG reduces zeta potential while improving gene silencing of siRNA nanoparticles. • Kinesin spindle protein can be an effective target for tumor vascular targeting. - Abstract: The ideal siRNA delivery system should selectively deliver the construct to the target cell, avoid enzymatic degradation, and evade uptake by phagocytes. In the present study, we evaluated the importance of polyethylene glycol (PEG) on lipid-based carrier systems for encapsulating, and delivering, siRNA to tumor vessels using cellular models. Lipid nanoparticles containing different percentage of PEG were evaluated based on their physical chemical properties, density compared to water, siRNA encapsulation, toxicity, targeting efficiency and gene silencing in vitro. siRNA can be efficiently loaded into lipid nanoparticles (LNPs) when DOTAP is included in the formulation mixture. However, the total amount encapsulated decreased with increase in PEG content. In the presence of siRNA, the final formulations contained a mixed population of particles based on density. The major population which contains the majority of siRNA exhibited a density of 4% glucose, and the minor fraction associated with a decreased amount of siRNA had a density less than PBS. The inclusion of 10 mol% PEG resulted in a greater amount of siRNA associated with the minor fraction. Finally, when kinesin spindle protein (KSP) siRNA was encapsulated in lipid nanoparticles containing a modest amount of PEG, the proliferation of endothelial cells was inhibited due to the efficient knock down of KSP mRNA. The presence of siRNA resulted in the formation of solid lipid nanoparticles when prepared using the thin film and hydration method. LNPs with a relatively modest amount of

Goldberg-Shprintzen megacolon syndrome with associated sensory motor axonal neuropathy.

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Dafsari, Hormos Salimi; Byrne, Susan; Lin, Jean-Pierre; Pitt, Matthew; Jongbloed, Jan Dh; Flinter, Frances; Jungbluth, Heinz

2015-06-01

Goldberg-Shprintzen megacolon syndrome (GOSHS) (OMIM 609460) is characterized by a combination of learning difficulties, characteristic dysmorphic features and Hirschsprung's disease. Variable clinical features include iris coloboma, congenital heart defects and central nervous system abnormalities, in particular polymicrogyria. GOSHS has been attributed to recessive mutations in KIAA1279, encoding kinesin family member (KIF)-binding protein (KBP) with a crucial role in neuronal microtubule dynamics. Here we report on a 7-year-old girl with GOSHS as a result of a homozygous deletion of exons 5 and 6 of the KIAA1279 gene. She had been referred with the suspicion of an underlying neuromuscular disorder before the genetic diagnosis was established, prompted by the findings of motor developmental delay, hypotonia, ptosis and absent reflexes. Neurophysiological studies revealed unequivocal evidence of a peripheral axonal sensory motor neuropathy. We hypothesize that an axonal sensory motor neuropathy may be part of the phenotypical spectrum of KIAA1279-related GOSHS, probably reflecting the effects of reduced KBP protein expression on peripheral neuronal function. © 2015 Wiley Periodicals, Inc.

Anomalous inhibition of c-Met by the kinesin inhibitor aurintricarboxylic acid.

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Milanovic, Mina; Radtke, Simone; Peel, Nick; Howell, Michael; Carrière, Virginie; Joffre, Carine; Kermorgant, Stéphanie; Parker, Peter J

2012-03-01

c-Met [the hepatocyte growth factor (HGF) receptor] is a receptor tyrosine kinase playing a role in various biological events. Overexpression of the receptor has been observed in a number of cancers, correlating with increased metastatic tendency and poor prognosis. Additionally, activating mutations in c-Met kinase domain have been reported in a subset of familial cancers causing resistance to treatment. Receptor trafficking, relying on the integrity of the microtubule network, plays an important role in activation of downstream targets and initiation of signalling events. Aurintricarboxylic acid (ATA) is a triphenylmethane derivative that has been reported to inhibit microtubule motor proteins kinesins. Additional reported properties of this inhibitor include inhibition of protein tyrosine phosphatases, nucleases and members of the Jak family. Here we demonstrate that ATA prevents HGF-induced c-Met phosphorylation, internalisation, subsequent receptor trafficking and degradation. In addition, ATA prevented HGF-induced downstream signalling which also affected cellular function, as assayed by collective cell migration of A549 cells. Surprisingly, the inhibitory effect of ATA on HGF-induced phosphorylation and signalling in vivo was associated with an increase in basal c-Met kinase activity in vitro. It is concluded that the inhibitory effects of ATA on c-Met in vivo is an allosteric effect mediated through the kinase domain of the receptor. As the currently tested adenosine triphosphate competitive tyrosine kinase inhibitors (TKIs) may lead to tumor resistance (McDermott U, et al., Cancer Res 2010;70:1625-34), our findings suggest that novel anti-c-Met therapies could be developed in the future for cancer treatment. Copyright © 2011 UICC.

S. pombe kinesins-8 promote both nucleation and catastrophe of microtubules.

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Muriel Erent

Full Text Available The kinesins-8 were originally thought to be microtubule depolymerases, but are now emerging as more versatile catalysts of microtubule dynamics. We show here that S. pombe Klp5-436 and Klp6-440 are non-processive plus-end-directed motors whose in vitro velocities on S. pombe microtubules at 7 and 23 nm s(-1 are too slow to keep pace with the growing tips of dynamic interphase microtubules in living S. pombe. In vitro, Klp5 and 6 dimers exhibit a hitherto-undescribed combination of strong enhancement of microtubule nucleation with no effect on growth rate or catastrophe frequency. By contrast in vivo, both Klp5 and Klp6 promote microtubule catastrophe at cell ends whilst Klp6 also increases the number of interphase microtubule arrays (IMAs. Our data support a model in which Klp5/6 bind tightly to free tubulin heterodimers, strongly promoting the nucleation of new microtubules, and then continue to land as a tubulin-motor complex on the tips of growing microtubules, with the motors then dissociating after a few seconds residence on the lattice. In vivo, we predict that only at cell ends, when growing microtubule tips become lodged and their growth slows down, will Klp5/6 motor activity succeed in tracking growing microtubule tips. This mechanism would allow Klp5/6 to detect the arrival of microtubule tips at cells ends and to amplify the intrinsic tendency for microtubules to catastrophise in compression at cell ends. Our evidence identifies Klp5 and 6 as spatial regulators of microtubule dynamics that enhance both microtubule nucleation at the cell centre and microtubule catastrophe at the cell ends.

Protein Interaction Analysis Provides a Map of the Spatial and Temporal Organization of the Ciliary Gating Zone.

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Takao, Daisuke; Wang, Liang; Boss, Allison; Verhey, Kristen J

2017-08-07

The motility and signaling functions of the primary cilium require a unique protein and lipid composition that is determined by gating mechanisms localized at the base of the cilium. Several protein complexes localize to the gating zone and may regulate ciliary protein composition; however, the mechanisms of ciliary gating and the dynamics of the gating components are largely unknown. Here, we used the BiFC (bimolecular fluorescence complementation) assay and report for the first time on the protein-protein interactions that occur between ciliary gating components and transiting cargoes during ciliary entry. We find that the nucleoporin Nup62 and the C termini of the nephronophthisis (NPHP) proteins NPHP4 and NPHP5 interact with the axoneme-associated kinesin-2 motor KIF17 and thus spatially map to the inner region of the ciliary gating zone. Nup62 and NPHP4 exhibit rapid turnover at the transition zone and thus define dynamic components of the gate. We find that B9D1, AHI1, and the N termini of NPHP4 and NPHP5 interact with the transmembrane protein SSTR3 and thus spatially map to the outer region of the ciliary gating zone. B9D1, AHI1, and NPHP5 exhibit little to no turnover at the transition zone and thus define components of a stable gating structure. These data provide the first comprehensive map of the molecular orientations of gating zone components along the inner-to-outer axis of the ciliary gating zone. These results advance our understanding of the functional roles of gating zone components in regulating ciliary protein composition. Copyright © 2017 Elsevier Ltd. All rights reserved.

Kinesin Khc-73/KIF13B modulates retrograde BMP signaling by influencing endosomal dynamics at the Drosophila neuromuscular junction.

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Liao, Edward H; Gray, Lindsay; Tsurudome, Kazuya; El-Mounzer, Wassim; Elazzouzi, Fatima; Baim, Christopher; Farzin, Sarah; Calderon, Mario R; Kauwe, Grant; Haghighi, A Pejmun

2018-01-01

Retrograde signaling is essential for neuronal growth, function and survival; however, we know little about how signaling endosomes might be directed from synaptic terminals onto retrograde axonal pathways. We have identified Khc-73, a plus-end directed microtubule motor protein, as a regulator of sorting of endosomes in Drosophila larval motor neurons. The number of synaptic boutons and the amount of neurotransmitter release at the Khc-73 mutant larval neuromuscular junction (NMJ) are normal, but we find a significant decrease in the number of presynaptic release sites. This defect in Khc-73 mutant larvae can be genetically enhanced by a partial genetic loss of Bone Morphogenic Protein (BMP) signaling or suppressed by activation of BMP signaling in motoneurons. Consistently, activation of BMP signaling that normally enhances the accumulation of phosphorylated form of BMP transcription factor Mad in the nuclei, can be suppressed by genetic removal of Khc-73. Using a number of assays including live imaging in larval motor neurons, we show that loss of Khc-73 curbs the ability of retrograde-bound endosomes to leave the synaptic area and join the retrograde axonal pathway. Our findings identify Khc-73 as a regulator of endosomal traffic at the synapse and modulator of retrograde BMP signaling in motoneurons.

Orientations of Iron-Sulfur Clusters FA and FB in the Homodimeric Type-I Photosynthetic Reaction Center of Heliobacterium modesticaldum.

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Kondo, Toru; Matsuoka, Masahiro; Azai, Chihiro; Itoh, Shigeru; Oh-Oka, Hirozo

2016-05-12

Orientations of the FA and FB iron-sulfur (FeS) clusters in a structure-unknown type-I homodimeric heriobacterial reaction center (hRC) were studied in oriented membranes of the thermophilic anaerobic photosynthetic bacterium Heliobacterium modesticaldum by electron paramagnetic resonance (EPR), and compared with those in heterodimeric photosystem I (PS I). The Rieske-type FeS center in the cytochrome b/c complex showed a well-oriented EPR signal. Illumination at 14 K induced an FB(-) signal with g-axes of gz = 2.066, gy = 1.937, and gx = 1.890, tilted at angles of 60°, 60°, and 45°, respectively, with respect to the membrane normal. Chemical reduction with dithionite produced an additional signal of FA(-), which magnetically interacted with FB(-), with gz = 2.046, gy = 1.942, and gx = 1.911 at 30°, 60°, and 90°, respectively. The angles and redox properties of FA(-) and FB(-) in hRC resemble those of FB(-) and FA(-), respectively, in PS I. Therefore, FA and FB in hRC, named after their g-value similarities, seem to be located like FB and FA, not like FA and FB, respectively, in PS I. The reducing side of hRC could resemble those in PS I, if the names of FA and FB are interchanged with each other.

Structural and functional characterization of a novel homodimeric three-finger neurotoxin from the venom of Ophiophagus hannah (king cobra).

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Roy, Amrita; Zhou, Xingding; Chong, Ming Zhi; D'hoedt, Dieter; Foo, Chun Shin; Rajagopalan, Nandhakishore; Nirthanan, Selvanayagam; Bertrand, Daniel; Sivaraman, J; Kini, R Manjunatha

2010-03-12

Snake venoms are a mixture of pharmacologically active proteins and polypeptides that have led to the development of molecular probes and therapeutic agents. Here, we describe the structural and functional characterization of a novel neurotoxin, haditoxin, from the venom of Ophiophagus hannah (King cobra). Haditoxin exhibited novel pharmacology with antagonism toward muscle (alphabetagammadelta) and neuronal (alpha(7), alpha(3)beta(2), and alpha(4)beta(2)) nicotinic acetylcholine receptors (nAChRs) with highest affinity for alpha(7)-nAChRs. The high resolution (1.5 A) crystal structure revealed haditoxin to be a homodimer, like kappa-neurotoxins, which target neuronal alpha(3)beta(2)- and alpha(4)beta(2)-nAChRs. Interestingly however, the monomeric subunits of haditoxin were composed of a three-finger protein fold typical of curaremimetic short-chain alpha-neurotoxins. Biochemical studies confirmed that it existed as a non-covalent dimer species in solution. Its structural similarity to short-chain alpha-neurotoxins and kappa-neurotoxins notwithstanding, haditoxin exhibited unique blockade of alpha(7)-nAChRs (IC(50) 180 nm), which is recognized by neither short-chain alpha-neurotoxins nor kappa-neurotoxins. This is the first report of a dimeric short-chain alpha-neurotoxin interacting with neuronal alpha(7)-nAChRs as well as the first homodimeric three-finger toxin to interact with muscle nAChRs.

Mutations in Human Tubulin Proximal to the Kinesin-Binding Site Alter Dynamic Instability at Microtubule Plus- and Minus-Ends

Energy Technology Data Exchange (ETDEWEB)

Ti, Shih-Chieh; Pamula, Melissa C.; Howes, Stuart C.; Duellberg, Christian; Cade, Nicholas I.; Kleiner, Ralph E.; Forth, Scott; Surrey, Thomas; Nogales, Eva; Kapoor, Tarun M.

2016-04-01

The assembly of microtubule-based cellular structures depends on regulated tubulin polymerization and directional transport. In this research, we have purified and characterized tubulin heterodimers that have human β-tubulin isotype III (TUBB3), as well as heterodimers with one of two β-tubulin mutations (D417H or R262H). Both point mutations are proximal to the kinesin-binding site and have been linked to an ocular motility disorder in humans. Compared to wild-type, microtubules with these mutations have decreased catastrophe frequencies and increased average lifetimes of plus- and minus-end-stabilizing caps. Importantly, the D417H mutation does not alter microtubule lattice structure or Mal3 binding to growing filaments. Instead, this mutation reduces the affinity of tubulin for TOG domains and colchicine, suggesting that the distribution of tubulin heterodimer conformations is changed. Together, our findings reveal how residues on the surface of microtubules, distal from the GTP-hydrolysis site and inter-subunit contacts, can alter polymerization dynamics at the plus- and minus-ends of microtubules.

Chemical and thermal modulation of molecular motor activities

Science.gov (United States)

Hong, Weili

Molecular motors of kinesin and dynein families are responsible for various intracellular activities, from long distance movement of organelles, vesicles, protein complexes, and mRNAs to powering mitotic processes. They can take nanometer steps using chemical energy from the hydrolysis of ATP (adenosine triphosphate), and their dysfunction is involved in many neurodegenerative diseases that require long distance transport of cargos. Here I report on the study of the properties of molecular motors at a single-molecule level using optical trappings. I first studied the inhibition properties of kinesin motors by marine natural compound adociasulfates. I showed that adociasulfates compete with microtubules for binding to kinesins and thus inhibit kinesins' activity. Although adociasulfates are a strong inhibitor for all kinesin members, they show a much higher inhibition effect for conventional kinesins than for mitotic kinesins. Thus adociasulfates can be used to specifically inhibit conventional kinesins. By comparing the inhibition of kinesins by two structurally similar adociasulfates, one can see that the negatively charged sulfate residue of adociasulfates can be replaced by other negative residues and thus make it possible for adociasulfate-derived compounds to be more cell permeable. Kinesins and dyneins move cargos towards opposite directions along a microtubule. Cargos with both kinesins and dyneins attached often move bidirectionally due to undergoing a tug-of-war between the oppositely moving kinesin and dynein motors. Here I studied the effect of temperature on microtubule-based kinesin and dynein motor transport. While kinesins' and dyneins' velocities are closely matched above 15 °C, below this temperature the dyneins' velocity decreases much faster than the kinesins'. The kinesins' and dyneins' forces do not measurably change with temperature. The results suggest that temperature has significant effects on bidirectional transport and can be used to

Crystal structures of the F and pSLT plasmid TraJ N-terminal regions reveal similar homodimeric PAS folds with functional interchangeability

Energy Technology Data Exchange (ETDEWEB)

Lu, Jun; Wu, Ruiying; Adkins, Joshua N.; Joachimiak, Andrzej; Glover, Mark

2014-09-16

In the F-family of conjugative plasmids, TraJ is an essential transcriptional activator of the tra operon that encodes most of the proteins required for conjugation. Here we report for the first time the X-ray crystal structures of the TraJ N-terminal regions from the prototypic F plasmid (TraJF11-130) and from the Salmonella virulence plasmid pSLT (TraJpSLT 1-128). Both proteins form similar homodimeric Per-ARNT-Sim (PAS) fold structures. Mutational analysis reveals that the observed dimeric interface is critical for TraJF transcriptional activation, indicating that dimerization of TraJ is required for its in vivo function. An artificial ligand (oxidized dithiothreitol) occupies a cavity in the TraJF dimer interface, while a smaller cavity in corresponding region of the TraJpSLT structure lacks a ligand. Gas chromatography/mass spectrometry-electron ionization analysis of dithiothreitol-free TraJF suggests indole may be the natural TraJ ligand; however, disruption of the indole biosynthetic pathway does not affect TraJF function. Heterologous PAS domains from pSLT and R100 TraJ can functionally replace the TraJF PAS domain, suggesting that TraJ allelic specificity is mediated by the region C-terminal to the PAS domain.

Negative regulation of ciliary length by ciliary male germ cell-associated kinase (Mak) is required for retinal photoreceptor survival.

Science.gov (United States)

Omori, Yoshihiro; Chaya, Taro; Katoh, Kimiko; Kajimura, Naoko; Sato, Shigeru; Muraoka, Koichiro; Ueno, Shinji; Koyasu, Toshiyuki; Kondo, Mineo; Furukawa, Takahisa

2010-12-28

Cilia function as cell sensors in many organs, and their disorders are referred to as "ciliopathies." Although ciliary components and transport machinery have been well studied, regulatory mechanisms of ciliary formation and maintenance are poorly understood. Here we show that male germ cell-associated kinase (Mak) regulates retinal photoreceptor ciliary length and subcompartmentalization. Mak was localized both in the connecting cilia and outer-segment axonemes of photoreceptor cells. In the Mak-null retina, photoreceptors exhibit elongated cilia and progressive degeneration. We observed accumulation of intraflagellar transport 88 (IFT88) and IFT57, expansion of kinesin family member 3A (Kif3a), and acetylated α-tubulin signals in the Mak-null photoreceptor cilia. We found abnormal rhodopsin accumulation in the Mak-null photoreceptor cell bodies at postnatal day 14. In addition, overexpression of retinitis pigmentosa 1 (RP1), a microtubule-associated protein localized in outer-segment axonemes, induced ciliary elongation, and Mak coexpression rescued excessive ciliary elongation by RP1. The RP1 N-terminal portion induces ciliary elongation and increased intensity of acetylated α-tubulin labeling in the cells and is phosphorylated by Mak. These results suggest that Mak is essential for the regulation of ciliary length and is required for the long-term survival of photoreceptors.

Selective synaptic targeting of the excitatory and inhibitory presynaptic organizers FGF22 and FGF7.

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Terauchi, Akiko; Timmons, Kendall M; Kikuma, Koto; Pechmann, Yvonne; Kneussel, Matthias; Umemori, Hisashi

2015-01-15

Specific formation of excitatory and inhibitory synapses is crucial for proper functioning of the brain. Fibroblast growth factor 22 (FGF22) and FGF7 are postsynaptic-cell-derived presynaptic organizers necessary for excitatory and inhibitory presynaptic differentiation, respectively, in the hippocampus. For the establishment of specific synaptic networks, these FGFs must localize to appropriate synaptic locations - FGF22 to excitatory and FGF7 to inhibitory postsynaptic sites. Here, we show that distinct motor and adaptor proteins contribute to intracellular microtubule transport of FGF22 and FGF7. Excitatory synaptic targeting of FGF22 requires the motor proteins KIF3A and KIF17 and the adaptor protein SAP102 (also known as DLG3). By contrast, inhibitory synaptic targeting of FGF7 requires the motor KIF5 and the adaptor gephyrin. Time-lapse imaging shows that FGF22 moves with SAP102, whereas FGF7 moves with gephyrin. These results reveal the basis of selective targeting of the excitatory and inhibitory presynaptic organizers that supports their different synaptogenic functions. Finally, we found that knockdown of SAP102 or PSD95 (also known as DLG4), which impairs the differentiation of excitatory synapses, alters FGF7 localization, suggesting that signals from excitatory synapses might regulate inhibitory synapse formation by controlling the distribution of the inhibitory presynaptic organizer. © 2015. Published by The Company of Biologists Ltd.

Structural differences between glycosylated, disulfide-linked heterodimeric Knob-into-Hole Fc fragment and its homodimeric Knob-Knob and Hole-Hole side products.

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Kuglstatter, A; Stihle, M; Neumann, C; Müller, C; Schaefer, W; Klein, C; Benz, J

2017-09-01

An increasing number of bispecific therapeutic antibodies are progressing through clinical development. The Knob-into-Hole (KiH) technology uses complementary mutations in the CH3 region of the antibody Fc fragment to achieve heavy chain heterodimerization. Here we describe the X-ray crystal structures of glycosylated and disulfide-engineered heterodimeric KiH Fc fragment and its homodimeric Knob-Knob and Hole-Hole side products. The heterodimer structure confirms the KiH design principle and supports the hypothesis that glycosylation stabilizes a closed Fc conformation. Both homodimer structures show parallel Fc fragment architectures, in contrast to recently reported crystal structures of the corresponding aglycosylated Fc fragments which in the absence of disulfide mutations show an unexpected antiparallel arrangement. The glycosylated Knob-Knob Fc fragment is destabilized as indicated by variability in the relative orientation of its CH3 domains. The glycosylated Hole-Hole Fc fragment shows an unexpected intermolecular disulfide bond via the introduced Y349C Hole mutation which results in a large CH3 domain shift and a new CH3-CH3 interface. The crystal structures of glycosylated, disulfide-linked KiH Fc fragment and its Knob-Knob and Hole-Hole side products reported here will facilitate further design of highly efficient antibody heterodimerization strategies. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

PPARG, KCNJ11, CDKAL1, CDKN2A-CDKN2B, IDE-KIF11-HHEX, IGF2BP2 and SLC30A8 are associated with type 2 diabetes in a Chinese population.

Directory of Open Access Journals (Sweden)

Cheng Hu

2009-10-01

Full Text Available Recent advance in genetic studies added the confirmed susceptible loci for type 2 diabetes to eighteen. In this study, we attempt to analyze the independent and joint effect of variants from these loci on type 2 diabetes and clinical phenotypes related to glucose metabolism.Twenty-one single nucleotide polymorphisms (SNPs from fourteen loci were successfully genotyped in 1,849 subjects with type 2 diabetes and 1,785 subjects with normal glucose regulation. We analyzed the allele and genotype distribution between the cases and controls of these SNPs as well as the joint effects of the susceptible loci on type 2 diabetes risk. The associations between SNPs and type 2 diabetes were examined by logistic regression. The associations between SNPs and quantitative traits were examined by linear regression. The discriminative accuracy of the prediction models was assessed by area under the receiver operating characteristic curves. We confirmed the effects of SNPs from PPARG, KCNJ11, CDKAL1, CDKN2A-CDKN2B, IDE-KIF11-HHEX, IGF2BP2 and SLC30A8 on risk for type 2 diabetes, with odds ratios ranging from 1.114 to 1.406 (P value range from 0.0335 to 1.37E-12. But no significant association was detected between SNPs from WFS1, FTO, JAZF1, TSPAN8-LGR5, THADA, ADAMTS9, NOTCH2-ADAM30 and type 2 diabetes. Analyses on the quantitative traits in the control subjects showed that THADA SNP rs7578597 was association with 2-h insulin during oral glucose tolerance tests (P = 0.0005, empirical P = 0.0090. The joint effect analysis of SNPs from eleven loci showed the individual carrying more risk alleles had a significantly higher risk for type 2 diabetes. And the type 2 diabetes patients with more risk allele tended to have earlier diagnostic ages (P = 0.0006.The current study confirmed the association between PPARG, KCNJ11, CDKAL1, CDKN2A-CDKN2B, IDE-KIF11-HHEX, IGF2BP2 and SLC30A8 and type 2 diabetes. These type 2 diabetes risk loci contributed to the disease additively.

Recombinant expression of homodimeric 660 kDa human thyroglobulin in soybean seeds: an alternative source of human thyroglobulin.

Science.gov (United States)

Powell, Rebecca; Hudson, Laura C; Lambirth, Kevin C; Luth, Diane; Wang, Kan; Bost, Kenneth L; Piller, Kenneth J

2011-07-01

Soybean seeds possess many qualities that make them ideal targets for the production of recombinant proteins. However, one quality often overlooked is their ability to stockpile large amounts of complex storage proteins. Because of this characteristic, we hypothesized that soybean seeds would support recombinant expression of large and complex proteins that are currently difficult or impossible to express using traditional plant and non-plant-based host systems. To test this hypothesis, we transformed soybeans with a synthetic gene encoding human thyroglobulin (hTG)-a 660 kDa homodimeric protein that is widely used in the diagnostic industry for screening and detection of thyroid disease. In the absence of a recombinant system that can produce recombinant hTG, research and diagnostic grade hTG continues to be purified from cadaver and surgically removed thyroid tissue. These less-than-ideal tissue sources lack uniform glycosylation and iodination and therefore introduce variability when purified hTG is used in sensitive ELISA screens. In this study, we report the successful expression of recombinant hTG in soybean seeds. Authenticity of the soy-derived protein was demonstrated using commercial ELISA kits developed specifically for the detection of hTG in patient sera. Western analyses and gel filtration chromatography demonstrated that recombinant hTG and thyroid-purified hTG are biologically similar with respect to size, mass, charge and subunit interaction. The recombinant protein was stable over three generations and accumulated to ~1.5% of total soluble seed protein. These results support our hypothesis that soybeans represent a practical alternative to traditional host systems for the expression of large and complex proteins.

Photocontrol of the mitotic kinesin Eg5 using a novel S-trityl-L-cysteine analogue as a photochromic inhibitor.

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Ishikawa, Kumiko; Tohyama, Kanako; Mitsuhashi, Shinya; Maruta, Shinsaku

2014-04-01

Because the mitotic kinesin Eg5 is essential for the formation of bipolar spindles during eukaryotic cell division, it has been considered as a potential target for cancer treatment. A number of specific and potent inhibitors of Eg5 are known. S-trityl-L-cysteine is one of the inhibitors of Eg5 whose molecular mechanism of inhibition was well studied. The trityl group of S-trityl-L-cysteine was shown to be a key moiety required for potent inhibition. In this study, we synthesized a novel photochromic S-trityl-L-cysteine analogue, 4-(N-(2-(N-acetylcysteine-S-yl) acetyl) amino)-4'- (N-(2-(N-(triphenylmethyl)amino)acetyl)amino)azobenzene (ACTAB), composed of a trityl group, azobenzene and N-acetyl-L-cysteine, which exhibits cis-trans photoisomerization in order to photocontrol the function of Eg5. ACTAB exhibited cis-trans photoisomerization upon alternating irradiation at two different wavelengths in the visible range, 400 and 480 nm. ACTAB induced reversible changes in the inhibitory activity of ATPase and motor activities correlating with the cis-trans photoisomerization. Compared with cis-ACTAB, trans-ACTAB reduced ATPase activity and microtubule gliding velocity more significantly. These results suggest that ACTAB could be used as photochromic inhibitor of Eg5 to achieve photocontrol of living cells.

The HRM practices of innovative knowledge-intensive firms

DEFF Research Database (Denmark)

Jørgensen, Frances; Becker, Karen; Matthews, Judy

2011-01-01

, is the basis of this research. The study investigates the HRM practices being used by four innovative firms: two KIFs and two traditional production companies. The findings suggest that there are some similarities between the HRM practices of the case firms, but there are also important differences......There is little research on HRM practices in knowledge-intensive firms (KIFs), especially in comparison to that conducted in traditional production environments. Whether such HRM practices are appropriate for KIFs, particularly ones that differentiate themselves on their ability to innovate...

A cAMP/PKA/Kinesin-1 Axis Promotes the Axonal Transport of Mitochondria in Aging Drosophila Neurons.

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Vagnoni, Alessio; Bullock, Simon L

2018-04-23

Mitochondria play fundamental roles within cells, including energy provision, calcium homeostasis, and the regulation of apoptosis. The transport of mitochondria by microtubule-based motors is critical for neuronal structure and function. This process allows local requirements for mitochondrial functions to be met and also facilitates recycling of these organelles [1, 2]. An age-related reduction in mitochondrial transport has been observed in neurons of mammalian and non-mammalian organisms [3-6], and has been proposed to contribute to the broader decline in neuronal function that occurs during aging [3, 5-7]. However, the factors that influence mitochondrial transport in aging neurons are poorly understood. Here we provide evidence using the tractable Drosophila wing nerve system that the cyclic AMP/protein kinase A (cAMP/PKA) pathway promotes the axonal transport of mitochondria in adult neurons. The level of the catalytic subunit of PKA decreases during aging, and acute activation of the cAMP/PKA pathway in aged flies strongly stimulates mitochondrial motility. Thus, the age-related impairment of transport is reversible. The expression of many genes is increased by PKA activation in aged flies. However, our results indicate that elevated mitochondrial transport is due in part to upregulation of the heavy chain of the kinesin-1 motor, the level of which declines during aging. Our study identifies evolutionarily conserved factors that can strongly influence mitochondrial motility in aging neurons. Copyright © 2018 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Discovery of (+)-N-(3-aminopropyl)-N-[1-(5-benzyl-3-methyl-4-oxo-[1,2]thiazolo[5,4-d]pyrimidin-6-yl)-2-methylpropyl]-4-methylbenzamide (AZD4877), a kinesin spindle protein inhibitor and potential anticancer agent.

Science.gov (United States)

Theoclitou, Maria-Elena; Aquila, Brian; Block, Michael H; Brassil, Patrick J; Castriotta, Lillian; Code, Erin; Collins, Michael P; Davies, Audrey M; Deegan, Tracy; Ezhuthachan, Jayachandran; Filla, Sandra; Freed, Ellen; Hu, Haiqing; Huszar, Dennis; Jayaraman, Muthusamy; Lawson, Deborah; Lewis, Paula M; Nadella, Murali V P; Oza, Vibha; Padmanilayam, Maniyan; Pontz, Timothy; Ronco, Lucienne; Russell, Daniel; Whitston, David; Zheng, Xiaolan

2011-10-13

Structure-activity relationship analysis identified (+)-N-(3-aminopropyl)-N-[1-(5-benzyl-3-methyl-4-oxo-[1,2]thiazolo[5,4-d]pyrimidin-6-yl)-2-methylpropyl]-4-methylbenzamide (AZD4877), from a series of novel kinesin spindle protein (KSP) inhibitors, as exhibiting both excellent biochemical potency and pharmaceutical properties suitable for clinical development. The selected compound arrested cells in mitosis leading to the formation of the monopolar spindle phenotype characteristic of KSP inhibition and induction of cellular death. A favorable pharmacokinetic profile and notable in vivo efficacy supported the selection of this compound as a clinical candidate for the treatment of cancer.

The kinesin–tubulin complex: considerations in structural and functional complexity

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Olmsted ZT

2015-02-01

Full Text Available Zachary T Olmsted, Andrew G Colliver, Janet L Paluh State University of New York Polytechnic Institute, Colleges of Nanoscale Science and Engineering, College of Nanoscale Science, Nanobioscience Constellation, Albany, NY, USA Abstract: The ability of cells to respond to external cues by appropriately manipulating their internal environment requires a dynamic microtubule cytoskeleton that is facilitated by associated kinesin motor interactions. The evolutionary adaptations of kinesins and tubulins when merged generate a highly adaptable communication and infrastructure cellular network that is important to understanding specialized cell functions, human disease, and disease therapies. Here, we review the state of the field in the complex relationship of kinesin–tubulin interactions. We propose 12 mechanistic specializations of kinesins. In one category, referred to as sortability, we describe how kinesin interactions with tubulin isoforms, isotypes, or posttranslationally modified tubulins contribute to diverse cellular roles. Fourteen kinesin families have previously been described. Here, we illustrate the great depth of functional complexity that is possible in members within a single kinesin family by mechanistic specialization through discussion of the well-studied Kinesin-14 family. This includes new roles of Kinesin-14 in regulating supramolecular structures such as the microtubule-organizing center γ-tubulin ring complex of centrosomes. We next explore the value of an improved mechanistic understanding of kinesin–tubulin interactions in regard to human development, disease mechanisms, and improving treatments that target kinesin–tubulin complexes. The ability to combine the current kinesin nomenclature along with a more precisely defined kinesin and tubulin molecular toolbox is needed to support more detailed exploration of kinesin–tubulin interaction mechanisms including functional uniqueness, redundancy, or adaptations to new

Pushing the size limit of de novo structure ensemble prediction guided by sparse SDSL-EPR restraints to 200 residues: The monomeric and homodimeric forms of BAX

Science.gov (United States)

Fischer, Axel W.; Bordignon, Enrica; Bleicken, Stephanie; García-Sáez, Ana J.; Jeschke, Gunnar; Meiler, Jens

2016-01-01

Structure determination remains a challenge for many biologically important proteins. In particular, proteins that adopt multiple conformations often evade crystallization in all biologically relevant states. Although computational de novo protein folding approaches often sample biologically relevant conformations, the selection of the most accurate model for different functional states remains a formidable challenge, in particular, for proteins with more than about 150 residues. Electron paramagnetic resonance (EPR) spectroscopy can obtain limited structural information for proteins in well-defined biological states and thereby assist in selecting biologically relevant conformations. The present study demonstrates that de novo folding methods are able to accurately sample the folds of 192-residue long soluble monomeric Bcl-2-associated X protein (BAX). The tertiary structures of the monomeric and homodimeric forms of BAX were predicted using the primary structure as well as 25 and 11 EPR distance restraints, respectively. The predicted models were subsequently compared to respective NMR/X-ray structures of BAX. EPR restraints improve the protein-size normalized root-mean-square-deviation (RMSD100) of the most accurate models with respect to the NMR/crystal structure from 5.9 Å to 3.9 Å and from 5.7 Å to 3.3 Å, respectively. Additionally, the model discrimination is improved, which is demonstrated by an improvement of the enrichment from 5% to 15% and from 13% to 21%, respectively. PMID:27129417

Singlet-triplet splittings from the virial theorem and single-particle excitation energies

Science.gov (United States)

Becke, Axel D.

2018-01-01

The zeroth-order (uncorrelated) singlet-triplet energy difference in single-particle excited configurations is 2Kif, where Kif is the Coulomb self-energy of the product of the transition orbitals. Here we present a non-empirical, virial-theorem argument that the correlated singlet-triplet energy difference should be half of this, namely, Kif. This incredibly simple result gives vertical HOMO-LUMO excitation energies in small-molecule benchmarks as good as the popular TD-B3LYP time-dependent approach to excited states. For linear acenes and nonlinear polycyclic aromatic hydrocarbons, the performance is significantly better than TD-B3LYP. In addition to the virial theorem, the derivation borrows intuitive pair-density concepts from density-functional theory.

Cell Signaling and Differential Protein Expression in Neuronal Differentiation of Bone Marrow Mesenchymal Stem Cells with Hypermethylated Salvador/Warts/Hippo (SWH Pathway Genes.

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Hui-Hung Tzeng

Full Text Available Human mesenchymal stem cells (MSCs modified by targeting DNA hypermethylation of genes in the Salvador/Warts/Hippo pathway were induced to differentiate into neuronal cells in vitro. The differentiated cells secreted a significant level of brain-derived neurotrophy factor (BDNF and the expression of BDNF receptor tyrosine receptor kinase B (TrkB correlated well with the secretion of BDNF. In the differentiating cells, CREB was active after the binding of growth factors to induce phosphorylation of ERK in the MAPK/ERK pathway. Downstream of phosphorylated CREB led to the functional maturation of differentiated cells and secretion of BDNF, which contributed to the sustained expression of pERK and pCREB. In summary, both PI3K/Akt and MAPK/ERK signaling pathways play important roles in the neuronal differentiation of MSCs. The main function of the PI3K/Akt pathway is to maintain cell survival during neural differentiation; whereas the role of the MAPK/ERK pathway is probably to promote the maturation of differentiated MSCs. Further, cellular levels of protein kinase C epsilon type (PKC-ε and kinesin heavy chain (KIF5B increased with time of induction, whereas the level of NME/NM23 nucleoside diphosphate kinase 1 (Nm23-H1 decreased during the time course of differentiation. The correlation between PKC-ε and TrkB suggested that there is cross-talk between PKC-ε and the PI3K/Akt signaling pathway.

The KAC family of kinesin-like proteins is essential for the association of chloroplasts with the plasma membrane in land plants.

Science.gov (United States)

Suetsugu, Noriyuki; Sato, Yoshikatsu; Tsuboi, Hidenori; Kasahara, Masahiro; Imaizumi, Takato; Kagawa, Takatoshi; Hiwatashi, Yuji; Hasebe, Mitsuyasu; Wada, Masamitsu

2012-11-01

Chloroplasts require association with the plasma membrane for movement in response to light and for appropriate positioning within the cell to capture photosynthetic light efficiently. In Arabidopsis, CHLOROPLAST UNUSUAL POSITIONING 1 (CHUP1), KINESIN-LIKE PROTEIN FOR ACTIN-BASED CHLOROPLAST MOVEMENT 1 (KAC1) and KAC2 are required for both the proper movement of chloroplasts and the association of chloroplasts with the plasma membrane, through the reorganization of short actin filaments located on the periphery of the chloroplasts. Here, we show that KAC and CHUP1 orthologs (AcKAC1, AcCHUP1A and AcCHUP1B, and PpKAC1 and PpKAC2) play important roles in chloroplast positioning in the fern Adiantum capillus-veneris and the moss Physcomitrella patens. The knockdown of AcKAC1 and two AcCHUP1 genes induced the aggregation of chloroplasts around the nucleus. Analyses of A. capillus-veneris mutants containing perinuclear-aggregated chloroplasts confirmed that AcKAC1 is required for chloroplast-plasma membrane association. In addition, P. patens lines in which two KAC genes had been knocked out showed an aggregated chloroplast phenotype similar to that of the fern kac1 mutants. These results indicate that chloroplast positioning and movement are mediated through the activities of KAC and CHUP1 proteins, which are conserved in land plants.

Novel Serum Proteomic Signatures in a Non-Human Primate Model of Retinal Injury

Science.gov (United States)

2011-01-01

public release; distribution unlimited 13. SUPPLEMENTARY NOTES 14. ABSTRACT 15. SUBJECT TERMS 16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF...OS=Macaca mulatta GN=TMEM57 PE=2 SV=1 DB=sp 11 17 3 4 0.860319 0.768643 Q6UIN1 Protein kinase C iota (Fragment) OS=Macaca mulatta GN=PRKCI PE=2 SV=1...06 Q6UIN1 Protein kinase C iota (Fragment) OS=Macaca mulatta GN=PRKCI PE=2 SV=1 DB=tr 7 26 6 3 0.300602 0.002578 Q3YAL2 Kinesin family member 27

Hsp90 inhibitor 17-AAG reduces ErbB2 levels and inhibits proliferation of the trastuzumab resistant breast tumor cell line JIMT-1.

Science.gov (United States)

Zsebik, Barbara; Citri, Ami; Isola, Jorma; Yarden, Yosef; Szöllosi, János; Vereb, György

2006-04-15

ErbB2, a member of the EGF receptor family of tyrosine kinases is overexpressed on many tumor cells of epithelial origin and is the molecular target of trastuzumab (Herceptin), the first humanized antibody used in the therapy of solid tumors. Trastuzumab, which is thought to act, at least in part, by downregulating ErbB2 expression is only effective in approximately 30-40% of ErbB2 positive breast tumors. Geldanamycin and its derivative 17-AAG are potential antitumor agents capable of downregulating client proteins of Hsp90, including ErbB2. To investigate the ability of 17-AAG to downregulate ErbB2 in trastuzumab resistant breast cancer cells and the possibility of 17-AAG and trastuzumab potentiating each other's effect, the recently established trastuzumab resistant breast cancer cell line, JIMT-1 was compared to the known trastuzumab sensitive SKBR-3 line. Baseline and stimulus-evoked dimerization and activation levels of ErbB2, and the effects of trastuzumab and 17-AAG alone and in combination on cell proliferation and apoptosis, as well as on ErbB2 expression and phosphorylation have been measured. Baseline activation and amenability to activation and downregulation by trastuzumab was much lower in the resistant line. However, 17-AAG enhanced ErbB2 homodimerization after 5-10 min of treatment in both cell lines, and decreased proliferation with an IC50 of 70 nM for SKBR-3 and 10nM for JIMT-1. Thus, 17-AAG may be a useful drug in trastuzumab resistant ErbB2 overexpressing tumors. The antiproliferative effect of 17-AAG was positively correlated with phosphorylation and downregulation of ErbB2 and was dominated by apoptosis, although, especially at higher doses, necrosis was also present. Interestingly, IC50 values for ErbB2 downregulation and phosphorylation, in the 30-40 nM range, were not significantly different for the two cell lines. This observation and the negative correlation between resting ErbB2 levels and the antiproliferative effect of 17-AAG may

Cooperative protein structural dynamics of homodimeric hemoglobin linked to water cluster at subunit interface revealed by time-resolved X-ray solution scattering

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Jong Goo Kim

2016-03-01

Full Text Available Homodimeric hemoglobin (HbI consisting of two subunits is a good model system for investigating the allosteric structural transition as it exhibits cooperativity in ligand binding. In this work, as an effort to extend our previous study on wild-type and F97Y mutant HbI, we investigate structural dynamics of a mutant HbI in solution to examine the role of well-organized interfacial water cluster, which has been known to mediate intersubunit communication in HbI. In the T72V mutant of HbI, the interfacial water cluster in the T state is perturbed due to the lack of Thr72, resulting in two less interfacial water molecules than in wild-type HbI. By performing picosecond time-resolved X-ray solution scattering experiment and kinetic analysis on the T72V mutant, we identify three structurally distinct intermediates (I1, I2, and I3 and show that the kinetics of the T72V mutant are well described by the same kinetic model used for wild-type and F97Y HbI, which involves biphasic kinetics, geminate recombination, and bimolecular CO recombination. The optimized kinetic model shows that the R-T transition and bimolecular CO recombination are faster in the T72V mutant than in the wild type. From structural analysis using species-associated difference scattering curves for the intermediates, we find that the T-like deoxy I3 intermediate in solution has a different structure from deoxy HbI in crystal. In addition, we extract detailed structural parameters of the intermediates such as E-F distance, intersubunit rotation angle, and heme-heme distance. By comparing the structures of protein intermediates in wild-type HbI and the T72V mutant, we reveal how the perturbation in the interfacial water cluster affects the kinetics and structures of reaction intermediates of HbI.

47 CFR 17.17 - Existing structures.

Science.gov (United States)

2010-10-01

... 47 Telecommunication 1 2010-10-01 2010-10-01 false Existing structures. 17.17 Section 17.17... STRUCTURES Federal Aviation Administration Notification Criteria § 17.17 Existing structures. (a) The requirements found in § 17.23 relating to painting and lighting of antenna structures shall not apply to those...

Purification and properties of the glutathione S-transferases from the anoxia-tolerant turtle, Trachemys scripta elegans.

Science.gov (United States)

Willmore, William G; Storey, Kenneth B

2005-07-01

Glutathione S-transferases (GSTs) play critical roles in detoxification, response to oxidative stress, regeneration of S-thiolated proteins, and catalysis of reactions in nondetoxification metabolic pathways. Liver GSTs were purified from the anoxia-tolerant turtle, Trachemys scripta elegans. Purification separated a homodimeric (subunit relative molecular mass =34 kDa) and a heterodimeric (subunit relative molecular mass = 32.6 and 36.8 kDa) form of GST. The enzymes were purified 23-69-fold and 156-174-fold for homodimeric and heterodimeric GSTs, respectively. Kinetic data gathered using a variety of substrates and inhibitors suggested that both homodimeric and heterodimeric GSTs were of the alpha class although they showed significant differences in substrate affinities and responses to inhibitors. For example, homodimeric GST showed activity with known alpha class substrates, cumene hydroperoxide and p-nitrobenzylchloride, whereas heterodimeric GST showed no activity with cumene hydroperoxide. The specific activity of liver GSTs with chlorodinitrobenzene (CDNB) as the substrate was reduced by 2.6- and 8.7-fold for homodimeric and heterodimeric GSTs isolated from liver of anoxic turtles as compared with aerobic controls, suggesting an anoxia-responsive stable modification of the protein that may alter its function during natural anaerobiosis.

Engineering intracellular active transport systems as in vivo biomolecular tools.

Energy Technology Data Exchange (ETDEWEB)

Bachand, George David; Carroll-Portillo, Amanda

2006-11-01

Active transport systems provide essential functions in terms of cell physiology and metastasis. These systems, however, are also co-opted by invading viruses, enabling directed transport of the virus to and from the cell's nucleus (i.e., the site of virus replication). Based on this concept, fundamentally new approaches for interrogating and manipulating the inner workings of living cells may be achievable by co-opting Nature's active transport systems as an in vivo biomolecular tool. The overall goal of this project was to investigate the ability to engineer kinesin-based transport systems for in vivo applications, specifically the collection of effector proteins (e.g., transcriptional regulators) within single cells. In the first part of this project, a chimeric fusion protein consisting of kinesin and a single chain variable fragment (scFv) of an antibody was successfully produced through a recombinant expression system. The kinesin-scFv retained both catalytic and antigenic functionality, enabling selective capture and transport of target antigens. The incorporation of a rabbit IgG-specific scFv into the kinesin established a generalized system for functionalizing kinesin with a wide range of target-selective antibodies raised in rabbits. The second objective was to develop methods of isolating the intact microtubule network from live cells as a platform for evaluating kinesin-based transport within the cytoskeletal architecture of a cell. Successful isolation of intact microtubule networks from two distinct cell types was demonstrated using glutaraldehyde and methanol fixation methods. This work provides a platform for inferring the ability of kinesin-scFv to function in vivo, and may also serve as a three-dimensional scaffold for evaluating and exploiting kinesin-based transport for nanotechnological applications. Overall, the technology developed in this project represents a first-step in engineering active transport system for in vivo

Plus- and minus-end directed microtubule motors bind simultaneously to herpes simplex virus capsids using different inner tegument structures.

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Kerstin Radtke

2010-07-01

Full Text Available Many viruses depend on host microtubule motors to reach their destined intracellular location. Viral particles of neurotropic alphaherpesviruses such as herpes simplex virus 1 (HSV1 show bidirectional transport towards the cell center as well as the periphery, indicating that they utilize microtubule motors of opposing directionality. To understand the mechanisms of specific motor recruitment, it is necessary to characterize the molecular composition of such motile viral structures. We have generated HSV1 capsids with different surface features without impairing their overall architecture, and show that in a mammalian cell-free system the microtubule motors dynein and kinesin-1 and the dynein cofactor dynactin could interact directly with capsids independent of other host factors. The capsid composition and surface was analyzed with respect to 23 structural proteins that are potentially exposed to the cytosol during virus assembly or cell entry. Many of these proteins belong to the tegument, the hallmark of all herpesviruses located between the capsid and the viral envelope. Using immunoblots, quantitative mass spectrometry and quantitative immunoelectron microscopy, we show that capsids exposing inner tegument proteins such as pUS3, pUL36, pUL37, ICP0, pUL14, pUL16, and pUL21 recruited dynein, dynactin, kinesin-1 and kinesin-2. In contrast, neither untegumented capsids exposing VP5, VP26, pUL17 and pUL25 nor capsids covered by outer tegument proteins such as vhs, pUL11, ICP4, ICP34.5, VP11/12, VP13/14, VP16, VP22 or pUS11 bound microtubule motors. Our data suggest that HSV1 uses different structural features of the inner tegument to recruit dynein or kinesin-1. Individual capsids simultaneously accommodated motors of opposing directionality as well as several copies of the same motor. Thus, these associated motors either engage in a tug-of-war or their activities are coordinately regulated to achieve net transport either to the nucleus during

Arabidopsis CDS blastp result: AK059635 [KOME

Lifescience Database Archive (English)

Full Text Available AK059635 001-031-A08 At4g14330.1 phragmoplast-associated kinesin-related protein 2 ...(PAKRP2) identical to cDNA phragmoplast-associated kinesin-related protein 2 (PAKRP2) GI:16973450 6e-18 ...

Clinical value of Xenopus kinesin-like protein 2 as a prognostic marker in patients with digestive system cancers: a systematic review and meta-analysis.

Science.gov (United States)

Wang, Gang; Wang, Qian; Li, Zhengyan; Liu, Chaoxu; He, Xianli

2018-01-01

Xenopus kinesin-like protein 2 (TPX2) is a microtubule-associated protein that plays an important role in spindle assembly and dynamics. However, the clinical and prognostic value of TPX2 in the digestive system cancers remains unclear. The objective of this review was to evaluate the association of TPX2 expression with disease-free survival (DFS), overall survival (OS), and clinicopathological features of digestive system cancers. The software Stata 12.0 was used to analyze the outcomes, including OS, disease-free survival (DFS), and clinicopathological characteristics. A total of 10 eligible studies with 906 patients were included. Elevated TPX2 expression was significantly associated with poor DFS (pooled hazard ratio [HR] =2.48, 95% confidence interval [CI]: 1.96-3.13) and OS (pooled HR =2.66, 95% CI: 2.04-3.48) of digestive system malignancies. Subgroup analyses showed that cancer type, sample size, study quality, and laboratory detection methods did not alter the significant prognostic value of TPX2. Additionally, TPX2 expression was found to be an independent predictive factor for DFS (HR =2.31, 95% CI: 1.78-3.01). TPX2 expression might be associated with TNM stage and pathological grade in digestive system cancer. In conclusion, TPX2 is an independent prognostic factor for survival of patients with digestive system cancer. Furthermore, its overexpression is associated with TNM stage and pathological grade in digestive system cancer.

Arabidopsis CDS blastp result: AK242767 [KOME

Lifescience Database Archive (English)

Full Text Available AK242767 J090053D16 At4g14330.1 68417.m02207 phragmoplast-associated kinesin-relate...d protein 2 (PAKRP2) identical to cDNA phragmoplast-associated kinesin-related protein 2 (PAKRP2) GI:16973450 1e-20 ...

Arabidopsis CDS blastp result: AK242756 [KOME

Lifescience Database Archive (English)

Full Text Available AK242756 J090051D19 At4g14330.1 68417.m02207 phragmoplast-associated kinesin-relate...d protein 2 (PAKRP2) identical to cDNA phragmoplast-associated kinesin-related protein 2 (PAKRP2) GI:16973450 4e-22 ...

Dissecting the molecular mechanisms of intraflagellar transport in Chlamydomonas

DEFF Research Database (Denmark)

Pedersen, L. B.; Geimer, S.; Rosenbaum, J. L.

2006-01-01

Background The assembly and maintenance of eukaryotic cilia and flagella are mediated by intraflagellar transport (IFT), a bidirectional microtubule (MT)-based transport system. The IFT system consists of anterograde (kinesin-2) and retrograde (cDynein1b) motor complexes and IFT particles...... comprising two complexes, A and B. In the current model for IFT, kinesin-2 carries cDynein1b, IFT particles, and axonemal precursors from the flagellar base to the tip, and cDynein1b transports kinesin-2, IFT particles, and axonemal turnover products from the tip back to the base. Most of the components...... of the IFT system have been identified and characterized, but the mechanisms by which these different components are coordinated and regulated at the flagellar base and tip are unclear. Results Using a variety of Chlamydomonas mutants, we confirm that cDynein1b requires kinesin-2 for transport toward the tip...

A single amino acid substitution in the exoplasmic domain of the human growth hormone (GH) receptor confers familial GH resistance (Laron syndrome) with positive GH-binding activity by abolishing receptor homodimerization.

Science.gov (United States)

Duquesnoy, P; Sobrier, M L; Duriez, B; Dastot, F; Buchanan, C R; Savage, M O; Preece, M A; Craescu, C T; Blouquit, Y; Goossens, M

1994-01-01

Growth hormone (GH) elicits a variety of biological activities mainly mediated by the GH receptor (GHR), a transmembrane protein that, based on in vitro studies, seemed to function as a homodimer. To test this hypothesis directly, we investigated patients displaying the classic features of Laron syndrome (familial GH resistance characterized by severe dwarfism and metabolic dysfunction), except for the presence of normal binding activity of the plasma GH-binding protein, a molecule that derives from the exoplasmic-coding domain of the GHR gene. In two unrelated families, the same GHR mutation was identified, resulting in the substitution of a highly conserved aspartate residue by histidine at position 152 (D152H) of the exoplasmic domain, within the postulated interface sequence involved in homodimerization. The recombinant mutated receptor protein was correctly expressed at the plasma membrane. It displayed subnormal GH-binding activity, a finding in agreement with the X-ray crystal structure data inferring this aspartate residue outside the GH-binding domain. However, mAb-based studies suggested the critical role of aspartate 152 in the proper folding of the interface area. We show that a recombinant soluble form of the mutant receptor is unable to dimerize, the D152H substitution also preventing the formation of heterodimers of wild-type and mutant molecules. These results provide in vivo evidence that monomeric receptors are inactive and that receptor dimerization is involved in the primary signalling of the GH-associated growth-promoting and metabolic actions. Images PMID:8137822

Regulation of microtubule-based transport by MAP4

Science.gov (United States)

Semenova, Irina; Ikeda, Kazuho; Resaul, Karim; Kraikivski, Pavel; Aguiar, Mike; Gygi, Steven; Zaliapin, Ilya; Cowan, Ann; Rodionov, Vladimir

2014-01-01

Microtubule (MT)-based transport of organelles driven by the opposing MT motors kinesins and dynein is tightly regulated in cells, but the underlying molecular mechanisms remain largely unknown. Here we tested the regulation of MT transport by the ubiquitous protein MAP4 using Xenopus melanophores as an experimental system. In these cells, pigment granules (melanosomes) move along MTs to the cell center (aggregation) or to the periphery (dispersion) by means of cytoplasmic dynein and kinesin-2, respectively. We found that aggregation signals induced phosphorylation of threonine residues in the MT-binding domain of the Xenopus MAP4 (XMAP4), thus decreasing binding of this protein to MTs. Overexpression of XMAP4 inhibited pigment aggregation by shortening dynein-dependent MT runs of melanosomes, whereas removal of XMAP4 from MTs reduced the length of kinesin-2–dependent runs and suppressed pigment dispersion. We hypothesize that binding of XMAP4 to MTs negatively regulates dynein-dependent movement of melanosomes and positively regulates kinesin-2–based movement. Phosphorylation during pigment aggregation reduces binding of XMAP4 to MTs, thus increasing dynein-dependent and decreasing kinesin-2–dependent motility of melanosomes, which stimulates their accumulation in the cell center, whereas dephosphorylation of XMAP4 during dispersion has an opposite effect. PMID:25143402

The role of the Hedgehog signaling pathway in cancer: A comprehensive review

Directory of Open Access Journals (Sweden)

Ana Marija Skoda

2018-02-01

Full Text Available The Hedgehog (Hh signaling pathway was first identified in the common fruit fly. It is a highly conserved evolutionary pathway of signal transmission from the cell membrane to the nucleus. The Hh signaling pathway plays an important role in the embryonic development. It exerts its biological effects through a signaling cascade that culminates in a change of balance between activator and repressor forms of glioma-associated oncogene (Gli transcription factors. The components of the Hh signaling pathway involved in the signaling transfer to the Gli transcription factors include Hedgehog ligands (Sonic Hh [SHh], Indian Hh [IHh], and Desert Hh [DHh], Patched receptor (Ptch1, Ptch2, Smoothened receptor (Smo, Suppressor of fused homolog (Sufu, kinesin protein Kif7, protein kinase A (PKA, and cyclic adenosine monophosphate (cAMP. The activator form of Gli travels to the nucleus and stimulates the transcription of the target genes by binding to their promoters. The main target genes of the Hh signaling pathway are PTCH1, PTCH2, and GLI1. Deregulation of the Hh signaling pathway is associated with developmental anomalies and cancer, including Gorlin syndrome, and sporadic cancers, such as basal cell carcinoma, medulloblastoma, pancreatic, breast, colon, ovarian, and small-cell lung carcinomas. The aberrant activation of the Hh signaling pathway is caused by mutations in the related genes (ligand-independent signaling or by the excessive expression of the Hh signaling molecules (ligand-dependent signaling – autocrine or paracrine. Several Hh signaling pathway inhibitors, such as vismodegib and sonidegib, have been developed for cancer treatment. These drugs are regarded as promising cancer therapies, especially for patients with refractory/advanced cancers.

Design and Evaluation of Micellar Nanocarriers for 17-allyamino-17-demethoxygeldanamycin (17-AAG)

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17-Allyamino-17-demethoxygeldanamycin (17-AAG) is a potent anticancer agent currently undergoing phases I and II clinical trials. However, the clinical development of 17-AAG has been hindered by its poor aqueous solubility and hepatotoxicity. This study aimed to devise novel mice...

Regulation of the PI3K pathway through a p85α monomer–homodimer equilibrium

KAUST Repository

Cheung, Lydia W T; Walkiewicz, Katarzyna Wiktoria; Besong, Tabot M.D.; Guo, Huifang; Hawke, David H.; Arold, Stefan T.; Mills, Gordon B.

2015-01-01

The canonical action of the p85α regulatory subunit of phosphatidylinositol 3-kinase (PI3K) is to associate with the p110α catalytic subunit to allow stimuli-dependent activation of the PI3K pathway. We elucidate a p110α-independent role of homodimerized p85α in the positive regulation of PTEN stability and activity. p110α-free p85α homodimerizes via two intermolecular interactions (SH3:proline-rich region and BH:BH) to selectively bind unphosphorylated activated PTEN. As a consequence, homodimeric but not monomeric p85α suppresses the PI3K pathway by protecting PTEN from E3 ligase WWP2-mediated proteasomal degradation. Further, the p85α homodimer enhances the lipid phosphatase activity and membrane association of PTEN. Strikingly, we identified cancer patient-derived oncogenic p85α mutations that target the homodimerization or PTEN interaction surface. Collectively, our data suggest the equilibrium of p85α monomerdimers regulates the PI3K pathway and disrupting this equilibrium could lead to disease development. © Cheung et al.

Regulation of the PI3K pathway through a p85α monomer–homodimer equilibrium

KAUST Repository

Cheung, Lydia W T

2015-07-29

The canonical action of the p85α regulatory subunit of phosphatidylinositol 3-kinase (PI3K) is to associate with the p110α catalytic subunit to allow stimuli-dependent activation of the PI3K pathway. We elucidate a p110α-independent role of homodimerized p85α in the positive regulation of PTEN stability and activity. p110α-free p85α homodimerizes via two intermolecular interactions (SH3:proline-rich region and BH:BH) to selectively bind unphosphorylated activated PTEN. As a consequence, homodimeric but not monomeric p85α suppresses the PI3K pathway by protecting PTEN from E3 ligase WWP2-mediated proteasomal degradation. Further, the p85α homodimer enhances the lipid phosphatase activity and membrane association of PTEN. Strikingly, we identified cancer patient-derived oncogenic p85α mutations that target the homodimerization or PTEN interaction surface. Collectively, our data suggest the equilibrium of p85α monomerdimers regulates the PI3K pathway and disrupting this equilibrium could lead to disease development. © Cheung et al.

ICK is essential for cell type-specific ciliogenesis and the regulation of ciliary transport.

Science.gov (United States)

Chaya, Taro; Omori, Yoshihiro; Kuwahara, Ryusuke; Furukawa, Takahisa

2014-06-02

Cilia and flagella are formed and maintained by intraflagellar transport (IFT) and play important roles in sensing and moving across species. At the distal tip of the cilia/flagella, IFT complexes turn around to switch from anterograde to retrograde transport; however, the underlying regulatory mechanism is unclear. Here, we identified ICK localization at the tip of cilia as a regulator of ciliary transport. In ICK-deficient mice, we found ciliary defects in neuronal progenitor cells with Hedgehog signal defects. ICK-deficient cells formed cilia with mislocalized Hedgehog signaling components. Loss of ICK caused the accumulation of IFT-A, IFT-B, and BBSome components at the ciliary tips. In contrast, overexpression of ICK induced the strong accumulation of IFT-B, but not IFT-A or BBSome components at ciliary tips. In addition, ICK directly phosphorylated Kif3a, while inhibition of this Kif3a phosphorylation affected ciliary formation. Our results suggest that ICK is a Kif3a kinase and essential for proper ciliogenesis in development by regulating ciliary transport at the tip of cilia. © 2014 The Authors.

Acquired resistance to 17-allylamino-17-demethoxygeldanamycin (17-AAG, tanespimycin) in glioblastoma cells.

Science.gov (United States)

Gaspar, Nathalie; Sharp, Swee Y; Pacey, Simon; Jones, Chris; Walton, Michael; Vassal, Gilles; Eccles, Suzanne; Pearson, Andrew; Workman, Paul

2009-03-01

Heat shock protein 90 (HSP90) inhibitors, such as 17-allylamino-17-demethoxygeldanamycin (17-AAG, tanespimycin), which is currently in phase II/phase III clinical trials, are promising new anticancer agents. Here, we explored acquired resistance to HSP90 inhibitors in glioblastoma (GB), a primary brain tumor with poor prognosis. GB cells were exposed continuously to increased 17-AAG concentrations. Four 17-AAG-resistant GB cell lines were generated. High-resistance levels with resistance indices (RI = resistant line IC(50)/parental line IC(50)) of 20 to 137 were obtained rapidly (2-8 weeks). After cessation of 17-AAG exposure, RI decreased and then stabilized. Cross-resistance was found with other ansamycin benzoquinones but not with the structurally unrelated HSP90 inhibitors, radicicol, the purine BIIB021, and the resorcinylic pyrazole/isoxazole amide compounds VER-49009, VER-50589, and NVP-AUY922. An inverse correlation between NAD(P)H/quinone oxidoreductase 1 (NQO1) expression/activity and 17-AAG IC(50) was observed in the resistant lines. The NQO1 inhibitor ES936 abrogated the differential effects of 17-AAG sensitivity between the parental and resistant lines. NQO1 mRNA levels and NQO1 DNA polymorphism analysis indicated different underlying mechanisms: reduced expression and selection of the inactive NQO1*2 polymorphism. Decreased NQO1 expression was also observed in a melanoma line with acquired resistance to 17-AAG. No resistance was generated with VER-50589 and NVP-AUY922. In conclusion, low NQO1 activity is a likely mechanism of acquired resistance to 17-AAG in GB, melanoma, and, possibly, other tumor types. Such resistance can be overcome with novel HSP90 inhibitors.

Unifying principles in homodimeric type I photosynthetic reaction centers: properties of PscB and the FA, FB and FX iron-sulfur clusters in green sulfur bacteria.

Science.gov (United States)

Jagannathan, Bharat; Golbeck, John H

2008-12-01

The photosynthetic reaction center from the green sulfur bacterium Chlorobium tepidum (CbRC) was solubilized from membranes using Triton X-100 and isolated by sucrose density ultra-centrifugation. The CbRC complexes were subsequently treated with 0.5 M NaCl and ultrafiltered over a 100 kDa cutoff membrane. The resulting CbRC cores did not exhibit the low-temperature EPR resonances from FA- and FB- and were unable to reduce NADP+. SDS-PAGE and mass spectrometric analysis showed that the PscB subunit, which harbors the FA and FB clusters, had become dissociated, and was now present in the filtrate. Attempts to rebind PscB onto CbRC cores were unsuccessful. Mössbauer spectroscopy showed that recombinant PscB contains a heterogeneous mixture of [4Fe-4S]2+,1+ and other types of Fe/S clusters tentatively identified as [2Fe-2S]2+,1+ clusters and rubredoxin-like Fe3+,2+ centers, and that the [4Fe-4S]2+,1+ clusters which were present were degraded at high ionic strength. Quantitative analysis confirmed that the amount of iron and sulfide in the recombinant protein was sub-stoichiometric. A heme-staining assay indicated that cytochrome c551 remained firmly attached to the CbRC cores. Low-temperature EPR spectroscopy of photoaccumulated CbRC complexes and CbRC cores showed resonances between g=5.4 and 4.4 assigned to a S=3/2 ground spin state [4Fe-4S]1+ cluster and at g=1.77 assigned to a S=1/2 ground spin state [4Fe-4S]1+ cluster, both from FX-. These results unify the properties of the acceptor side of the Type I homodimeric reaction centers found in green sulfur bacteria and heliobacteria: in both, the FA and FB iron-sulfur clusters are present on a salt-dissociable subunit, and FX is present as an interpolypeptide [4Fe-4S]2+,1+ cluster with a significant population in a S=3/2 ground spin state.

Molecular mechanism of 17-allylamino-17-demethoxygeldanamycin (17-AAG)-induced AXL receptor tyrosine kinase degradation.

Science.gov (United States)

Krishnamoorthy, Gnana Prakasam; Guida, Teresa; Alfano, Luigi; Avilla, Elvira; Santoro, Massimo; Carlomagno, Francesca; Melillo, Rosa Marina

2013-06-14

The receptor tyrosine kinase AXL is overexpressed in many cancer types including thyroid carcinomas and has well established roles in tumor formation and progression. Proper folding, maturation, and activity of several oncogenic receptor tyrosine kinases require HSP90 chaperoning. HSP90 inhibition by the antibiotic geldanamycin or its derivative 17-allylamino-17-demethoxygeldanamycin (17-AAG) causes destabilization of its client proteins. Here we show that AXL is a novel client protein of HSP90. 17-AAG induced a time- and dose-dependent down-regulation of endogenous or ectopically expressed AXL protein, thereby inhibiting AXL-mediated signaling and biological activity. 17-AAG-induced AXL down-regulation specifically affected fully glycosylated mature receptor present on cell membrane. By using biotin and [(35)S]methionine labeling, we showed that 17-AAG caused depletion of membrane-localized AXL by mediating its degradation in the intracellular compartment, thus restricting its exposure on the cell surface. 17-AAG induced AXL polyubiquitination and subsequent proteasomal degradation; under basal conditions, AXL co-immunoprecipitated with HSP90. Upon 17-AAG treatment, AXL associated with the co-chaperone HSP70 and the ubiquitin E3 ligase carboxyl terminus of HSC70-interacting protein (CHIP). Overexpression of CHIP, but not of the inactive mutant CHIP K30A, induced accumulation of AXL polyubiquitinated species upon 17-AAG treatment. The sensitivity of AXL to 17-AAG required its intracellular domain because an AXL intracellular domain-deleted mutant was insensitive to the compound. Active AXL and kinase-dead AXL were similarly sensitive to 17-AAG, implying that 17-AAG sensitivity does not require receptor phosphorylation. Overall our data elucidate the molecular basis of AXL down-regulation by HSP90 inhibitors and suggest that HSP90 inhibition in anticancer therapy can exert its effect through inhibition of multiple kinases including AXL.

Molecular Mechanism of 17-Allylamino-17-demethoxygeldanamycin (17-AAG)-induced AXL Receptor Tyrosine Kinase Degradation*

Science.gov (United States)

Krishnamoorthy, Gnana Prakasam; Guida, Teresa; Alfano, Luigi; Avilla, Elvira; Santoro, Massimo; Carlomagno, Francesca; Melillo, Rosa Marina

2013-01-01

The receptor tyrosine kinase AXL is overexpressed in many cancer types including thyroid carcinomas and has well established roles in tumor formation and progression. Proper folding, maturation, and activity of several oncogenic receptor tyrosine kinases require HSP90 chaperoning. HSP90 inhibition by the antibiotic geldanamycin or its derivative 17-allylamino-17-demethoxygeldanamycin (17-AAG) causes destabilization of its client proteins. Here we show that AXL is a novel client protein of HSP90. 17-AAG induced a time- and dose-dependent down-regulation of endogenous or ectopically expressed AXL protein, thereby inhibiting AXL-mediated signaling and biological activity. 17-AAG-induced AXL down-regulation specifically affected fully glycosylated mature receptor present on cell membrane. By using biotin and [35S]methionine labeling, we showed that 17-AAG caused depletion of membrane-localized AXL by mediating its degradation in the intracellular compartment, thus restricting its exposure on the cell surface. 17-AAG induced AXL polyubiquitination and subsequent proteasomal degradation; under basal conditions, AXL co-immunoprecipitated with HSP90. Upon 17-AAG treatment, AXL associated with the co-chaperone HSP70 and the ubiquitin E3 ligase carboxyl terminus of HSC70-interacting protein (CHIP). Overexpression of CHIP, but not of the inactive mutant CHIP K30A, induced accumulation of AXL polyubiquitinated species upon 17-AAG treatment. The sensitivity of AXL to 17-AAG required its intracellular domain because an AXL intracellular domain-deleted mutant was insensitive to the compound. Active AXL and kinase-dead AXL were similarly sensitive to 17-AAG, implying that 17-AAG sensitivity does not require receptor phosphorylation. Overall our data elucidate the molecular basis of AXL down-regulation by HSP90 inhibitors and suggest that HSP90 inhibition in anticancer therapy can exert its effect through inhibition of multiple kinases including AXL. PMID:23629654

Coin Tossing Explains the Activity of Opposing Microtubule Motors on Phagosomes.

Science.gov (United States)

Sanghavi, Paulomi; D'Souza, Ashwin; Rai, Ashim; Rai, Arpan; Padinhatheeri, Ranjith; Mallik, Roop

2018-05-07

How the opposing activity of kinesin and dynein motors generates polarized distribution of organelles inside cells is poorly understood and hotly debated [1, 2]. Possible explanations include stochastic mechanical competition [3, 4], coordinated regulation by motor-associated proteins [5-7], mechanical activation of motors [8], and lipid-induced organization [9]. Here, we address this question by using phagocytosed latex beads to generate early phagosomes (EPs) that move bidirectionally along microtubules (MTs) in an in vitro assay [9]. Dynein/kinesin activity on individual EPs is recorded as real-time force generation of the motors against an optical trap. Activity of one class of motors frequently coincides with, or is rapidly followed by opposite motors. This leads to frequent and rapid reversals of EPs in the trap. Remarkably, the choice between dynein and kinesin can be explained by the tossing of a coin. Opposing motors therefore appear to function stochastically and independently of each other, as also confirmed by observing no effect on kinesin function when dynein is inhibited on the EPs. A simple binomial probability calculation based on the geometry of EP-microtubule contact explains the observed activity of dynein and kinesin on phagosomes. This understanding of intracellular transport in terms of a hypothetical coin, if it holds true for other cargoes, provides a conceptual framework to explain the polarized localization of organelles inside cells. Copyright © 2018 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Soluble N-ethylmaleimide-sensitive factor attachment protein receptors required during Trypanosoma cruzi parasitophorous vacuole development.

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Cueto, Juan Agustín; Vanrell, María Cristina; Salassa, Betiana Nebaí; Nola, Sébastien; Galli, Thierry; Colombo, María Isabel; Romano, Patricia Silvia

2017-06-01

Trypanosoma cruzi, the etiologic agent of Chagas disease, is an obligate intracellular parasite that exploits different host vesicular pathways to invade the target cells. Vesicular and target soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) are key proteins of the intracellular membrane fusion machinery. During the early times of T. cruzi infection, several vesicles are attracted to the parasite contact sites in the plasma membrane. Fusion of these vesicles promotes the formation of the parasitic vacuole and parasite entry. In this work, we study the requirement and the nature of SNAREs involved in the fusion events that take place during T. cruzi infection. Our results show that inhibition of N-ethylmaleimide-sensitive factor protein, a protein required for SNARE complex disassembly, impairs T. cruzi infection. Both TI-VAMP/VAMP7 and cellubrevin/VAMP3, two v-SNAREs of the endocytic and exocytic pathways, are specifically recruited to the parasitophorous vacuole membrane in a synchronized manner but, although VAMP3 is acquired earlier than VAMP7, impairment of VAMP3 by tetanus neurotoxin fails to reduce T. cruzi infection. In contrast, reduction of VAMP7 activity by expression of VAMP7's longin domain, depletion by small interfering RNA or knockout, significantly decreases T. cruzi infection susceptibility as a result of a minor acquisition of lysosomal components to the parasitic vacuole. In addition, overexpression of the VAMP7 partner Vti1b increases the infection, whereas expression of a KIF5 kinesin mutant reduces VAMP7 recruitment to vacuole and, concomitantly, T. cruzi infection. Altogether, these data support a key role of TI-VAMP/VAMP7 in the fusion events that culminate in the T. cruzi parasitophorous vacuole development. © 2016 John Wiley & Sons Ltd.

IL-17A, IL-17RC polymorphisms and IL17 plasma levels in Tunisian patients with rheumatoid arthritis

Science.gov (United States)

Chahbi, Mayssa; Haouami, Youssra; Sfar, Imen; Abdelmoula, Leila; Ben Abdallah, Taieb; Gorgi, Yousr

2018-01-01

Background Interleukin-17 (IL-17), a cytokine mainly secreted by Th17 cells, seems to play a significant role in the pathogenesis of rheumatoid arthritis (RA). Functional genetic polymorphisms in IL-17 and its receptor genes can influence either qualitatively or quantitatively their functions. Therefore, we aimed to study the impact of IL17-A and IL17RC polymorphisms on plasma level of IL-17 and RA susceptibility and severity. Methods In this context, IL-17A*rs2275913 and IL-17RC*rs708567 polymorphisms were investigated together with the quantification of IL17 plasma level in 115 RA patients and 91 healthy control subjects matched in age, sex and ethnic origin. Results There were no statistically significant associations between IL-17A and IL-17RC studied polymorphisms and RA susceptibility. In contrast, IL-17A plasma levels were significantly higher in patients (55.07 pg/ml) comparatively to controls (4.75 pg/ml), p<10E-12. A ROC curve was used to evaluate the performance of plasma IL-17 in detecting RA. Given 100% specificity, the highest sensitivity of plasma IL-17A was 61.7% at a cut-off value of 18.25 pg/ml; p < 10E-21, CI = [0.849–0.939]. Analytic results showed that the IgM-rheumatoid factor and anti-CCP antibodies were significantly less frequent in patients with the IL-17RC*A/A genotype than those carrying *G/G and *G/A genotypes; p = 0.013 and p = 0.015, respectively. Otherwise, IL-17 plasma levels’ analysis showed a significant association with the activity of RA (DAS28≥5.1 = 74.71 pg/ml vs. DAS28<5.1 = 11.96 pg/ml), p<10E-6. Conclusion IL-17A*rs2275913 (G/A) and IL-17RC*rs708567 (G/A) polymorphisms did not seem to influence RA susceptibility in Tunisian population. This result agrees with those reported previously. Plasma IL-17A level seems to be predictive of severe RA occurrence. PMID:29584788

Albumin Homodimers in Patients with Cirrhosis: Clinical and Prognostic Relevance of a Novel Identified Structural Alteration of the Molecule.

Science.gov (United States)

Baldassarre, Maurizio; Domenicali, Marco; Naldi, Marina; Laggetta, Maristella; Giannone, Ferdinando A; Biselli, Maurizio; Patrono, Daniela; Bertucci, Carlo; Bernardi, Mauro; Caraceni, Paolo

2016-10-26

Decompensated cirrhosis is associated to extensive post-transcriptional changes of human albumin (HA). This study aims to characterize the occurrence of HA homodimerization in a large cohort of patients with decompensated cirrhosis and to evaluate its association with clinical features and prognosis. HA monomeric and dimeric isoforms were identified in peripheral blood by using a HPLC-ESI-MS technique in 123 cirrhotic patients hospitalized for acute decompensation and 50 age- and sex-comparable healthy controls. Clinical and biochemical parameters were recorded and patients followed up to one year. Among the monomeric isoforms identified, the N- and C-terminal truncated and the native HA underwent homodimerization. All three homodimers were significantly more abundant in patients with cirrhosis, acute-on-chronic liver failure and correlate with the prognostic scores. The homodimeric N-terminal truncated isoform was independently associated to disease complications and was able to stratify 1-year survival. As a result of all these changes, the monomeric native HA was significantly decreased in patients with cirrhosis, being also associated with a poorer prognosis. In conclusion homodimerization is a novel described structural alteration of the HA molecule in decompensated cirrhosis and contributes to the progressive reduction of the monomeric native HA, the only isoform provided of structural and functional integrity.

A Phase II trial of 17-allylamino, 17-demethoxygeldanamycin (17-AAG, tanespimycin) in patients with metastatic melanoma.

Science.gov (United States)

Pacey, Simon; Gore, Martin; Chao, David; Banerji, Udai; Larkin, James; Sarker, Sarah; Owen, Karen; Asad, Yasmin; Raynaud, Florence; Walton, Mike; Judson, Ian; Workman, Paul; Eisen, Tim

2012-02-01

A Phase II study to screen for anti-melanoma activity of the heat shock protein 90 (HSP90) inhibitor, 17-AAG (17-allylamino-17-demethoxygeldanamycin) was performed. The primary endpoint was the rate of disease stabilisation in patients with progressive, metastatic melanoma treated with 17-AAG. Secondary endpoints were to determine: the toxicity of 17-AAG, the duration of response(s), median survival and further study the pharmacokinetics and pharmacodynamics of 17-AAG. Patients with metastatic melanoma (progressive disease documented ≤6 months of entering study) were treated with weekly, intravenous 17-AAG. A Simon one sample two stage minimax design was used. A stable disease rate of ≥25% at 6 months was considered compatible with 17-AAG having activity. Fourteen patients (8 male: 6 female) were entered, eleven received 17-AAG (performance status 0 or 1). Median age was 60 (range 29-81) years. The majority (93%) received prior chemotherapy and had stage M1c disease (71%). Toxicity was rarely ≥ Grade 2 in severity and commonly included fatigue, headache and gastrointestinal disturbances. One of eleven patients treated with 17-AAG had stable disease for 6 months and median survival for all patients was 173 days. The study was closed prematurely prior to completion of the first stage of recruitment and limited planned pharmacokinetic and pharmacodynamic analyses. Some evidence of 17-AAG activity was observed although early study termination meant study endpoints were not reached. Stable disease rates can be incorporated into trials screening for anti-melanoma activity and further study of HSP90 inhibitors in melanoma should be considered.

Multigeneration family with dominant SPG30 hereditary spastic paraplegia.

Science.gov (United States)

Roda, Ricardo H; Schindler, Alice B; Blackstone, Craig

2017-11-01

Autosomal recessive KIF1A missense mutations cause hereditary spastic paraplegia (HSP) type SPG30, while recessive truncations lead to sensory and autonomic neuropathy (HSN2C) and many de novo missense mutations are associated with cognitive impairment. Here, we describe family members across three generations with pure HSP. A heterozygous p.Ser69Leu KIF1A mutation segregates with those afflicted. The same variant was previously reported in a Finnish father and son with pure HSP as well as four members of a Sicilian kindred with more intrafamilial phenotypic variability. This further validates the pathogenicity of the p.Ser69Leu mutation and suggests that it may represent a mutation hot spot.

17 CFR 12.17 - Satisfaction of complaint.

Science.gov (United States)

2010-04-01

... 17 Commodity and Securities Exchanges 1 2010-04-01 2010-04-01 false Satisfaction of complaint. 12... RELATING TO REPARATIONS General Information and Preliminary Consideration of Pleadings § 12.17 Satisfaction... as the complainant will accept in satisfaction of his claim; and (b) by submitting to the Commission...

Folate receptor targeted 17-allylamino-17-demethoxygeldanamycin (17-AAG) loaded polymeric nanoparticles for breast cancer.

Science.gov (United States)

Saxena, Vipin; Naguib, Youssef; Hussain, M Delwar

2012-06-01

Low water solubility and hepatotoxicity limited the clinical use of 17-allylamino-17-demethoxy geldanamycin (17-AAG), an inhibitor of heat shock protein 90 (HSP90). Folate targeted polylactide-co-glycolide-polyethylene glycol-folic acid (PLGA-PEG-FA) nanoparticles containing 17-AAG were prepared and characterized. Cellular uptake and in vitro cytotoxicity of the prepared nanoparticles were determined in MCF-7 human breast cancer cells. The particle size of 17-AAG loaded folate targeted nanoparticles was 238.67±3.52 nm, drug loading was 8.25±2.49% and about 80% of drug was released from the nanoparticles over 10 days. Cellular uptake studies showed much higher intracellular uptake of folate targeted nanoparticle as compared to nontargeted nanoparticles. Cytotoxicity study showed 2 fold increase (PAAG loaded PLGA-PEG-FA nanoparticles might be developed as a targeted delivery system for breast and other cancer treatment. Copyright © 2012 Elsevier B.V. All rights reserved.

New 17x17 assembly takes the heat

International Nuclear Information System (INIS)

Reparaz, A.; Brown, C.A. Sr.

1992-01-01

Advanced Nuclear Fuels' (ANF's) new 17 x 17 fuel assembly described in this article offers advanced features which provide increased thermal margins, higher burnup, responsiveness to electrical grid demands, improved seismic performance and superior corrosion resistance. (Author)

Growth hormone (GH)-independent dimerization of GH receptor by a leucine zipper results in constitutive activation

DEFF Research Database (Denmark)

Behncken, S N; Billestrup, Nils; Brown, R

2000-01-01

Growth hormone initiates signaling by inducing homodimerization of two GH receptors. Here, we have sought to determine whether constitutively active receptor can be created in the absence of the extracellular domain by substituting it with high affinity leucine zippers to create dimers of the gro......Growth hormone initiates signaling by inducing homodimerization of two GH receptors. Here, we have sought to determine whether constitutively active receptor can be created in the absence of the extracellular domain by substituting it with high affinity leucine zippers to create dimers...

Magnetic excitations in Ho2Co17 and Ho2Fe17

International Nuclear Information System (INIS)

Clausen, K.N.

1981-01-01

The low energy part ( 2 Co 17 and Ho 2 Fe 17 have been measured along the three high symmetry directions at a temperature of 4.2 K, using the inelastic neutron scattering technique. The resulting magnon dispersion relations have been interpreted using linear spin wave theory with a Hamiltonian including single ion crystal field anisotropy and isotropic exchange between spatially well localized spins. The R 2 T 17 structure contains two different Ho sites, with the same point symmetry, and from the spin wave results it was concluded that the crystal field anisotropy of the two Ho sites in both Ho 2 Co 17 and Ho 2 Fe 17 were identical. The deduced crystal field parameters for Ho 2 Fe 17 were slightly larger than for Ho 2 Co 17 , and the parameters were of the same order of magnitude as for pure Ho. For Ho 2 Fe 17 the Fe-Fe exchange was found to be anisotropic, and for both compounds the magnetic ordering temperatures of 1178 K for Ho 2 Co 17 and 335 K for Ho 2 Fe 17 were determined by the strong positive 3d-3d exchange. (Auth.)

IL-17/Th17 Pathway Is Activated in Acne Lesions

Science.gov (United States)

Kelhälä, Hanna-Leena; Palatsi, Riitta; Fyhrquist, Nanna; Lehtimäki, Sari; Väyrynen, Juha P.; Kallioinen, Matti; Kubin, Minna E.; Greco, Dario; Tasanen, Kaisa; Alenius, Harri; Bertino, Beatrice; Carlavan, Isabelle; Mehul, Bruno; Déret, Sophie; Reiniche, Pascale; Martel, Philippe; Marty, Carine; Blume-Peytavi, Ulrike; Voegel, Johannes J.; Lauerma, Antti

2014-01-01

The mechanisms of inflammation in acne are currently subject of intense investigation. This study focused on the activation of adaptive and innate immunity in clinically early visible inflamed acne lesions and was performed in two independent patient populations. Biopsies were collected from lesional and non-lesional skin of acne patients. Using Affymetrix Genechips, we observed significant elevation of the signature cytokines of the Th17 lineage in acne lesions compared to non-lesional skin. The increased expression of IL-17 was confirmed at the RNA and also protein level with real-time PCR (RT-PCR) and Luminex technology. Cytokines involved in Th17 lineage differentiation (IL-1β, IL-6, TGF-β, IL23p19) were remarkably induced at the RNA level. In addition, proinflammatory cytokines and chemokines (TNF-α, IL-8, CSF2 and CCL20), Th1 markers (IL12p40, CXCR3, T-bet, IFN-γ), T regulatory cell markers (Foxp3, IL-10, TGF-β) and IL-17 related antimicrobial peptides (S100A7, S100A9, lipocalin, hBD2, hBD3, hCAP18) were induced. Importantly, immunohistochemistry revealed significantly increased numbers of IL-17A positive T cells and CD83 dendritic cells in the acne lesions. In summary our results demonstrate the presence of IL-17A positive T cells and the activation of Th17-related cytokines in acne lesions, indicating that the Th17 pathway is activated and may play a pivotal role in the disease process, possibly offering new targets of therapy. PMID:25153527

Electrochemistry of cytochrome P450 17α-hydroxylase/17,20-lyase (P450c17).

Science.gov (United States)

Martin, Lisandra L; Kubeil, Clemens; Simonov, Alexandr N; Kuznetsov, Vladimir L; Corbin, C Jo; Auchus, Richard J; Conley, Alan J; Bond, Alan M; Rodgers, Raymond J

2017-02-05

Within the superfamily of cytochrome P450 enzymes (P450s), there is a small class which is functionally employed for steroid biosynthesis. The enzymes in this class appear to have a small active site to accommodate the steroid substrates specifically and snuggly, prior to the redox transformation or hydroxylation to form a product. Cytochrome P450c17 is one of these and is also a multi-functional P450, with two activities, the first 17α-hydroxylation of pregnenolone is followed by a subsequent 17,20-lyase transformation to dehydroepiandrosterone (DHEA) as the dominant pathways to cortisol precursors or androgens in humans, respectively. How P450c17 regulates these two redox reactions is of special interest. There is a paucity of direct electrochemical studies on steroidogenic P450s, and in this mini-review we provide an overview of these studies with P450c17. Historical consideration as to the difficulties in obtaining reliable electrochemistry due to issues of handling proteins on an electrode, together with advances in the electrochemical techniques are addressed. Recent work using Fourier transformed alternating current voltammetry is highlighted as this technique can provide both catalytic information simultaneously with the underlying redox transfer with the P450 haem. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Preparation and evaluation of 17-allyamino-17-demethoxygeldanamycin (17-AAG)-loaded poly(lactic acid-co-glycolic acid) nanoparticles.

Science.gov (United States)

Pradhan, Roshan; Poudel, Bijay Kumar; Choi, Ju Yeon; Choi, Im Soon; Shin, Beom Soo; Choi, Han-Gon; Yong, Chul Soon; Kim, Jong Oh

2015-01-01

In the present study, we developed the novel 17-allyamino-17-demethoxygeldanamycin (17-AAG)-loaded poly(lactic acid-co-glycolic acid) (PLGA) nanoparticles (NPs) using the combination of sodium lauryl sulfate and poloxamer 407 as the anionic and non-ionic surfactant for stabilization. The PLGA NPs were prepared by emulsification/solvent evaporation method. Both the drug/polymer ratio and phase ratio were 1:10 (w/w). The optimized formulation of 17-AAG-loaded PLGA NPs had a particle size and polydispersity index of 151.6 ± 2.0 and 0.152 ± 0.010 nm, respectively, which was further supported by TEM image. The encapsulation efficiency and drug loading capacity were 69.9 and 7.0%, respectively. In vitro release study showed sustained release. When in vitro release data were fitted to Korsmeyer-Peppas model, the n value was 0.468, which suggested that the drug was released by anomalous or non-Fickian diffusion. In addition, 17-AAG-loaded PLGA NPs in 72 h, displayed approximately 60% cell viability reduction at 10 µg/ml 17-AAG concentration, in MCF-7 cell lines, indicating sustained release from NPs. Therefore, our results demonstrated that incorporation of 17-AAG into PLGA NPs could provide a novel effective nanocarrier for the treatment of cancer.

Effect of 17 x 17 fuel assembly geometry on interchannel thermal mixing

International Nuclear Information System (INIS)

Motley, F.E.; Wenzell, A.H.; Cadek, F.F.

1975-01-01

A test to determine the value of the thermal diffusion coefficient (TDC) in the 17 x 17 fuel assembly geometry was conducted. The test section was a 5 x 5 rod bundle with a radial power difference of 4.5 to 1. The rod OD and pitch are identical to the 17 x 17 fuel assembly, as is the mixing vane grid design. The value of thermal diffusion coefficient (TDC) was determined by matching the experimental exit enthalpy distribution to that predicted by the THINC computer code. The mean value of TDC for the 17 x 17 fuel assembly geometry is TDC = .059. 6 references

Effects of 17-allylamino-17-demethoxygeldanamycin (17-AAG) in transgenic mouse models of frontotemporal lobar degeneration and Alzheimer's disease.

Science.gov (United States)

Ho, Shuk Wai; Tsui, Yuk Tung Chanel; Wong, Ting Ting; Cheung, Stanley Kwok-Kuen; Goggins, William B; Yi, Lau Ming; Cheng, Kwok Kin; Baum, Larry

2013-12-17

Alzheimer's disease (AD), the most common dementia, is characterized by potentially neurotoxic aggregation of Aβ peptide and tau protein, and their deposition as amyloid plaques and neurofibrillary tangles (NFTs). Tau aggregation also occurs in other common neurodegenerative diseases. Frontotemporal dementia (FTD) can be caused by tau mutations that increase the susceptibility of tau to hyperphosphorylation and aggregation, which may cause neuronal dysfunction and deposition of NFTs. 17-allylamino-17-demethoxygeldanamycin (17-AAG) is a potent inhibitor of heat shock protein 90 (Hsp90), a cytosolic chaperone implicated in the proper folding and functions of a repertoire of client proteins. 17-AAG binds to Hsp90 and enhances degradation of Hsp90 client protein. We sought to determine whether 17-AAG can reduce Aβ and tau pathology in the brains of AD and FTD model mice expressing Aβ or P301L mutant tau, respectively. Mice were randomized to receive 25, 5, or 0 mg/kg 17-AAG thrice weekly from age eight to 11 months. Analysis was performed by rotarod test on motor function, on the area occupied by plaques in hippocampus or NFTs in medulla tissue sections, and on mortality. A high dose of 17-AAG tended to decrease NFTs in male mice (p = 0.08). Further studies are required to confirm the effect of 17-AAG in diseases of tau aggregation.

Th17 cells and IL-17 in protective immunity to vaginal candidiasis.

Science.gov (United States)

Pietrella, Donatella; Rachini, Anna; Pines, Mark; Pandey, Neelam; Mosci, Paolo; Bistoni, Francesco; d'Enfert, Cristophe; Vecchiarelli, Anna

2011-01-01

Th17 cells play a major role in coordinating the host defence in oropharyngeal candidiasis. In this study we investigated the involvement of the Th17 response in an animal model of vulvovaginal candidiasis (VVC). To monitor the course of infection we exploited a new in vivo imaging technique. i) The progression of VVC leads to a strong influx of neutrophils in the vagina soon after the challenge which persisted despite the resolution of infection; ii) IL-17, produced by vaginal cells, particularly CD4 T cells, was detected in the vaginal wash during the infection, reaching a maximum 14 days after the challenge; iii) The amount and kinetics of IL-23 in vaginal fluids were comparable to those in vaginal cells; iv) The inhibition of Th17 differentiation led to significant inhibition of IL-17 production with consequent exacerbation of infection; v) An increased production of βdefensin 2 was manifested in cells of infected mice. This production was strongly reduced when Th17 differentiation was inhibited and was increased by rIL-17 treatment. These results imply that IL-17 and Th17, along with innate antimicrobial factors, have a role in the immune response to vaginal candidiasis.

17 CFR 17.02 - Form, manner and time of filing reports.

Science.gov (United States)

2010-04-01

... 17 Commodity and Securities Exchanges 1 2010-04-01 2010-04-01 false Form, manner and time of filing reports. 17.02 Section 17.02 Commodity and Securities Exchanges COMMODITY FUTURES TRADING... markets located in that time zone, and central time for information concerning all other markets. (b...

Formulation and in vitro evaluation of 17-allyamino-17-demethoxygeldanamycin (17-AAG) loaded polymeric mixed micelles for glioblastoma multiforme.

Science.gov (United States)

Saxena, Vipin; Hussain, Muhammad Delwar

2013-12-01

Glioblastoma multiforme (GBM) is the most common and aggressive malignant primary brain tumor in human. 17-Allylamino-17-demethoxy geldanamycin (17-AAG) is an inhibitor of heat shock protein 90 (HSP90). The highly lipophilic nature and selective targeting of tumor cells makes 17-AAG a promising candidate for therapy of GBMs but poor water solubility, short biological half-life and hepatotoxicity limited its clinical use. Polymeric mixed micelles composed of Pluronic® P-123 and F-127 (2:1 (w/w)) containing 17-AAG were prepared and characterized. Cellular uptake and in vitro cytotoxicity of the prepared micelles were determined in U87MG human glioblastoma cells. The particle size of 17-AAG loaded Pluronic(®) P-123 and F-127 mixed micelles was 22.2 ± 0.1 nm; drug loading was about 4.0 ± 0.5% (w/w) with 88.2 ± 3.1% (w/w) encapsulation efficiency. About 90% of drug was released from the nanoparticles over 8 days. Cellular uptake studies showed intracellular uptake of mixed micelles. Cytotoxicity study showed 5-fold increase (P AAG-loaded mixed micelles to free 17-AAG. Due to their targeting ability, size, high drug loading and controlled release behavior, 17-AAG loaded Pluronic(®) P-123 and F-127 mixed micelles might be developed as a delivery system for GBM treatment. © 2013 Elsevier B.V. All rights reserved.

The IL-17F/IL-17RC Axis Promotes Respiratory Allergy in the Proximal Airways

Directory of Open Access Journals (Sweden)

Antonella De Luca

2017-08-01

Full Text Available The interleukin 17 (IL-17 cytokine and receptor family is central to antimicrobial resistance and inflammation in the lung. Mice lacking IL-17A, IL-17F, or the IL-17RA subunit were compared with wild-type mice for susceptibility to airway inflammation in models of infection and allergy. Signaling through IL-17RA was required for efficient microbial clearance and prevention of allergy; in the absence of IL-17RA, signaling through IL-17RC on epithelial cells, predominantly by IL-17F, significantly exacerbated lower airway Aspergillus or Pseudomonas infection and allergic airway inflammation. In contrast, following infection with the upper respiratory pathogen Staphylococcus aureus, the IL-17F/IL-17RC axis mediated protection. Thus, IL-17A and IL-17F exert distinct biological effects during pulmonary infection; the IL-17F/IL-17RC signaling axis has the potential to significantly worsen pathogen-associated inflammation of the lower respiratory tract in particular, and should be investigated further as a therapeutic target for treating pathological inflammation in the lung.

Effects of 17-allylamino-17-demethoxygeldanamycin (17-AAG) in transgenic mouse models of frontotemporal lobar degeneration and Alzheimer’s disease

Science.gov (United States)

2013-01-01

Alzheimer’s disease (AD), the most common dementia, is characterized by potentially neurotoxic aggregation of Aβ peptide and tau protein, and their deposition as amyloid plaques and neurofibrillary tangles (NFTs). Tau aggregation also occurs in other common neurodegenerative diseases. Frontotemporal dementia (FTD) can be caused by tau mutations that increase the susceptibility of tau to hyperphosphorylation and aggregation, which may cause neuronal dysfunction and deposition of NFTs. 17-allylamino-17-demethoxygeldanamycin (17-AAG) is a potent inhibitor of heat shock protein 90 (Hsp90), a cytosolic chaperone implicated in the proper folding and functions of a repertoire of client proteins. 17-AAG binds to Hsp90 and enhances degradation of Hsp90 client protein. We sought to determine whether 17-AAG can reduce Aβ and tau pathology in the brains of AD and FTD model mice expressing Aβ or P301L mutant tau, respectively. Mice were randomized to receive 25, 5, or 0 mg/kg 17-AAG thrice weekly from age eight to 11 months. Analysis was performed by rotarod test on motor function, on the area occupied by plaques in hippocampus or NFTs in medulla tissue sections, and on mortality. A high dose of 17-AAG tended to decrease NFTs in male mice (p = 0.08). Further studies are required to confirm the effect of 17-AAG in diseases of tau aggregation. PMID:24344631

17 CFR 8.17 - Hearing.

Science.gov (United States)

2010-04-01

... Commodity and Securities Exchanges COMMODITY FUTURES TRADING COMMISSION EXCHANGE PROCEDURES FOR DISCIPLINARY, SUMMARY, AND MEMBERSHIP DENIAL ACTIONS Disciplinary Procedure § 8.17 Hearing. (a) The following minimum... conducted before members of the disciplinary committee. The hearing may be conducted before all of the...

17 CFR 240.17Ad-3 - Limitations on expansion.

Science.gov (United States)

2010-04-01

... 17 Commodity and Securities Exchanges 3 2010-04-01 2010-04-01 false Limitations on expansion. 240... expansion. (a) Any registered transfer agent which is required to file any notice pursuant to § 240.17Ad-2 (c) or (d) for each of three consecutive months shall not from the fifth business day after the end...

A Phase II Trial of 17-Allylamino-17-Demethoxygeldanamycin (17-AAG) in Patients with Hormone-Refractory Metastatic Prostate Cancer

Science.gov (United States)

Heath, Elisabeth I.; Hillman, David W.; Vaishampayan, Ulka; Sheng, Shijie; Sarkar, Fazlul; Harper, Felicity; Gaskins, Melvin; Pitot, Henry C.; Tan, Winston; Ivy, S. Percy; Pili, Roberto; Carducci, Michael A.; Liu, Glenn

2011-01-01

Purpose 17-Allylamino-17-Demethoxygeldanamycin (17-AAG) is a benzoquinone ansamycin antibiotic with anti-proliferative activity in several mouse xenograft models including prostate cancer models. A two-stage phase II study was conducted to assess the activity and toxicity profile of 17-AAG administered to patients with metastatic, hormone-refractory prostate cancer. Experimental Design Patients with at least one prior systemic therapy and a rising PSA were eligible. Patients received 17-AAG at a dose of 300 mg/m2 IV weekly for three out of four weeks. The primary objective was to assess the PSA response. Secondary objectives were to determine overall survival, to assess toxicity, to measure IL-6, IL-8 and maspin levels and quality of life. Results Fifteen eligible patients were enrolled. The median age was 68 years and the median PSA was 261 ng/mL. Patients received 17-AAG for a median number of 2 cycles. Severe adverse events included: grade 3 fatigue (4 pts), grade 3 lymphopenia (2 pts) and grade 3 back pain (2 pts). The median PSA progression free survival was 1.8 months (95% CI: 1.3–3.4 months). The six-month overall survival was 71% (95% CI: 52%–100%). Conclusion 17-AAG did not show any activity with regards to PSA response. Due to insufficient PSA response, enrollment was stopped at end of first stage per study design. The most significant severe toxicity was grade 3 fatigue. Further evaluation of 17-AAG at a dose of 300 mg/m2 IV weekly as a single agent in patients with metastatic, hormone-refractory prostate cancer who received at least one prior systemic therapy is not warranted. PMID:19047126

Inorganic arsenic represses interleukin-17A expression in human activated Th17 lymphocytes

Energy Technology Data Exchange (ETDEWEB)

Morzadec, Claudie; Macoch, Mélinda; Robineau, Marc; Sparfel, Lydie [UMR INSERM U1085, Institut de Recherche sur la Santé, l' Environnement et le Travail (IRSET), Université de Rennes 1, 2 avenue du Professeur Léon Bernard, 35043 Rennes (France); Fardel, Olivier [UMR INSERM U1085, Institut de Recherche sur la Santé, l' Environnement et le Travail (IRSET), Université de Rennes 1, 2 avenue du Professeur Léon Bernard, 35043 Rennes (France); Pôle Biologie, Centre Hospitalier Universitaire (CHU) Rennes, 2 rue Henri Le Guilloux, 35033 Rennes (France); Vernhet, Laurent, E-mail: laurent.vernhet@univ-rennes1.fr [UMR INSERM U1085, Institut de Recherche sur la Santé, l' Environnement et le Travail (IRSET), Université de Rennes 1, 2 avenue du Professeur Léon Bernard, 35043 Rennes (France)

2012-08-01

Trivalent inorganic arsenic [As(III)] is an efficient anticancer agent used to treat patients suffering from acute promyelocytic leukemia. Recently, experimental studies have clearly demonstrated that this metalloid can also cure lymphoproliferative and/or pro-inflammatory syndromes in different murine models of chronic immune-mediated diseases. T helper (Th) 1 and Th17 lymphocytes play a central role in development of these diseases, in mice and humans, especially by secreting the potent pro-inflammatory cytokine interferon-γ and IL-17A, respectively. As(III) impairs basic functions of human T cells but its ability to modulate secretion of pro-inflammatory cytokines by differentiated Th lymphocytes is unknown. In the present study, we demonstrate that As(III), used at concentrations clinically achievable in plasma of patients, has no effect on the secretion of interferon-γ from Th1 cells but almost totally blocks the expression and the release of IL-17A from human Th17 lymphocytes co-stimulated for five days with anti-CD3 and anti-CD28 antibodies, in the presence of differentiating cytokines. In addition, As(III) specifically reduces mRNA levels of the retinoic-related orphan receptor (ROR)C gene which encodes RORγt, a key transcription factor controlling optimal IL-17 expression in fully differentiated Th17 cells. The metalloid also blocks initial expression of IL-17 gene induced by the co-stimulation, probably in part by impairing activation of the JNK/c-Jun pathway. In conclusion, our results demonstrate that As(III) represses expression of the major pro-inflammatory cytokine IL-17A produced by human Th17 lymphocytes, thus strengthening the idea that As(III) may be useful to treat inflammatory immune-mediated diseases in humans. -- Highlights: ► Arsenic inhibits secretion of IL-17A from human naïve and memory Th17 lymphocytes. ► Arsenic represses early expression of IL-17A gene in human activated T lymphocytes. ► Arsenic interferes with activation of

21 CFR 862.1385 - 17-Hydroxycorticosteroids (17-ketogenic steroids) test system.

Science.gov (United States)

2010-04-01

... the adrenal gland. Measurements of 17-hydroxycorticosteroids (17-ketogenic steroids) are used in the diagnosis and treatment of various diseases of the adrenal or pituitary glands and gonadal disorders. (b...

17 CFR 240.17a-11 - Notification provisions for brokers and dealers.

Science.gov (United States)

2010-04-01

... brokers and dealers. 240.17a-11 Section 240.17a-11 Commodity and Securities Exchanges SECURITIES AND... Stabilizing Activities § 240.17a-11 Notification provisions for brokers and dealers. (a) This section shall apply to every broker or dealer registered with the Commission pursuant to section 15 of the Act. (b)(1...

Th17/IL-17A might play a protective role in chronic lymphocytic leukemia immunity.

Directory of Open Access Journals (Sweden)

Iwona Hus

Full Text Available Th17 cells, a recently discovered subset of T helper cells that secrete IL-17A, can affect the inflammation process autoimmune and cancer diseases development. The purpose of this study was to evaluate the role of Th17 cells and IL17A in biology of CLL. The study group included 294 untreated CLL patients in different clinical stages. Here, we show that higher Th17 and IL-17A values were associated with less advanced clinical stage of CLL. Th17 cells' percentages in PB were lower in patients who died due to CLL during follow-up due to CLL (as compared to surviving patients and in patients responding to first-line therapy with fludarabine-based regimens (as compared to non-responders. IL-17A inversely correlated with the time from CLL diagnosis to the start of therapy and was lower in patients who required treatment during follow-up. Th-17 and IL-17A values were lower in patients with adverse prognostic factors (17p and 11q deletion, CD38 and ZAP-70 expression. CLL patients with detectable IL-17A mRNA in T cells were in Rai Stage 0 and negative for both ZAP-70 and CD38 expression. Th17 percentages positively correlated with iNKT and adversely with Treg cells. The results of this study suggest that Th17 may play a beneficial role in CLL immunity.

17 CFR 210.3-17 - Financial statements of natural persons.

Science.gov (United States)

2010-04-01

... 17 Commodity and Securities Exchanges 2 2010-04-01 2010-04-01 false Financial statements of... COMMISSION FORM AND CONTENT OF AND REQUIREMENTS FOR FINANCIAL STATEMENTS, SECURITIES ACT OF 1933, SECURITIES... Financial Statements § 210.3-17 Financial statements of natural persons. (a) In lieu of the financial...

Zebrafish homologs of genes within 16p11.2, a genomic region associated with brain disorders, are active during brain development, and include two deletion dosage sensor genes

Directory of Open Access Journals (Sweden)

Alicia Blaker-Lee

2012-11-01

Deletion or duplication of one copy of the human 16p11.2 interval is tightly associated with impaired brain function, including autism spectrum disorders (ASDs, intellectual disability disorder (IDD and other phenotypes, indicating the importance of gene dosage in this copy number variant region (CNV. The core of this CNV includes 25 genes; however, the number of genes that contribute to these phenotypes is not known. Furthermore, genes whose functional levels change with deletion or duplication (termed ‘dosage sensors’, which can associate the CNV with pathologies, have not been identified in this region. Using the zebrafish as a tool, a set of 16p11.2 homologs was identified, primarily on chromosomes 3 and 12. Use of 11 phenotypic assays, spanning the first 5 days of development, demonstrated that this set of genes is highly active, such that 21 out of the 22 homologs tested showed loss-of-function phenotypes. Most genes in this region were required for nervous system development – impacting brain morphology, eye development, axonal density or organization, and motor response. In general, human genes were able to substitute for the fish homolog, demonstrating orthology and suggesting conserved molecular pathways. In a screen for 16p11.2 genes whose function is sensitive to hemizygosity, the aldolase a (aldoaa and kinesin family member 22 (kif22 genes were identified as giving clear phenotypes when RNA levels were reduced by ∼50%, suggesting that these genes are deletion dosage sensors. This study leads to two major findings. The first is that the 16p11.2 region comprises a highly active set of genes, which could present a large genetic target and might explain why multiple brain function, and other, phenotypes are associated with this interval. The second major finding is that there are (at least two genes with deletion dosage sensor properties among the 16p11.2 set, and these could link this CNV to brain disorders such as ASD and IDD.

17 CFR 270.17a-8 - Mergers of affiliated companies.

Science.gov (United States)

2010-04-01

... 17 Commodity and Securities Exchanges 3 2010-04-01 2010-04-01 false Mergers of affiliated... (CONTINUED) RULES AND REGULATIONS, INVESTMENT COMPANY ACT OF 1940 § 270.17a-8 Mergers of affiliated companies. (a) Exemption of affiliated mergers. A Merger of a registered investment company (or a series thereof...

Synthesis of (131)I-labeled-[(131)I]iodo-17-allylamino-17-demethoxy geldanamycin ([(131)I]iodo-17-AAG) and its biodistribution in mice.

Science.gov (United States)

Daozhen, Chen; Lu, Liu; Min, Yang; Xinyu, Jiang; Ying, Huang

2007-10-01

The aim of this study was to examine the radioiodinating condition of 17-allylamino-17-demethoxy geldanamycin (17-AAG) and observe its biodistribution in the hepatoma cell line HepA tumorearing ICR mice for understanding the possibility of its application in nuclear medicine. [(131)I]iodo-17-AAG was prepared by the reaction of 17-AAG with Na[(131)I] in the presence of hydrogen peroxide. [(131)I]iodo-17-AAG was purified by high-performance liquid chromatography (HPLC). The stability of [(131)I]iodo-17-AAG was measured by thin-layer chromatography (TLC). The distributions in HepA tumor-bearing ICR mice at 0.5, 1, 4, 8, 24, and 48 hours after injection of [(131)I]iodo-17-AAG were measured. Tumor uptake studies were performed in HepA tumor-bearing ICR mice. The labeling yield was over 83%. The radiochemical purity of [(131)I]iodo-17-AAG was 99.6% after purification. The specific activity was greater than 4 Ci/micromol. The labeled compound was stable for at least 120 hours in saline at 4 degrees C. It was initially in blood at 5 minutes with 4.79% of injected dose per g of tissue (%ID/g), and then dropped 0.33% ID/g at 24 hours. The uptake in liver, lung, and kidney at 4.44% ID/g, 2.03% ID/g, and 2.17% ID/g decreased with time, and less than 1% ID/g was measured after 24 hours in those organs. There was rapid tumor uptake, which reached 1.26% ID/g at 0.5 hours, the highest uptake at 8 hours. Yet, the [(131)I]iodo-17-AAG in the contralateral muscle was at a low level during the 48 hours. The tumor-contralateral muscle (T/CM) radioactivity ratio for [(131)I]iodo-17-AAG remained constant at all time points. [(131)I]iodo-17-AAG can be efficiently radiolabeled at high specific activity, purified by HPLC and stored with little radiolysis at this specific activities. [(131)I]iodo-17-AAG is a promising radiopharmaceutical in nuclear medicine, especially for tumor-targeted radionuclide brachytherapy.

17 CFR 240.17Ad-21T - Operational capability in a Year 2000 environment.

Science.gov (United States)

2010-04-01

... 17 Commodity and Securities Exchanges 3 2010-04-01 2010-04-01 false Operational capability in a Year 2000 environment. 240.17Ad-21T Section 240.17Ad-21T Commodity and Securities Exchanges SECURITIES... Company Rules § 240.17Ad-21T Operational capability in a Year 2000 environment. (a) This section applies...

Role of interleukin (IL)-17 and T-helper (Th)17 cells in cancer.

Science.gov (United States)

Song, Yang; Yang, Jian Ming

2017-11-04

Interleukin-17 (IL-17), a pleiotropic proinflammatory cytokine, is reported to be significantly generated by a distinct subset of CD4 + T-cells, upgrading cancer-elicited inflammation and preventing cancer cells from immune surveillance. T-helper (Th)17 cells produced from naive CD4 + T cells have recently been renowned and generally accepted, gaining eminence in cancer studies and playing the effective role in context of cancer. Th17 cells are the main source of IL-17-secreting cells, It was found that other cell types produced this cytokine as well, including Group 3 innate lymphoid cells (ILC3), δγT cells, invariant natural killer T (iNKT) cells, lymphoid-tissue inducer (LTi)-like cells and Natural killer (NK) cells. Th17-associated cytokines give impetus to tumor progression, or inducing angiogenesis and metastasis. This review demonstrates an understanding on how the pro- or antitumor function of Th17 cells and IL-17 may change cancer progression, leading to the appearance of complex and pivotal biologic activities in tumor. Copyright © 2017 Elsevier Inc. All rights reserved.

17Beta-hydroxysteroid dehydrogenase (17beta-HSD) in scleractinian corals and zooxanthellae.

Science.gov (United States)

Blomquist, Charles H; Lima, P H; Tarrant, A M; Atkinson, M J; Atkinson, S

2006-04-01

Steroid metabolism studies have yielded evidence of 17beta-hydroxysteroid dehydrogenase (17beta-HSD) activity in corals. This project was undertaken to clarify whether there are multiple isoforms of 17beta-HSD, whether activity levels vary seasonally, and if zooxanthellae contribute to activity. 17Beta-HSD activity was characterized in zooxanthellate and azooxanthellate coral fragments collected in summer and winter and in zooxanthellae cultured from Montipora capitata. More specifically, 17beta-HSD activity was characterized with regard to steroid substrate and inhibitor specificity, coenzyme specificity, and Michaelis constants for estradiol (E2) and NADP+. Six samples each of M. capitata and Tubastrea coccinea (three summers, three winters) were assayed with E2 and NADP+. Specific activity levels (pmol/mg protein) varied 10-fold among M. capitata samples and 6-fold among T. coccinea samples. There was overlap of activity levels between summer and winter samples. NADP+/NAD+ activity ratios varied from 1.6 to 22.2 for M. capatita, 2.3 to 3.8 for T. coccinea and 0.7 to 1.1 for zooxanthellae. Coumestrol was the most inhibitory of the steroids and phytoestrogens tested. Our data confirm that corals and zooxanthellae contain 17beta-HSD and are consistent with the presence of more than one isoform of the enzyme.

17 CFR 240.17a-7 - Records of non-resident brokers and dealers.

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2010-04-01

... brokers and dealers. 240.17a-7 Section 240.17a-7 Commodity and Securities Exchanges SECURITIES AND... Stabilizing Activities § 240.17a-7 Records of non-resident brokers and dealers. (a)(1) Except as provided in paragraphs (b) and (c) of this section, each non-resident broker or dealer registered or applying for...

Inhibitory effects of 131I labeled 17-allylamino-17-demethoxygeldanamycin on breast cancer cell line

International Nuclear Information System (INIS)

Chen Daozhen; Liu Lu; Jiang Xinyu; Huang Ying; Yang Min; Yu Huixin; Luo Shineng; Lin Xiufeng

2007-01-01

Objective: 17-allylamino-17-demethoxygeldanamycin(17-AAG) is a less toxic analogue of geldanamycin (GA) that retains the tumoricidal features of GA. Same as its parent compound, 17-AAG inhibits several signaling pathways through binding to heat shock protein (HSP) 90, which results in destabilization of signaling complexes and degradation of client proteins in a variety of tumor cell growth. Treatment with 17-AAG was effective to inhibit tumor growth and induce apoptosis in colon cancer, glioblastoma, and breast cancer cell lines. This study aimed at exploring the anti-proliferation effects and mechanism of 131 I labeled 17-AAG on human breast cancer cell line MCF-7. Methods: 131 I-17-AAG was prepared by the reaction of 17-AAG with Na 131 I in the presence of hydrogen peroxide. The MCF-7 cells were divided into 5 groups with different additional drugs: group A, dimethyl sulfoxide (DMSO); group B, 370 kBq Na 131 I; group C, 2.5 mg/L 17-AAG; group D, 370 kBq 131 I-17-AAG; group E, 370 kBq 131 I-17-AAG + 2.5 mg/L 17-AAG. 3- (4,5-dimethylthiazol-2-yl)-2,5, diphenylte-trazolium bromide (MTT) assay was used to evaluate the effect of growth inhibition of MCF-7 cells. Cell cycle and apoptosis were analyzed by flow cytometry. The change of the expression of Akt2 mRNA in MCF-7 cells was examined by RT-PCR. Results: The labeling yield of 131 I-17-AAG was 83%. The radiochemical purity of 131 I-17-AAG after purification was 96.6%. The specific activity was 1.48 x 10 5 MBq/μmol. All drugs could significantly inhibit the growth of MCF-7 cells in vitro as the duration lasts longer, especially for group E. After 48 h, sub-G1 peaks detected by flow cytometry were(1.54±0.13)%, (5.72±1.05)%, (12.97±1.44)%, (20.65±1.36)%, (35.39±4.15)% for group A, B, C, D and E, respectively. The experimental groups (B-E) were all significantly higher than the control group (A, all P 131 I-17-AAG could suppress the growth of human breast cancer cell line MCF-7 and hasten the apoptosis. It could

IL-17 for therapy.

Science.gov (United States)

Kurschus, Florian C; Moos, Sonja

2017-09-01

The cytokine IL-17 is now a target for an array of therapeutic monoclonal antibodies supposed to treat a variety of inflammatory diseases. The forerunner Secukinumab, an IL-17A neutralizing antibody, is meanwhile approved as first-line treatments for moderate-to-severe plaque psoriasis, and as second-line treatment for psoriatic arthritis and ankylosing spondylitis. Ixekizumab and Brodalumab, both also targeting the IL-17 pathway, were also recently approved by the FDA for plaque psoriasis. Using mice overexpressing IL-17A in a tissue of choice, we showed that the ectopic expression of this cytokine in keratinocytes resulted in a spontaneous and very strong form of psoriasis-like dermatitis. Interestingly, this model showed some typical comorbidities found in humans with psoriasis. In this review, we will discuss why IL-17 is a good target especially in psoriasis and what we learned from mouse models about its functions in pathological situations. Copyright © 2017 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.

Hsp90 inhibitor 17-allylamino-17-demethoxygeldanamycin inhibits the proliferation of ARPE-19 cells

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Wang Lin

2010-04-01

Full Text Available Abstract Background The antiproliferative effect of the Hsp90 inhibitor 17-AAG (17-allylamino-17-demethoxygeldanamycin on human retinal pigment epithelial cells is investigated. Methods MTT and flow cytometry were used to study the antiproliferative effects of the 17-AAG treatment of ARPE-19 cells. 2D gel electrophoresis (2-DE and mass spectrometry were applied to detect the altered expression of proteins, which was verified by real-time PCR. Gene Ontology analysis and Ingenuity Pathway Analysis (IPA were utilized to analyze the signaling pathways, cellular location, function, and network connections of the identified proteins. And SOD assay was employed to confirm the analysis. Results 17-AAG suppressed the proliferation of ARPE-19 cells by inducing cell cycle arrest and apoptosis. Proteomic analysis revealed that the expression of 94 proteins was altered by a factor of more than 1.5 following exposure to 17-AAG. Of these 94, 87 proteins were identified. Real-time PCR results indicated that Hsp90 and Hsp70, which were not identified by proteomic analysis, were both upregulated upon 17-AAG treatment. IPA revealed that most of the proteins have functions that are related to oxidative stress, as verified by SOD assay, while canonical pathway analysis revealed glycolysis/gluconeogenesis. Conclusions 17-AAG suppressed the proliferation of ARPE-19 cells by inducing cell cycle arrest and apoptosis, and possibly by oxidative stress.

17 CFR 240.17h-2T - Risk assessment reporting requirements for brokers and dealers.

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2010-04-01

... requirements for brokers and dealers. 240.17h-2T Section 240.17h-2T Commodity and Securities Exchanges... Organizations § 240.17h-2T Risk assessment reporting requirements for brokers and dealers. (a) Reporting requirements of risk assessment information required to be maintained by section 240.17h-1T. (1) Every broker...

Relationship between female genital tract infections, mucosal interleukin-17 production and local T helper type 17 cells.

Science.gov (United States)

Masson, Lindi; Salkinder, Amy L; Olivier, Abraham Jacobus; McKinnon, Lyle R; Gamieldien, Hoyam; Mlisana, Koleka; Scriba, Thomas J; Lewis, David A; Little, Francesca; Jaspan, Heather B; Ronacher, Katharina; Denny, Lynette; Abdool Karim, Salim S; Passmore, Jo-Ann S

2015-12-01

T helper type 17 (Th17) cells play an important role in immunity to fungal and bacterial pathogens, although their role in the female genital tract, where exposure to these pathogens is common, is not well understood. We investigated the relationship between female genital tract infections, cervicovaginal interleukin-17 (IL-17) concentrations and Th17 cell frequencies. Forty-two cytokines were measured in cervicovaginal lavages from HIV-uninfected and HIV-infected women. Frequencies of Th17 cells (CD3(+) CD4(+) IL-17a(+)) were evaluated in cervical cytobrushes and blood by flow cytometry. Women were screened for Chlamydia trachomatis, Neisseria gonorrhoeae, Mycoplasma genitalium, Trichomonas vaginalis and herpes simplex virus 2 by PCR, and candidal infections and bacterial vaginosis by Gram stain. Women with bacterial sexually transmitted infections (STIs), specifically chlamydia and gonorrhoea, had higher genital IL-17 concentrations than women with no STI, whereas women with candidal pseudohyphae/spores had lower IL-17 concentrations compared with women without candidal infections. Viral STIs (herpes simplex virus 2 and HIV) were not associated with significant changes in genital IL-17 concentrations. Genital IL-17 concentrations correlated strongly with other inflammatory cytokines and growth factors. Although Th17 cells were depleted from blood during HIV infection, cervical Th17 cell frequencies were similar in HIV-uninfected and HIV-infected women. Cervical Th17 cell frequencies were also not associated with STIs or candida, although few women had a STI. These findings suggest that IL-17 production in the female genital tract is induced in response to bacterial but not viral STIs. Decreased IL-17 associated with candidal infections suggests that candida may actively suppress IL-17 production or women with dampened IL-17 responses may be more susceptible to candidal outgrowth. © 2015 John Wiley & Sons Ltd.

MAP17, a ROS-dependent oncogene

International Nuclear Information System (INIS)

Carnero, Amancio

2012-01-01

MAP17 is a small 17 kDa non-glycosylated membrane protein previously identified as being overexpressed in carcinomas. Breast tumor cells that overexpress MAP17 show an increased tumoral phenotype with enhanced proliferative capabilities both in the presence or the absence of contact inhibition, decreased apoptotic sensitivity, and increased migration. MAP17-expressing clones also grow better in nude mice. The increased malignant cell behavior induced by MAP17 is associated with an increase in reactive oxygen species (ROS) production, and the treatment of MAP17-expressing cells with antioxidants results in a reduction in the tumorigenic properties of these cells. The MAP17-dependent increase in ROS and tumorigenesis relies on its PDZ-binding domain because disruption of this sequence by point mutations abolishes the ability of MAP17 to enhance ROS production and tumorigenesis. MAP17 is overexpressed in a great variety of human carcinomas, including breast tumors. Immunohistochemical analysis of MAP17 during cancer progression demonstrates that overexpression of the protein strongly correlates with tumoral progression. Generalized MAP17 overexpression in human carcinomas indicates that MAP17 can be a good marker for tumorigenesis and, especially, for malignant progression.

Signaling through IL-17C/IL-17RE is dispensable for immunity to systemic, oral and cutaneous candidiasis.

Science.gov (United States)

Conti, Heather R; Whibley, Natasha; Coleman, Bianca M; Garg, Abhishek V; Jaycox, Jillian R; Gaffen, Sarah L

2015-01-01

Candida albicans is a commensal fungal microbe of the human orogastrointestinal tract and skin. C. albicans causes multiple forms of disease in immunocompromised patients, including oral, vaginal, dermal and disseminated candidiasis. The cytokine IL-17 (IL-17A) and its receptor subunits, IL-17RA and IL-17RC, are required for protection to most forms of candidiasis. The importance of the IL-17R pathway has been observed not only in knockout mouse models, but also in humans with rare genetic mutations that impact generation of Th17 cells or the IL-17 signaling pathway, including Hyper-IgE Syndrome (STAT3 or TYK2 mutations) or IL17RA or ACT1 gene deficiency. The IL-17 family of cytokines is a distinct subclass of cytokines with unique structural and signaling properties. IL-17A is the best-characterized member of the IL-17 family to date, but far less is known about other IL-17-related cytokines. In this study, we sought to determine the role of a related IL-17 cytokine, IL-17C, in protection against oral, dermal and disseminated forms of C. albicans infection. IL-17C signals through a heterodimeric receptor composed of the IL-17RA and IL-17RE subunits. We observed that IL-17C mRNA was induced following oral C. albicans infection. However, mice lacking IL-17C or IL-17RE cleared C. albicans infections in the oral mucosa, skin and bloodstream at rates similar to WT littermate controls. Moreover, these mice demonstrated similar gene transcription profiles and recovery kinetics as WT animals. These findings indicate that IL-17C and IL-17RE are dispensable for immunity to the forms of candidiasis evaluated, and illustrate a surprisingly limited specificity of the IL-17 family of cytokines with respect to systemic, oral and cutaneous Candida infections.

Signaling through IL-17C/IL-17RE Is Dispensable for Immunity to Systemic, Oral and Cutaneous Candidiasis

Science.gov (United States)

Conti, Heather R.; Whibley, Natasha; Coleman, Bianca M.; Garg, Abhishek V.; Jaycox, Jillian R.; Gaffen, Sarah L.

2015-01-01

Candida albicans is a commensal fungal microbe of the human orogastrointestinal tract and skin. C. albicans causes multiple forms of disease in immunocompromised patients, including oral, vaginal, dermal and disseminated candidiasis. The cytokine IL-17 (IL-17A) and its receptor subunits, IL-17RA and IL-17RC, are required for protection to most forms of candidiasis. The importance of the IL-17R pathway has been observed not only in knockout mouse models, but also in humans with rare genetic mutations that impact generation of Th17 cells or the IL-17 signaling pathway, including Hyper-IgE Syndrome (STAT3 or TYK2 mutations) or IL17RA or ACT1 gene deficiency. The IL-17 family of cytokines is a distinct subclass of cytokines with unique structural and signaling properties. IL-17A is the best-characterized member of the IL-17 family to date, but far less is known about other IL-17-related cytokines. In this study, we sought to determine the role of a related IL-17 cytokine, IL-17C, in protection against oral, dermal and disseminated forms of C. albicans infection. IL-17C signals through a heterodimeric receptor composed of the IL-17RA and IL-17RE subunits. We observed that IL-17C mRNA was induced following oral C. albicans infection. However, mice lacking IL-17C or IL-17RE cleared C. albicans infections in the oral mucosa, skin and bloodstream at rates similar to WT littermate controls. Moreover, these mice demonstrated similar gene transcription profiles and recovery kinetics as WT animals. These findings indicate that IL-17C and IL-17RE are dispensable for immunity to the forms of candidiasis evaluated, and illustrate a surprisingly limited specificity of the IL-17 family of cytokines with respect to systemic, oral and cutaneous Candida infections. PMID:25849644

17 CFR 240.17Ad-13 - Annual study and evaluation of internal accounting control.

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2010-04-01

... internal accounting control. 240.17Ad-13 Section 240.17Ad-13 Commodity and Securities Exchanges SECURITIES... Company Rules § 240.17Ad-13 Annual study and evaluation of internal accounting control. (a) Accountant's... accountant concerning the transfer agent's system of internal accounting control and related procedures for...

17 CFR 270.17f-2 - Custody of investments by registered management investment company.

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2010-04-01

... registered management investment company. 270.17f-2 Section 270.17f-2 Commodity and Securities Exchanges....17f-2 Custody of investments by registered management investment company. (a) The securities and similar investments of a registered management investment company may be maintained in the custody of such...

A cytosolic juxtamembrane interface modulates plexin A3 oligomerization and signal transduction.

Directory of Open Access Journals (Sweden)

Rachael Barton

Full Text Available Plexins (plxns are transmembrane (TM receptors involved in the guidance of vascular, lymphatic vessel, and neuron growth as well as cancer metastasis. Plxn signaling results in cytosolic GTPase-activating protein activity, and previous research implicates dimerization as important for activation of plxn signaling. Purified, soluble plxn extracellular and cytosolic domains exhibit only weak homomeric interactions, suggesting a role for the plxn TM and juxtamembrane regions in homooligomerization. In this study, we consider a heptad repeat in the Danio rerio PlxnA3 cytosolic juxtamembrane domain (JM for its ability to influence PlxnA3 homooligomerization in TM-domain containing constructs. Site-directed mutagenesis in conjunction with the AraTM assay and bioluminescent energy transfer (BRET² suggest an interface involving a JM heptad repeat, in particular residue M1281, regulates PlxnA3 homomeric interactions when examined in constructs containing an ectodomain, TM and JM domain. In the presence of a neuropilin-2a co-receptor and semaphorin 3F ligand, disruption to PlxnA3 homodimerization caused by an M1281F mutation is eliminated, suggesting destabilization of the PlxnA3 homodimer in the JM is not sufficient to disrupt co-receptor complex formation. In contrast, enhanced homodimerization of PlxnA3 caused by mutation M1281L remains even in the presence of ligand semaphorin 3F and co-receptor neuropilin-2a. Consistent with this pattern of PlxnA3 dimerization in the presence of ligand and co-receptor, destabilizing mutations to PlxnA3 homodimerization (M1281F are able to rescue motor patterning defects in sidetracked zebrafish embryos, whereas mutations that enhance PlxnA3 homodimerization (M1281L are not. Collectively, our results indicate the JM heptad repeat, in particular residue M1281, forms a switchable interface that modulates both PlxnA3 homomeric interactions and signal transduction.

49 CFR 17.4 - [Reserved

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2010-10-01

... 49 Transportation 1 2010-10-01 2010-10-01 false [Reserved] 17.4 Section 17.4 Transportation Office of the Secretary of Transportation INTERGOVERNMENTAL REVIEW OF DEPARTMENT OF TRANSPORTATION PROGRAMS AND ACTIVITIES § 17.4 [Reserved] ...

Increased Circulating Th17 Cells, Serum IL-17A, and IL-23 in Takayasu Arteritis.

Science.gov (United States)

Misra, Durga Prasanna; Chaurasia, Smriti; Misra, Ramnath

2016-01-01

Introduction. Th17, γδT, NK, and NKT cells in peripheral blood and serum IL-17 and IL-23 in Takayasu arteritis (TA) were measured and correlated with disease activity. Methods. Th17 (anti-CD3APC, CD4PECy7, and IL-17PE), NKT, NK (anti-CD3APC, CD56FITC), and γδT (anti-CD3FITC and γδTCRAPC) cells were enumerated by flow cytometry in peripheral blood of 30 patients with TA (ACR1990 criteria) and 20 healthy controls, serum IL-17 and IL-23 measured by ELISA. Relation with disease activity (NIH criteria, ITAS2010) was analyzed (using nonparametric tests, median with interquartile range). Results. Mean age of patients was 33.47 ± 11.78 years (25 females); mean symptom duration was 7.1 ± 5.3 years. 13 were not on immunosuppressants; 12 were active (ITAS2010 ≥ 4). The percentage of Th17 cells was significantly expanded in TA (patients 2.1 (1.5-3.2) versus controls 0.75 (0.32-1.2); p < 0.0001) with no differences in other cell populations. Serum IL-17 and IL-23 (pg/mL) in patients (6.2 (4.6-8.5) and 15 (14.9-26.5), resp.) were significantly higher (p < 0.001) than controls (3.9 (3.9-7.3) and undetectable median value, resp.). Subgroup analysis revealed no correlation of Th17 cells, serum IL-17, and IL-23 with disease activity or medications, nor any significant difference before and after medication. Conclusions. There is significant expansion of Th17 cells and elevated serum IL-17 and IL-23 levels in TA patients compared to healthy controls.

Signaling through IL-17C/IL-17RE is dispensable for immunity to systemic, oral and cutaneous candidiasis.

Directory of Open Access Journals (Sweden)

Heather R Conti

Full Text Available Candida albicans is a commensal fungal microbe of the human orogastrointestinal tract and skin. C. albicans causes multiple forms of disease in immunocompromised patients, including oral, vaginal, dermal and disseminated candidiasis. The cytokine IL-17 (IL-17A and its receptor subunits, IL-17RA and IL-17RC, are required for protection to most forms of candidiasis. The importance of the IL-17R pathway has been observed not only in knockout mouse models, but also in humans with rare genetic mutations that impact generation of Th17 cells or the IL-17 signaling pathway, including Hyper-IgE Syndrome (STAT3 or TYK2 mutations or IL17RA or ACT1 gene deficiency. The IL-17 family of cytokines is a distinct subclass of cytokines with unique structural and signaling properties. IL-17A is the best-characterized member of the IL-17 family to date, but far less is known about other IL-17-related cytokines. In this study, we sought to determine the role of a related IL-17 cytokine, IL-17C, in protection against oral, dermal and disseminated forms of C. albicans infection. IL-17C signals through a heterodimeric receptor composed of the IL-17RA and IL-17RE subunits. We observed that IL-17C mRNA was induced following oral C. albicans infection. However, mice lacking IL-17C or IL-17RE cleared C. albicans infections in the oral mucosa, skin and bloodstream at rates similar to WT littermate controls. Moreover, these mice demonstrated similar gene transcription profiles and recovery kinetics as WT animals. These findings indicate that IL-17C and IL-17RE are dispensable for immunity to the forms of candidiasis evaluated, and illustrate a surprisingly limited specificity of the IL-17 family of cytokines with respect to systemic, oral and cutaneous Candida infections.

17 CFR 240.17a-19 - Form X-17A-19 Report by national securities exchanges and registered national securities...

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2010-04-01

... national securities exchanges and registered national securities associations of changes in the membership... Certain Stabilizing Activities § 240.17a-19 Form X-17A-19 Report by national securities exchanges and registered national securities associations of changes in the membership status of any of their members...

7 CFR 1215.17 - Research.

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2010-01-01

... 7 Agriculture 10 2010-01-01 2010-01-01 false Research. 1215.17 Section 1215.17 Agriculture... AND ORDERS; MISCELLANEOUS COMMODITIES), DEPARTMENT OF AGRICULTURE POPCORN PROMOTION, RESEARCH, AND CONSUMER INFORMATION Popcorn Promotion, Research, and Consumer Information Order Definitions § 1215.17...

7 CFR 1218.17 - Promotion.

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2010-01-01

... 7 Agriculture 10 2010-01-01 2010-01-01 false Promotion. 1218.17 Section 1218.17 Agriculture... AND ORDERS; MISCELLANEOUS COMMODITIES), DEPARTMENT OF AGRICULTURE BLUEBERRY PROMOTION, RESEARCH, AND INFORMATION ORDER Blueberry Promotion, Research, and Information Order Definitions § 1218.17 Promotion...

A comparative study of via drilling and scribing on PEN and PET substrates for flexible electronic applications using excimer and Nd:YAG laser sources

NARCIS (Netherlands)

Mandamparambil, R.; Fledderus, H.; Brand, J. van den; Saalmink, M.; Kusters, R.; Podprocky, T.; Steenberge, G. van; Baets, J. de; Dietzel, A.H.

2009-01-01

A study on via drilling and channel scribing on PEN and PET substrates for flexible electronic application is discussed in this paper. For the experiments, both KIF excimer laser (248 nm) and frequency tripled Nd:YAG (355 nm) laser are used. Different measurement techniques like optical microscopy,

In vivo control mechanisms of motor-cargo movement on microtubules

Science.gov (United States)

Gunawardena, Shermali

2014-03-01

Within axons, molecular motors transport essential components required for neuronal growth and viability. Although many levels of regulation must exist for proper anterograde and retrograde transport of vital proteins, little is known about these mechanisms. Previous work suggested that the amyloid precursor protein (APP) functions as a kinesin-1 receptor during transport. However, how APP vesicle motility is regulated is unclear. Using genetics and in vivo imaging in Drosophila we showed that reduction of presenilin (PS) substantially increased anterograde and retrograde APP vesicle velocities. Strikingly, PS deficiency had no effect on an unrelated cargo vesicle containing synaptotagmin, which is powered by a different kinesin motor. Increased PS-mediated velocities required functional kinesin-1 and dynein motors. We also found that these PS-mediated effects on motor protein function were mediated via a pathway that involves glycogen synthase kinase-3 β (GSK-3 β) . PS genetically interacted with GSK-3 β in an activity dependent manner. Excess of active GSK-3 β perturbed transport by causing axonal blockages, which were enhanced by reduction of kinesin-1 or dynein, while excess of non-functional GSK-3 β had no effect. Strikingly, GSK-3 β-activity dependent transport defects were enhanced by reduction of PS. Collectively, our findings suggest that PS and GSK-3 β are required for normal motor protein function, and we propose a model in which PS likely regulates GSK-3 β activity during transport. These findings have important implications for our understanding of the complex regulatory machinery that must exist in vivo and how this system is coordinated during vesicle motility on microtubules.

Limited replication of yellow fever 17DD and 17D-Dengue recombinant viruses in rhesus monkeys

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Gisela F. Trindade

2008-06-01

Full Text Available For the development of safe live attenuated flavivirus vaccines one of the main properties to be established is viral replication. We have used real-time reverse transcriptase-polymerase chain reaction and virus titration by plaque assay to determine the replication of yellow fever 17DD virus (YFV 17DD and recombinant yellow fever 17D viruses expressing envelope proteins of dengue virus serotypes 2 and 4 (17D-DENV-2 and 17D-DENV-4. Serum samples from rhesus monkeys inoculated with YFV 17DD and 17D-DENV chimeras by intracerebral or subcutaneous route were used to determine and compare the viremia induced by these viruses. Viral load quantification in samples from monkeys inoculated by either route with YFV 17DD virus suggested a restricted capability of the virus to replicate reaching not more than 2.0 log10 PFU mL-1 or 3.29 log10 copies mL-1. Recombinant 17D-dengue viruses were shown by plaquing and real-time PCR to be as attenuated as YF 17DD virus with the highest mean peak titer of 1.97 log10 PFU mL-1 or 3.53 log10 copies mL-1. These data serve as a comparative basis for the characterization of other 17D-based live attenuated candidate vaccines against other diseases.Uma das principais propriedades a serem estabelecidas para o desenvolvimento de vacinas seguras e atenuadas de flavivirus,é a taxa de replicação viral. Neste trabalho, aplicamos a metodologia de amplificação pela reação em cadeia da polimerase em tempo real e titulação viral por plaqueamento para determinação da replicação do vírus 17DD (FA 17DD e recombinantes, expressando proteínas do envelope de dengue sorotipos 2 e 4 (17D-DENV-2 e 17D-DENV-4. As amostras de soros de macacos inoculados por via intracerebral ou subcutânea com FA 17DD ou 17D-DENV foram usadas para determinar e comparar a viremia induzida por estes vírus. A quantificação da carga viral em amostras de macacos inoculados por ambas as vias com FA 17DD sugere restrita capacidade de replicação com

44 CFR 17.605 - Definitions.

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2010-10-01

... 44 Emergency Management and Assistance 1 2010-10-01 2010-10-01 false Definitions. 17.605 Section 17.605 Emergency Management and Assistance FEDERAL EMERGENCY MANAGEMENT AGENCY, DEPARTMENT OF HOMELAND SECURITY GENERAL GOVERNMENTWIDE REQUIREMENTS FOR DRUG-FREE WORKPLACE (GRANTS) § 17.605 Definitions...

48 CFR 17.101 - Authority.

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2010-10-01

... 48 Federal Acquisition Regulations System 1 2010-10-01 2010-10-01 false Authority. 17.101 Section 17.101 Federal Acquisition Regulations System FEDERAL ACQUISITION REGULATION CONTRACTING METHODS AND CONTRACT TYPES SPECIAL CONTRACTING METHODS Multiyear Contracting 17.101 Authority. This subpart implements...

Interplay between microtubule bundling and sorting factors ensures acentriolar spindle stability during C. elegans oocyte meiosis.

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Timothy J Mullen

2017-09-01

Full Text Available In many species, oocyte meiosis is carried out in the absence of centrioles. As a result, microtubule organization, spindle assembly, and chromosome segregation proceed by unique mechanisms. Here, we report insights into the principles underlying this specialized form of cell division, through studies of C. elegans KLP-15 and KLP-16, two highly homologous members of the kinesin-14 family of minus-end-directed kinesins. These proteins localize to the acentriolar oocyte spindle and promote microtubule bundling during spindle assembly; following KLP-15/16 depletion, microtubule bundles form but then collapse into a disorganized array. Surprisingly, despite this defect we found that during anaphase, microtubules are able to reorganize into a bundled array that facilitates chromosome segregation. This phenotype therefore enabled us to identify factors promoting microtubule organization during anaphase, whose contributions are normally undetectable in wild-type worms; we found that SPD-1 (PRC1 bundles microtubules and KLP-18 (kinesin-12 likely sorts those bundles into a functional orientation capable of mediating chromosome segregation. Therefore, our studies have revealed an interplay between distinct mechanisms that together promote spindle formation and chromosome segregation in the absence of structural cues such as centrioles.

44 CFR 17.600 - Purpose.

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2010-10-01

... 44 Emergency Management and Assistance 1 2010-10-01 2010-10-01 false Purpose. 17.600 Section 17.600 Emergency Management and Assistance FEDERAL EMERGENCY MANAGEMENT AGENCY, DEPARTMENT OF HOMELAND SECURITY GENERAL GOVERNMENTWIDE REQUIREMENTS FOR DRUG-FREE WORKPLACE (GRANTS) § 17.600 Purpose. (a) The...

6 CFR 17.540 - Advertising.

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2010-01-01

... 6 Domestic Security 1 2010-01-01 2010-01-01 false Advertising. 17.540 Section 17.540 Domestic... in Employment in Education Programs or Activities Prohibited § 17.540 Advertising. A recipient shall not in any advertising related to employment indicate preference, limitation, specification, or...

7 CFR 922.17 - Container.

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2010-01-01

... 7 Agriculture 8 2010-01-01 2010-01-01 false Container. 922.17 Section 922.17 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... IN WASHINGTON Order Regulating Handling Definitions § 922.17 Container. Container means a box, bag...

7 CFR 923.17 - Container.

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2010-01-01

... 7 Agriculture 8 2010-01-01 2010-01-01 false Container. 923.17 Section 923.17 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... COUNTIES IN WASHINGTON Order Regulating Handling Definitions § 923.17 Container. Container means a box, bag...

Structural Basis of Mec1-Ddc2-RPA Assembly and Activation on Single-Stranded DNA at Sites of Damage.

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Deshpande, Ishan; Seeber, Andrew; Shimada, Kenji; Keusch, Jeremy J; Gut, Heinz; Gasser, Susan M

2017-10-19

Mec1-Ddc2 (ATR-ATRIP) is a key DNA-damage-sensing kinase that is recruited through the single-stranded (ss) DNA-binding replication protein A (RPA) to initiate the DNA damage checkpoint response. Activation of ATR-ATRIP in the absence of DNA damage is lethal. Therefore, it is important that damage-specific recruitment precedes kinase activation, which is achieved at least in part by Mec1-Ddc2 homodimerization. Here, we report a structural, biochemical, and functional characterization of the yeast Mec1-Ddc2-RPA assembly. High-resolution co-crystal structures of Ddc2-Rfa1 and Ddc2-Rfa1-t11 (K45E mutant) N termini and of the Ddc2 coiled-coil domain (CCD) provide insight into Mec1-Ddc2 homodimerization and damage-site targeting. Based on our structural and functional findings, we present a Mec1-Ddc2-RPA-ssDNA composite structural model. By way of validation, we show that RPA-dependent recruitment of Mec1-Ddc2 is crucial for maintaining its homodimeric state at ssDNA and that Ddc2's recruitment domain and CCD are important for Mec1-dependent survival of UV-light-induced DNA damage. Copyright © 2017 Elsevier Inc. All rights reserved.

17 CFR 200.17 - Chief Management Analyst.

Science.gov (United States)

2010-04-01

...) Organizational structures and delegations of authority; (d) Management information systems and concepts; and (e... 17 Commodity and Securities Exchanges 2 2010-04-01 2010-04-01 false Chief Management Analyst. 200...; CONDUCT AND ETHICS; AND INFORMATION AND REQUESTS Organization and Program Management General Organization...

Cytoskeleton Molecular Motors: Structures and Their Functions in Neuron.

Science.gov (United States)

Xiao, Qingpin; Hu, Xiaohui; Wei, Zhiyi; Tam, Kin Yip

2016-01-01

Cells make use of molecular motors to transport small molecules, macromolecules and cellular organelles to target region to execute biological functions, which is utmost important for polarized cells, such as neurons. In particular, cytoskeleton motors play fundamental roles in neuron polarization, extension, shape and neurotransmission. Cytoskeleton motors comprise of myosin, kinesin and cytoplasmic dynein. F-actin filaments act as myosin track, while kinesin and cytoplasmic dynein move on microtubules. Cytoskeleton motors work together to build a highly polarized and regulated system in neuronal cells via different molecular mechanisms and functional regulations. This review discusses the structures and working mechanisms of the cytoskeleton motors in neurons.

48 CFR 17.107 - Options.

Science.gov (United States)

2010-10-01

... 48 Federal Acquisition Regulations System 1 2010-10-01 2010-10-01 false Options. 17.107 Section 17... CONTRACT TYPES SPECIAL CONTRACTING METHODS Multiyear Contracting 17.107 Options. Benefits may accrue by including options in a multiyear contract. In that event, contracting officers must follow the requirements...

9 CFR 352.17 - Transportation.

Science.gov (United States)

2010-01-01

... 9 Animals and Animal Products 2 2010-01-01 2010-01-01 false Transportation. 352.17 Section 352.17 Animals and Animal Products FOOD SAFETY AND INSPECTION SERVICE, DEPARTMENT OF AGRICULTURE AGENCY... CERTIFICATION EXOTIC ANIMALS AND HORSES; VOLUNTARY INSPECTION Exotic Animals § 352.17 Transportation. This shall...

14 CFR 183.17 - Reports.

Science.gov (United States)

2010-01-01

... 14 Aeronautics and Space 3 2010-01-01 2010-01-01 false Reports. 183.17 Section 183.17 Aeronautics... REGULATIONS REPRESENTATIVES OF THE ADMINISTRATOR Certification of Representatives § 183.17 Reports. Each representative designated under this part shall make such reports as are prescribed by the Administrator. ...

6 CFR 17.310 - Recruitment.

Science.gov (United States)

2010-01-01

... 6 Domestic Security 1 2010-01-01 2010-01-01 false Recruitment. 17.310 Section 17.310 Domestic... in Admission and Recruitment Prohibited § 17.310 Recruitment. (a) Nondiscriminatory recruitment. A... recruitment and admission of students. A recipient may be required to undertake additional recruitment efforts...

6 CFR 17.500 - Employment.

Science.gov (United States)

2010-01-01

... 6 Domestic Security 1 2010-01-01 2010-01-01 false Employment. 17.500 Section 17.500 Domestic... in Employment in Education Programs or Activities Prohibited § 17.500 Employment. (a) General. (1) No... subjected to discrimination in employment, or recruitment, consideration, or selection therefore, whether...

34 CFR 303.17 - Multidisciplinary.

Science.gov (United States)

2010-07-01

... 34 Education 2 2010-07-01 2010-07-01 false Multidisciplinary. 303.17 Section 303.17 Education... DISABILITIES General Purpose, Eligibility, and Other General Provisions § 303.17 Multidisciplinary. As used in this part, multidisciplinary means the involvement of two or more disciplines or professions in the...

16 CFR 309.17 - Labels.

Science.gov (United States)

2010-01-01

... 16 Commercial Practices 1 2010-01-01 2010-01-01 false Labels. 309.17 Section 309.17 Commercial... ALTERNATIVE FUELS AND ALTERNATIVE FUELED VEHICLES Requirements for Alternative Fuels Label Specifications § 309.17 Labels. All labels must meet the following specifications: (a) Layout: (1) Non-liquid...

38 CFR 17.801 - Definitions.

Science.gov (United States)

2010-07-01

... 38 Pensions, Bonuses, and Veterans' Relief 1 2010-07-01 2010-07-01 false Definitions. 17.801 Section 17.801 Pensions, Bonuses, and Veterans' Relief DEPARTMENT OF VETERANS AFFAIRS MEDICAL Transitional Housing Loan Program § 17.801 Definitions. (a) Applicant: A non-profit organization making application for...

49 CFR 219.17 - Construction.

Science.gov (United States)

2010-10-01

... 49 Transportation 4 2010-10-01 2010-10-01 false Construction. 219.17 Section 219.17 Transportation Other Regulations Relating to Transportation (Continued) FEDERAL RAILROAD ADMINISTRATION, DEPARTMENT OF TRANSPORTATION CONTROL OF ALCOHOL AND DRUG USE General § 219.17 Construction. Nothing in this part— (a) Restricts...

33 CFR 173.17 - Reciprocity.

Science.gov (United States)

2010-07-01

... 33 Navigation and Navigable Waters 2 2010-07-01 2010-07-01 false Reciprocity. 173.17 Section 173.17 Navigation and Navigable Waters COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) BOATING SAFETY VESSEL NUMBERING AND CASUALTY AND ACCIDENT REPORTING Numbering § 173.17 Reciprocity. (a) Section...

7 CFR 948.17 - Export.

Science.gov (United States)

2010-01-01

... 7 Agriculture 8 2010-01-01 2010-01-01 false Export. 948.17 Section 948.17 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and... Regulating Handling Definitions § 948.17 Export. Export means the shipment of potatoes to any destination...

7 CFR 947.17 - Export.

Science.gov (United States)

2010-01-01

... 7 Agriculture 8 2010-01-01 2010-01-01 false Export. 947.17 Section 947.17 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements and... Definitions § 947.17 Export. Export means shipment of potatoes beyond the boundaries of continental United...

42 CFR 402.17 - Settlement.

Science.gov (United States)

2010-10-01

... 42 Public Health 2 2010-10-01 2010-10-01 false Settlement. 402.17 Section 402.17 Public Health CENTERS FOR MEDICARE & MEDICAID SERVICES, DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL PROVISIONS CIVIL MONEY PENALTIES, ASSESSMENTS, AND EXCLUSIONS General Provisions § 402.17 Settlement. CMS or OIG has...

40 CFR 17.24 - Settlement.

Science.gov (United States)

2010-07-01

... 40 Protection of Environment 1 2010-07-01 2010-07-01 false Settlement. 17.24 Section 17.24... JUSTICE ACT IN EPA ADMINISTRATIVE PROCEEDINGS Procedures for Considering Applications § 17.24 Settlement. A prevailing party and EPA counsel may agree on a proposed settlement of an award before final...

43 CFR 17.201 - Application.

Science.gov (United States)

2010-10-01

... 43 Public Lands: Interior 1 2010-10-01 2010-10-01 false Application. 17.201 Section 17.201 Public Lands: Interior Office of the Secretary of the Interior NONDISCRIMINATION IN FEDERALLY ASSISTED PROGRAMS OF THE DEPARTMENT OF THE INTERIOR Nondiscrimination on the Basis of Handicap § 17.201 Application...

6 CFR 17.510 - Recruitment.

Science.gov (United States)

2010-01-01

... 6 Domestic Security 1 2010-01-01 2010-01-01 false Recruitment. 17.510 Section 17.510 Domestic... in Employment in Education Programs or Activities Prohibited § 17.510 Recruitment. (a) Nondiscriminatory recruitment and hiring. A recipient shall not discriminate on the basis of sex in the recruitment...

7 CFR 958.17 - Container.

Science.gov (United States)

2010-01-01

... 7 Agriculture 8 2010-01-01 2010-01-01 false Container. 958.17 Section 958.17 Agriculture... COUNTIES IN IDAHO, AND MALHEUR COUNTY, OREGON Order Regulating Handling Definitions § 958.17 Container. Container means a sack, box, bag, crate, hamper, basket, carton, package, or any other type of receptacle...

Interleukin (IL)-17A and IL-17F and asthma in Saudi Arabia: mRNA ...

African Journals Online (AJOL)

Win-07

IL17A and IL17F were significantly higher in asthma patients compared to controls [IL17A: 1.112 (2.088) vs 0.938 ... Asthma is a frequently encountered chronic airway in- ... transcription was performed, in 20 μl reaction volume using the.

The role of IL-17 in psoriasis.

Science.gov (United States)

Malakouti, Mona; Brown, Gabrielle Elena; Wang, Eva; Koo, John; Levin, Ethan C

2015-02-01

Psoriasis is a chronic skin condition traditionally believed to involve the Th1 pathway. Recently, the IL-23/Th17/IL-17 pathway has been highlighted in the pathogenesis of psoriasis and other autoimmune inflammatory conditions. From a clinician's perspective, we sought to review the basic science data relevant to IL-17's role in psoriasis pathogenesis. We performed a Pubmed and Web of Knowledge search for English articles starting from 1990 that discussed the Th17 pathway. Search terms such as "IL-17" and "psoriasis" were utilized. The IL-17 pathway is regulated by IL-23, a cytokine that is vital for the expansion and maintenance of the Th17 cell population. Th17 derived cytokines (IL-17A, IL-17F, IL-17A/F and IL-22) were elevated in both psoriasis-like murine models and human psoriatic lesional biopsies. Ixekizumab (anti-IL-17A) treatment of psoriasis was found to normalize levels of IL-17 downstream gene products. Both preclinical and clinical studies support the central role of IL-17 in the pathogenesis of psoriasis.

16 CFR 500.17 - Fractions.

Science.gov (United States)

2010-01-01

... 16 Commercial Practices 1 2010-01-01 2010-01-01 false Fractions. 500.17 Section 500.17 Commercial... LABELING ACT § 500.17 Fractions. (a) SI metric declarations of net quantity of contents of any consumer commodity may contain only decimal fractions. Other declarations of net quantity of contents may contain...

36 CFR 327.17 - Advertisment.

Science.gov (United States)

2010-07-01

... 36 Parks, Forests, and Public Property 3 2010-07-01 2010-07-01 false Advertisment. 327.17 Section 327.17 Parks, Forests, and Public Property CORPS OF ENGINEERS, DEPARTMENT OF THE ARMY RULES AND... § 327.17 Advertisment. (a) Advertising and the distribution of printed matter is allowed within project...

43 CFR 17.502 - Application.

Science.gov (United States)

2010-10-01

... 43 Public Lands: Interior 1 2010-10-01 2010-10-01 false Application. 17.502 Section 17.502 Public Lands: Interior Office of the Secretary of the Interior NONDISCRIMINATION IN FEDERALLY ASSISTED PROGRAMS... Programs or Activities Conducted by the Department of the Interior § 17.502 Application. This part applies...

6 CFR 17.405 - Housing.

Science.gov (United States)

2010-01-01

... 6 Domestic Security 1 2010-01-01 2010-01-01 false Housing. 17.405 Section 17.405 Domestic Security... Education Programs or Activities Prohibited § 17.405 Housing. (a) General. A recipient shall not, on the... different services or benefits related to housing, except as provided in this section (including housing...

43 CFR 17.540 - Employment.

Science.gov (United States)

2010-10-01

... 43 Public Lands: Interior 1 2010-10-01 2010-10-01 false Employment. 17.540 Section 17.540 Public... Programs or Activities Conducted by the Department of the Interior § 17.540 Employment. No qualified handicapped person shall, on the basis of handicap, be subjected to discrimination in employment under any...

43 CFR 17.332 - Mediation.

Science.gov (United States)

2010-10-01

... 43 Public Lands: Interior 1 2010-10-01 2010-10-01 false Mediation. 17.332 Section 17.332 Public..., and Enforcement Procedures § 17.332 Mediation. (a) Referral of complaints for mediation. DOI will... participate in the mediation process to the extent necessary to reach an agreement or make an informed...

Expression of T helper 17 cells and interleukin 17 in lupus nephritis patients

Directory of Open Access Journals (Sweden)

Nayera Z. Saber

2017-07-01

Conclusion: Peripheral blood Th17 cell frequencies and IL-17 in urine are highly linked to LN in SLE and are promising markers of disease activity in LN. Both are valuable targets for future therapeutic applications.

Targeting Th17-IL-17 Pathway in Prevention of Micro-Invasive Prostate Cancer in a Mouse Model.

Science.gov (United States)

Zhang, Qiuyang; Liu, Sen; Ge, Dongxia; Cunningham, David M; Huang, Feng; Ma, Lin; Burris, Thomas P; You, Zongbing

2017-06-01

Chronic inflammation has been associated with the development and progression of human cancers including prostate cancer. The exact role of the inflammatory Th17-IL-17 pathway in prostate cancer remains unknown. In this study, we aimed to determine the importance of Th17 cells and IL-17 in a Pten-null prostate cancer mouse model. The Pten-null mice were treated by Th17 inhibitor SR1001 or anti-mouse IL-17 monoclonal antibody from 6 weeks of age up to 12 weeks of age. For SR1001 treatment, the mice were injected intraperitoneally (i.p.) twice a day with vehicle or SR1001, which was dissolved in a dimethylsulfoxide (DMSO) solution. All mice were euthanized for necropsy at 12 weeks of age. For IL-17 antibody treatment, the mice were injected intravenously (i.v.) once every two weeks with control IgG or rat anti-mouse IL-17 monoclonal antibody, which was dissolved in PBS. The injection time points were at 6, 8, and 10 weeks old. All mice were analyzed for the prostate phenotypes at 12 weeks of age. We found that either SR1001 or anti-IL-17 antibody treatment decreased the formation of micro-invasive prostate cancer in Pten-null mice. The SR1001 or anti-IL-17 antibody treated mouse prostates had reduced proliferation, increased apoptosis, and reduced angiogenesis, as well as reduced inflammatory cell infiltration. By assessing the epithelial-to-mesenchymal transition (EMT) markers, we found that SR1001 or anti-IL-17 antibody treated prostate tissues had weaker EMT phenotype compared to the control treated prostates. These results demonstrated that Th17-IL-17 pathway plays a key role in prostate cancer progression in Pten-null mice. Targeting Th17-IL-17 pathway could prevent micro-invasive prostate cancer formation in mice. Prostate 77:888-899, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

39 CFR 960.17 - Settlement.

Science.gov (United States)

2010-07-01

... 39 Postal Service 1 2010-07-01 2010-07-01 false Settlement. 960.17 Section 960.17 Postal Service... ACT IN POSTAL SERVICE PROCEEDINGS Procedures for Considering Applications § 960.17 Settlement. The applicant and the Postal Service may agree on a proposed settlement of the award before final action on the...

29 CFR 401.17 - Act.

Science.gov (United States)

2010-07-01

... 29 Labor 2 2010-07-01 2010-07-01 false Act. 401.17 Section 401.17 Labor Regulations Relating to Labor OFFICE OF LABOR-MANAGEMENT STANDARDS, DEPARTMENT OF LABOR LABOR-MANAGEMENT STANDARDS MEANING OF TERMS USED IN THIS SUBCHAPTER § 401.17 Act. Act means the Labor-Management Reporting and Disclosure Act...

IL17eScan: A Tool for the Identification of Peptides Inducing IL-17 Response

Directory of Open Access Journals (Sweden)

Sudheer Gupta

2017-10-01

Full Text Available IL-17 cytokines are pro-inflammatory cytokines and are crucial in host defense against various microbes. Induction of these cytokines by microbial antigens has been investigated in the case of ischemic brain injury, gingivitis, candidiasis, autoimmune myocarditis, etc. In this study, we have investigated the ability of amino acid sequence of antigens to induce IL-17 response using machine-learning approaches. A total of 338 IL-17-inducing and 984 IL-17 non-inducing peptides were retrieved from Immune Epitope Database. 80% of the data were randomly selected as training dataset and rest 20% as validation dataset. To predict the IL-17-inducing ability of peptides/protein antigens, different sequence-based machine-learning models were developed. The performance of support vector machine (SVM and random forest (RF was compared with different parameters to predict IL-17-inducing epitopes (IIEs. The dipeptide composition-based SVM-model displayed an accuracy of 82.4% with Matthews correlation coefficient = 0.62 at polynomial (t = 1 kernel on 10-fold cross-validation and outperformed RF. Amino acid residues Leu, Ser, Arg, Asn, and Phe and dipeptides LL, SL, LK, IL, LI, NL, LR, FK, SF, and LE are abundant in IIEs. The present tool helps in the identification of IIEs using machine-learning approaches. The induction of IL-17 plays an important role in several inflammatory diseases, and identification of such epitopes would be of great help to the immunologists. It is freely available at http://metagenomics.iiserb.ac.in/IL17eScan/ and http://metabiosys.iiserb.ac.in/IL17eScan/.

40 CFR 22.17 - Default.

Science.gov (United States)

2010-07-01

... 40 Protection of Environment 1 2010-07-01 2010-07-01 false Default. 22.17 Section 22.17 Protection... Procedures § 22.17 Default. (a) Default. A party may be found to be in default: after motion, upon failure to... hearing. Default by respondent constitutes, for purposes of the pending proceeding only, an admission of...

42 CFR 86.17 - Nondiscrimination.

Science.gov (United States)

2010-10-01

... 42 Public Health 1 2010-10-01 2010-10-01 false Nondiscrimination. 86.17 Section 86.17 Public Health PUBLIC HEALTH SERVICE, DEPARTMENT OF HEALTH AND HUMAN SERVICES OCCUPATIONAL SAFETY AND HEALTH RESEARCH AND RELATED ACTIVITIES GRANTS FOR EDUCATION PROGRAMS IN OCCUPATIONAL SAFETY AND HEALTH Occupational Safety and Health Training Grants § 86.17...

22 CFR 17.3 - Fault.

Science.gov (United States)

2010-04-01

... 22 Foreign Relations 1 2010-04-01 2010-04-01 false Fault. 17.3 Section 17.3 Foreign Relations...) § 17.3 Fault. A recipient of an overpayment is without fault if he or she performed no act of... agency may have been at fault in initiating an overpayment will not necessarily relieve the individual...

22 CFR 120.17 - Export.

Science.gov (United States)

2010-04-01

... 22 Foreign Relations 1 2010-04-01 2010-04-01 false Export. 120.17 Section 120.17 Foreign Relations DEPARTMENT OF STATE INTERNATIONAL TRAFFIC IN ARMS REGULATIONS PURPOSE AND DEFINITIONS § 120.17 Export. (a) Export means: (1) Sending or taking a defense article out of the United States in any manner, except by...

17 CFR 170.10 - Proficiency examinations (sections 4p and 17(p) of the Act).

Science.gov (United States)

2010-04-01

... 17 Commodity and Securities Exchanges 1 2010-04-01 2010-04-01 false Proficiency examinations (sections 4p and 17(p) of the Act). 170.10 Section 170.10 Commodity and Securities Exchanges COMMODITY... examinations (sections 4p and 17(p) of the Act). A futures association may prescribe different training...

Labeling method of 17-allylamino, 17-demethoxygeldanamycin with 131I and its biodistribution in experimental animals

International Nuclear Information System (INIS)

Jiang Xinyu; Liu Lu; Gao Wen; Chen Daozhen; Huang Ying; Yang Min; Luo Shineng

2008-01-01

Objective: The aims of the study were to find out the optimal 131 I labeling method with 17-allylamino, 17-demethoxygeldanamycin (17-AAG) and also to study its biodistribution in animals. Methods: 131 I-17-AAG was prepared by the reaction of 17-AAG with Na 131 I in the presence of hydrogen peroxide. The labeling efficiency and the stability of 131 I-17-AAG were measured by paper chromatograph. The biodistribution in the ICR normal mice was observed by the blood samplings and major organs that were taken out from mice at 0.5, 1, 4, 8, 24 h after 131 I-17-AAG injection through tail veins. VX2 tumor was also implanted in rabbit liver for in vivo imaging with SPECT. Results: The optimal labeling conditions of 17-AAG with mi were determined. The labeling efficiency was 85.65%. The radiochemical purity of 131 I- 17-AAG in acetoacetate solution was (96.51 ± 0.80)% after purification and its radiochemical purity in normal saline solution was (95.57 ± 0.09)%. The radiochemical purity could keep to 90% in normal saline after 5 d at 4 degree C. The biodistribution study in normal mice showed that the uptake (percentage activity of injection dose per gram of tissue, % ID/g) in liver and kidney was less than that in cholecyst [(3.0963 ± 1.3394) %ID/g] at 0.5 h post-injection, and the uptake in stomach and intestine reached to the highest level at 4 h post-injection. The SPECT images showed that the 131 I-17-AAG was obviously concentrated in the tumor after injection at 2 h and 4 d, 6 d, 14 d with the highest tumor to non-tumor (T/NT) radioactivity ratio of 10.36. Conclusions: The labeling method of 17-AAG with 131 I was successfully established. The 131 I-17-AAG in normal saline had a good stability. The main biodistribution in mice was in digestive system and was excreted through the intestinal tract. The SPECT images showed that 131 I-17-AAG might be a potential target-directed agent to the tumor. (authors)

4 CFR 83.17 - Fees.

Science.gov (United States)

2010-01-01

... 4 Accounts 1 2010-01-01 2010-01-01 false Fees. 83.17 Section 83.17 Accounts GOVERNMENT ACCOUNTABILITY OFFICE RECORDS PRIVACY PROCEDURES FOR PERSONNEL RECORDS § 83.17 Fees. (a) Generally, GAO's policy... discretion may charge a fee when the cost for copying the record (at a rate of 20 cents per page) would be in...

21 CFR 17.23 - Discovery.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 1 2010-04-01 2010-04-01 false Discovery. 17.23 Section 17.23 Food and Drugs FOOD... HEARINGS § 17.23 Discovery. (a) No later than 60 days prior to the hearing, unless otherwise ordered by the..., depositions, and any forms of discovery, other than those permitted under paragraphs (a) and (e) of this...

50 CFR 17.94 - Critical habitats.

Science.gov (United States)

2010-10-01

... habitats. (a) The areas listed in § 17.95 (fish and wildlife) and § 17.96 (plants) and referred to in the... physical constituent elements within the defined area of Critical Habitat that are essential to the... 50 Wildlife and Fisheries 2 2010-10-01 2010-10-01 false Critical habitats. 17.94 Section 17.94...

49 CFR 1242.17 - Signals and interlockers (accounts XX-17-19 and XX-18-19).

Science.gov (United States)

2010-10-01

... 49 Transportation 9 2010-10-01 2010-10-01 false Signals and interlockers (accounts XX-17-19 and XX... RAILROADS 1 Operating Expenses-Way and Structures § 1242.17 Signals and interlockers (accounts XX-17-19 and XX-18-19). Separate common expenses on the basis of the total train-hours in running service, and/or...

16,17-Seco- and 2,3:16,17-di-seco-pregnanes from Guarea guidonia

Energy Technology Data Exchange (ETDEWEB)

Garcez, Walmir Silvan; Garcez, Fernanda Rodrigues [Universidade Federal de Mato Grosso do Sul (UFMS), Campo Grande, MS (Brazil). Dept. de Quimica]. E-mail: wgarcez@nin.ufms.br; Soares, Luzinatia Ramos [Universidade Federal de Mato Grosso do Sul (UFMS), Campo Grande, MS (Brazil). Dept. de Quimica; Universidade Estadual de Mato Grosso do Sul, Campo Grande, MS (Brazil)

2008-07-01

Two new seco- and di-seco-pregnanes, 2{alpha},3{beta}-dihydroxy-16,17-seco-pregn-17-ene-16-oic acid methyl ester 2{beta},19-hemiketal (1) and 2,3:16,17-di-seco-pregn-17-ene-3-oic acid-16-oic acid methyl ester-19-hydroxy-2-carboxylic acid-2,19-lactone (2), have been obtained from the trunk bark of Guarea guidonia. Their structures have been established by a combination of 1D- and 2D-NMR spectroscopic techniques and MS data. The unique seco- and di-seco-pregnane carbocyclic skeletal types as found in compounds 1 and 2 are being reported in the Meliaceae for the first time as well as the occurrence of pregnanes in the genus Guarea. (author)

17th International Conference on Textures of Materials (ICOTOM 17)

International Nuclear Information System (INIS)

Skrotzki, Werner; Oertel, Carl-Georg

2015-01-01

The 17th International Conference on Textures of Materials (ICOTOM 17) took place in Dresden, Germany, August 24-29, 2014. It belongs to the 'triennial' series of ICOTOM meetings with a long tradition, starting in 1969 - Clausthal, 1971 - Cracow, 1973 - Pont-à-Mousson, 1975 - Cambridge, 1978 - Aachen, 1981 - Tokyo, 1984 - Noordwijkerhout, 1987 - Santa Fe, 1990 - Avignon, 1993 - Clausthal, 1996 - Xian, 1999 - Montreal, 2002 - Seoul, 2005 - Leuven, 2008 - Pittsburgh, 2011 - Mumbai, 2014 - Dresden. ICOTOM 17 was hosted by the Dresden University of Technology, Institute of Structural Physics. Following the tradition of the ICOTOM conferences, the main focus of ICOTOM-17 was to promote and strengthen the fundamental understanding of the basic processes that govern the formation of texture and its relation to the properties of polycrystalline materials. Nonetheless, it was the aim to forge links between basic research on model materials and applied research on engineering materials of technical importance. Thus, ICOTOM 17 provided a forum for the presentation and discussion of recent progress in research of texture and related anisotropy of mechanical and functional properties of all kinds of polycrystalline materials including natural materials like rocks. Particular attention was paid to recent advances in texture measurement and analysis as well as modeling of texture development for all kinds of processes like solidification, plastic deformation, recrystallization and grain growth, phase transformations, thin film deposition, etc. Hence, ICOTOM 17 was of great interest to materials scientists, engineers from many different areas and geoscientists. The topics covered by ICOTOM 17 were: 1. Mathematical, numerical and statistical methods of texture analysis 2. Deformation textures 3. Crystallization, recrystallization and growth textures 4. Transformation textures 5. Textures in functional materials 6. Textures in advanced materials 7. Textures in rocks 8

50 CFR 17.7 - Raptor exemption.

Science.gov (United States)

2010-10-01

... 50 Wildlife and Fisheries 2 2010-10-01 2010-10-01 false Raptor exemption. 17.7 Section 17.7....7 Raptor exemption. (a) The prohibitions found in §§ 17.21 and 17.31 do not apply to any raptor [a... permittee's possession on November 10, 1978, or as the progeny of such a raptor. (b) This section does not...

On the anisotropy energies for YCo5, RCo5, Y2Co17, and R2Co17

International Nuclear Information System (INIS)

Takahashi, H.; Hikosaka, K.; Ohtsuka, S.; Seo, A.; Ukai, T.; Mori, N.

1988-01-01

The approximate d bands for YCo 5 , RCo 5 , Y 2 Co 17 , and R 2 Co 17 (Th 2 Zn 17 and Th 2 Ni 17 type) are formulated by Deegan's prescription and the formulas of Slater and Koster. The experimental results of YCo 5 and Y 2 Co 17 are discussed by using these approximate d bands. For RCo 5 and R 2 Co 17 the discussions are made by adopting the localized model and the band model for 4f electrons

Fate of 17β-estradiol and 17α-ethinylestradiol in batch and column studies simulating managed aquifer recharge

KAUST Repository

Maeng, Sungkyu

2013-11-01

Laboratory-scale batch and soil columns experiments were conducted to investigate the attenuation of estrogens (17β-estradiol and 17α-ethinylestradiol) during managed aquifer recharge. The role of microbial activity in the removal of selected estrogens was evaluated by comparing the results from biotic and abiotic batch experiments. Moreover, batch experiments were carried out using the sand media prepared over different acclimation periods to investigate the impact of acclimation periods on the removal of selected estrogens. Batch studies showed that adsorption was the dominant removal mechanism in the removal of 17β-estradiol and 17α-ethinylestradiol. 17β-estradiol and 17α-ethinylestradiol were attenuated by 99% and 96%, respectively, in batch experiments under oxic conditions. Redox conditions did not show any significant effect on the attenuation of 17β-estradiol. However, the net estrogenicity of 17β-estradiol remaining was lower under oxic conditions (130 ng estradiol-equivalents/L) than anoxic conditions (970 ng estradiol-equivalents/L) . Column studies operated at 17 h of empty bed contact time also demonstrated that removal mechanism of 17α-ethinylestradiol was more dependent on adsorption than biodegradation. © IWA Publishing 2013.

17-hydroxycorticosteroids

Science.gov (United States)

... level of cortisol. The urine volume and urine creatinine are often done with 17-OHCS test at ... Wisse, MD, Associate Professor of Medicine, Division of Metabolism, Endocrinology & Nutrition, University of Washington School of Medicine, ...

17 CFR 270.17g-1 - Bonding of officers and employees of registered management investment companies.

Science.gov (United States)

2010-04-01

... employees of registered management investment companies. 270.17g-1 Section 270.17g-1 Commodity and... ACT OF 1940 § 270.17g-1 Bonding of officers and employees of registered management investment companies. (a) Each registered management investment company shall provide and maintain a bond which shall...

Characterization of the March 2017 tank 10 surface sample (combination of HTF-10-17-30 AND HTF-10-17-31) and variable depth sample (combination of HTF-10-17-32 and HTF-10-17-33)

Energy Technology Data Exchange (ETDEWEB)

Reboul, S. H. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

2017-07-19

Two surface samples (HTF-10-17-30 and HTF-10-17-31) and two variable depth samples (HTF-10-17-32 and HTF-10-17-33) were collected from SRS Tank 10 during March 2017 and submitted to SRNL for characterization. At SRNL, the two surface samples were combined in one container, the two variable depth samples (VDSs) were combined in another container, and then the two composite samples were each characterized by a series of physical, ionic, radiological, and elemental analysis methods. The surface sample composite was characterized primarily for Tank Farm corrosion control purposes, while the VDS composite was characterized primarily for Tank Closure Cesium Removal (TCCR) purposes.

Comparison of inhibitory effects of 17-AAG nanoparticles and free 17-AAG on HSP90 gene expression in breast cancer.

Science.gov (United States)

Ghalhar, Masoud Gandomkar; Akbarzadeh, Abolfazl; Rahmati, Mohammad; Mellatyar, Hassan; Dariushnejad, Hassan; Zarghami, Nosratallah; Barkhordari, Amin

2014-01-01

HSP90 may be overexpressed in cancer cells which are greatly dependent on Hsp90 function. Geldanamycin derivative 17 allylamino-17-demethoxygeldanamycin (17-AAG) inhibits the function and expression of HSP90. 17-AAG has poor water-solubility which is a potential problem for clinical practice. In this study for improving the stability and solubility of molecules in drug delivery systems we used a β-cyclodextrin- 17AAG complex. To assess cytotoxic effects of β-cyclodextrin-17AAG complexes and free 17AAG, colorimetric cell viability (MTT) assays were performed. Cells were treated with equal concentrations of β-cyclodextrin- 17AAG complex and free 17AAG and Hsp90 gene expression levels in the two groups was compared by real-time PCR. MTT assay confirmed that β-cyclodextrin- 17AAG complex enhanced 17AAG cytotoxicity and drug delivery in T47D breast cancer cells. The level of Hsp90 gene expression in cells treated with β-cyclodextrin- 17AAG complex was lower than that of cells treated with free 17AAG (P=0.001). The results demonstrated that β-cyclodextrin- 17AAG complexes are more effective than free 17AAG in down-regulating HSP90 expression due to enhanced β-cyclodextrin-17AAG uptake by cells. Therefore, β-cyclodextrin could be superior carrier for this kind of hydrophobic agent.

The Myriad Roles of Miro in the Nervous System: Axonal Transport of Mitochondria and Beyond

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Bingwei eLu

2014-10-01

Full Text Available Mitochondrial rho GTPase (Miro is a mitochondrial outer membrane protein containing two GTPase domains and two helix-loop-helix Ca2+-binding domains called EF hands. Pioneering genetic studies in Drosophila first revealed a key function of Miro in regulating the axonal transport of mitochondria, during which Miro forms a multi-protein transport complex with Milton and Kinesin heavy chain (KHC to link trafficking mitochondria with the microtubule cytoskeleton. Recent studies showed that through binding to the EF hands of Miro and causing conformational changes of Miro and alteration of protein-protein interactions within the transport complex, Ca2+ can alter the engagement of mitochondria with the microtubule (MT/kinesin network, offering one mechanism to match mitochondrial distribution with neuronal activity. Despite the importance of the Miro/Milton/Kinesin complex in regulating mitochondrial transport in metazoans, not all components of the transport complex are conserved in lower organisms, and transport-independent functions of Miro are emerging. Here we review the diverse functions of the evolutionarily conserved Miro proteins that are relevant to the development, maintenance, and functioning of the nervous system and discuss the potential contribution of Miro dysfunction to the pathogenesis of diseases of the nervous system.

Atkinesin-13A Modulates Cell-Wall Synthesis and Cell Expansion in Arabidopsis thaliana via the THESEUS1 Pathway

Science.gov (United States)

Fujikura, Ushio; Elsaesser, Lore; Breuninger, Holger; Sánchez-Rodríguez, Clara; Ivakov, Alexander; Laux, Thomas; Findlay, Kim; Persson, Staffan; Lenhard, Michael

2014-01-01

Growth of plant organs relies on cell proliferation and expansion. While an increasingly detailed picture about the control of cell proliferation is emerging, our knowledge about the control of cell expansion remains more limited. We demonstrate here that the internal-motor kinesin AtKINESIN-13A (AtKIN13A) limits cell expansion and cell size in Arabidopsis thaliana, with loss-of-function atkin13a mutants forming larger petals with larger cells. The homolog, AtKINESIN-13B, also affects cell expansion and double mutants display growth, gametophytic and early embryonic defects, indicating a redundant role of the two genes. AtKIN13A is known to depolymerize microtubules and influence Golgi motility and distribution. Consistent with this function, AtKIN13A interacts genetically with ANGUSTIFOLIA, encoding a regulator of Golgi dynamics. Reduced AtKIN13A activity alters cell wall structure as assessed by Fourier-transformed infrared-spectroscopy and triggers signalling via the THESEUS1-dependent cell-wall integrity pathway, which in turn promotes the excess cell expansion in the atkin13a mutant. Thus, our results indicate that the intracellular activity of AtKIN13A regulates cell expansion and wall architecture via THESEUS1, providing a compelling case of interplay between cell wall integrity sensing and expansion. PMID:25232944

Reconstitution of Th17, Tc17 and Treg cells after paediatric haematopoietic stem cell transplantation

DEFF Research Database (Denmark)

Kielsen, Katrine; Ryder, Lars P; Lennox-Hvenekilde, David

2018-01-01

behind these associations have not been investigated previously. We hypothesized that increased levels of IL-7 post-transplant alters the balance between immune-regulatory T cell subsets during the post-transplant lymphocyte recovery towards a more pro-inflammatory profile. We quantified Th17 cells, Tc17.......025). The plasma level of IL-7 at day +90 correlated inversely with Th17 cell counts (rs=-0.65, P=0.0002) and the proportion of Tc17 cells (rs=0.64, P=0.0005) at day +90, but not with Tregs. Furthermore, high IL-7 levels at day +7 were predictive of a less naïve T-cell phenotype at day +90. These findings add...

Mechanistic Scrutiny Identifies a Kinetic Role for Cytochrome b5 Regulation of Human Cytochrome P450c17 (CYP17A1, P450 17A1.

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Alexandr N Simonov

Full Text Available Cytochrome P450c17 (P450 17A1, CYP17A1 is a critical enzyme in the synthesis of androgens and is now a target enzyme for the treatment of prostate cancer. Cytochrome P450c17 can exhibit either one or two physiological enzymatic activities differentially regulated by cytochrome b5. How this is achieved remains unknown. Here, comprehensive in silico, in vivo and in vitro analyses were undertaken. Fluorescence Resonance Energy Transfer analysis showed close interactions within living cells between cytochrome P450c17 and cytochrome b5. In silico modeling identified the sites of interaction and confirmed that E48 and E49 residues in cytochrome b5 are essential for activity. Quartz crystal microbalance studies identified specific protein-protein interactions in a lipid membrane. Voltammetric analysis revealed that the wild type cytochrome b5, but not a mutated, E48G/E49G cyt b5, altered the kinetics of electron transfer between the electrode and the P450c17. We conclude that cytochrome b5 can influence the electronic conductivity of cytochrome P450c17 via allosteric, protein-protein interactions.

DNA-mediated dimerization on a compact sequence signature controls enhancer engagement and regulation by FOXA1.

Science.gov (United States)

Wang, Xuecong; Srivastava, Yogesh; Jankowski, Aleksander; Malik, Vikas; Wei, Yuanjie; Del Rosario, Ricardo C H; Cojocaru, Vlad; Prabhakar, Shyam; Jauch, Ralf

2018-04-14

FOXA1 is a transcription factor capable to bind silenced chromatin to direct context-dependent cell fate conversion. Here, we demonstrate that a compact palindromic DNA element (termed 'DIV' for its diverging half-sites) induces the homodimerization of FOXA1 with strongly positive cooperativity. Alternative structural models are consistent with either an indirect DNA-mediated cooperativity or a direct protein-protein interaction. The cooperative homodimer formation is strictly constrained by precise half-site spacing. Re-analysis of chromatin immunoprecipitation sequencing data indicates that the DIV is effectively targeted by FOXA1 in the context of chromatin. Reporter assays show that FOXA1-dependent transcriptional activity declines when homodimeric binding is disrupted. In response to phosphatidylinositol-3 kinase inhibition DIV sites pre-bound by FOXA1 such as at the PVT1/MYC locus exhibit a strong increase in accessibility suggesting a role of the DIV configuration in the chromatin closed-open dynamics. Moreover, several disease-associated single nucleotide polymorphisms map to DIV elements and show allelic differences in FOXA1 homodimerization, reporter gene expression and are annotated as quantitative trait loci. This includes the rs541455835 variant at the MAPT locus encoding the Tau protein associated with Parkinson's disease. Collectively, the DIV guides chromatin engagement and regulation by FOXA1 and its perturbation could be linked to disease etiologies.

Kinase activation through dimerization by human SH2-B.

Science.gov (United States)

Nishi, Masahiro; Werner, Eric D; Oh, Byung-Chul; Frantz, J Daniel; Dhe-Paganon, Sirano; Hansen, Lone; Lee, Jongsoon; Shoelson, Steven E

2005-04-01

The isoforms of SH2-B, APS, and Lnk form a family of signaling proteins that have been described as activators, mediators, or inhibitors of cytokine and growth factor signaling. We now show that the three alternatively spliced isoforms of human SH2-B readily homodimerize in yeast two-hybrid and cellular transfections assays, and this is mediated specifically by a unique domain in its amino terminus. Consistent with previous reports, we further show that the SH2 domains of SH2-B and APS bind JAK2 at Tyr813. These findings suggested a model in which two molecules of SH2-B or APS homodimerize with their SH2 domains bound to two JAK2 molecules, creating heterotetrameric JAK2-(SH2-B)2-JAK2 or JAK2-(APS)2-JAK2 complexes. We further show that APS and SH2-B isoforms heterodimerize. At lower levels of SH2-B or APS expression, dimerization approximates two JAK2 molecules to induce transactivation. At higher relative concentrations of SH2-B or APS, kinase activation is blocked. SH2-B or APS homodimerization and SH2-B/APS heterodimerization thus provide direct mechanisms for activating and inhibiting JAK2 and other kinases from the inside of the cell and for potentiating or attenuating cytokine and growth factor receptor signaling when ligands are present.

28 CFR 17.26 - Derivative classification.

Science.gov (United States)

2010-07-01

... 28 Judicial Administration 1 2010-07-01 2010-07-01 false Derivative classification. 17.26 Section 17.26 Judicial Administration DEPARTMENT OF JUSTICE CLASSIFIED NATIONAL SECURITY INFORMATION AND ACCESS TO CLASSIFIED INFORMATION Classified Information § 17.26 Derivative classification. (a) Persons...

Th17 Response and Inflammatory Autoimmune Diseases

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Janelle C. Waite

2012-01-01

Full Text Available The proinflammatory activity of T helper 17 (Th17 cells can be beneficial to the host during infection. However, uncontrolled or inappropriate Th17 activation has been linked to several autoimmune and autoinflammatory pathologies. Indeed, preclinical and clinical data show that Th17 cells are associated with several autoimmune diseases such as arthritis, multiple sclerosis, psoriasis, and lupus. Furthermore, targeting the interleukin-17 (IL-17 pathway has attenuated disease severity in preclinical models of autoimmune diseases. Interestingly, a recent report brings to light a potential role for Th17 cells in the autoinflammatory disorder adult-onset Still's disease (AOSD. Whether Th17 cells are the cause or are directly involved in AOSD remains to be shown. In this paper, we discuss the biology of Th17 cells, their role in autoimmune disease development, and in AOSD in particular, as well as the growing interest of the pharmaceutical industry in their use as therapeutic targets.

14N-trinucleon cluster states in 17F and 17O

International Nuclear Information System (INIS)

Merchant, A.C.

1984-01-01

A cluster model is used to calculate the energies of those states in 17 F and 17 O which have a 14 N-trinucleon cluster-core structure. The non-central terms in the cluster-core potential are deduced phenomenologically and also calculated microscopically. They are found to be intimately related to equivalent terms in the potentials for similar cluster-core decompositions of neighbouring nuclei. The results are compared with the spectrum of states excited in a recent experimental study of three-particle transfer onto 14 N. (Author) [pt

44 CFR 17.625 - Exception provision.

Science.gov (United States)

2010-10-01

... 44 Emergency Management and Assistance 1 2010-10-01 2010-10-01 false Exception provision. 17.625 Section 17.625 Emergency Management and Assistance FEDERAL EMERGENCY MANAGEMENT AGENCY, DEPARTMENT OF HOMELAND SECURITY GENERAL GOVERNMENTWIDE REQUIREMENTS FOR DRUG-FREE WORKPLACE (GRANTS) § 17.625 Exception...

17 CFR 240.15c1-7 - Discretionary accounts.

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2010-04-01

... transactions or purchase or sale which are excessive in size or frequency in view of the financial resources... Securities Exchange Act of 1934 Rules Relating to Over-The-Counter Markets § 240.15c1-7 Discretionary...

17 CFR 240.17Ad-15 - Signature guarantees.

Science.gov (United States)

2010-04-01

... Securities Exchange Act of 1934 Supervised Investment Bank Holding Company Rules § 240.17Ad-15 Signature... Securities Exchange Act of 1934; (2) Eligible Guarantor Institution means: (i) Banks (as that term is defined... the transfer agent maintains a list of people authorized to act on behalf of that guarantor...

Inhibition of homologous recombination repair in irradiated tumor cells pretreated with Hsp90 inhibitor 17-allylamino-17-demethoxygeldanamycin

International Nuclear Information System (INIS)

Noguchi, Miho; Yu, Dong; Hirayama, Ryoichi; Ninomiya, Yasuharu; Sekine, Emiko; Kubota, Nobuo; Ando, Koichi; Okayasu, Ryuichi

2006-01-01

In order to investigate the mechanism of radio-sensitization by an Hsp90 inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG), we studied repair of DNA double strand breaks (DSBs) in irradiated human cells pre-treated with 17-AAG. DSBs are thought to be the critical target for radiation-induced cell death. Two human tumor cell lines DU145 and SQ-5 which showed clear radio-sensitization by 17-AAG revealed a significant inhibition of DSB repair, while normal human cells which did not show radio-sensitization by the drug indicated no change in the DSB repair kinetics with 17-AAG. We further demonstrated that BRCA2 was a novel client protein for Hsp90, and 17-AAG caused the degradation of BRCA2 and in turn altered the behavior of Rad51, a critical protein for homologous recombination (HR) pathway of DSB repair. Our data demonstrate for the first time that 17-AAG inhibits the HR repair process and could provide a new therapeutic strategy to selectively result in higher tumor cell killing

7Li(18O, 17N8Be reaction and the 17N + 8Be-potential

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A. T. Rudchik

2010-12-01

Full Text Available Angular distributions of the 7Li(18O, 17N8Be reaction were measured for the transitions to the ground states of 8Be and 17N and excited states of 17N at the energy Elab(18O = 114 MeV. The data were analyzed with coupled-reaction-channels method for one- and two-step transfers of nucleons and clusters. In the analysis, the 7Li + 18O potential de-duced in the analysis of the elastic 7Li + 18O-scattering data as well as shell-model spectroscopic amplitudes of trans-ferred nucleons and clusters were used. Parameters of the 8Be + 17N potential were deduced using the reaction data. Contributions of different one- and two-step transfers in the 7Li(18O, 17N8Be reaction cross-section was studied.

Solvent hold tank sample results for MCU-17-122-124 (March 2017), MCU-17-130-132 (April 2017), MCU-17-133-135 (May 2017), and MCU-17-141-149 (June 2017): Quarterly Report

Energy Technology Data Exchange (ETDEWEB)

Fondeur, F. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL); Jones, D. [Savannah River Site (SRS), Aiken, SC (United States). Savannah River National Lab. (SRNL)

2017-09-13

A trend summary of four Solvent Hold Tank (SHT) monthly samples; MCU-16-122-124 (March 2017), MCU-17-130-132 (April 2017), MCU-17-133-135 (May 2017), and MCU-17-141-149 (June 2017) are reported. Analyses of the June SHT sample (MCU-17-141-149) indicated that the modifier (CS-7SB) and the extractant (MaxCalix) concentrations were slightly below (4% each) their nominal recommended levels (169,000 mg/L and 46,400 mg/L respectively). The suppressor (TiDG) level has decreased since the January 2017 measurement but has remained steady in the range of 666 to 705 mg/L, well above the minimum recommended level (479 mg/L), but below the nominal level. The “flat” trends observed in the TiDG, MaxCalix, modifier, and Gamma measurement are consistent with the solvent being idle since January 10, 2017.

Electron scattering from 17O

International Nuclear Information System (INIS)

Kim, J.C.; Hicks, R.S.; Yen, R.; Auer, I.P.; Caplan, H.S.; Bergstrom, J.C.

1978-01-01

Cross sections for elastic and inelastic scattering of electrons from 17 O have been measured for momentum transfers up to 1.2 fm -1 . The elastic cross section indicates that the rms charge radii of 17 O and 16 O are equal to within a few parts in a thousand: 2 17 >sup(1/2)/ 2 16 >sub(1/2)=1.0015+-0.0025. Reduced transition probabilities and ground-state radiative widths are deduced for 17 O excited states below 9 MeV. Various aspects of the inelastic spectrum are discussed, with emphasis on the 'single-particle' levels at 0.871 (1/2 + ) and 5.083 (3/2 + ) MeV, the levels at 7.569 (7/2 - ) and 7.378 (5/2 + ) MeV, and the spectrum of electric octupole excitations. (Auth.)

29 CFR 1917.17 - Railroad facilities.

Science.gov (United States)

2010-07-01

... 29 Labor 7 2010-07-01 2010-07-01 false Railroad facilities. 1917.17 Section 1917.17 Labor Regulations Relating to Labor (Continued) OCCUPATIONAL SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR (CONTINUED) MARINE TERMINALS Marine Terminal Operations § 1917.17 Railroad facilities. (a) Work shall be...

7 CFR 982.17 - Marketing year.

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2010-01-01

... 7 Agriculture 8 2010-01-01 2010-01-01 false Marketing year. 982.17 Section 982.17 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... WASHINGTON Order Regulating Handling Definitions § 982.17 Marketing year. Marketing year means the 12 months...

47 CFR 13.17 - Replacement license.

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2010-10-01

... 47 Telecommunication 1 2010-10-01 2010-10-01 false Replacement license. 13.17 Section 13.17 Telecommunication FEDERAL COMMUNICATIONS COMMISSION GENERAL COMMERCIAL RADIO OPERATORS General § 13.17 Replacement... request a replacement. The application must be accompanied by the required fee and submitted to the...

7 CFR 1709.17 - Environmental review.

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2010-01-01

... 7 Agriculture 11 2010-01-01 2010-01-01 false Environmental review. 1709.17 Section 1709.17... AGRICULTURE ASSISTANCE TO HIGH ENERGY COST COMMUNITIES General Requirements § 1709.17 Environmental review. (a.... (b) Applicants must address environmental aspects of their projects in the grant application in...

49 CFR 241.17 - Preemptive effect.

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2010-10-01

... 49 Transportation 4 2010-10-01 2010-10-01 false Preemptive effect. 241.17 Section 241.17 Transportation Other Regulations Relating to Transportation (Continued) FEDERAL RAILROAD ADMINISTRATION... OPERATIONS § 241.17 Preemptive effect. Under 49 U.S.C. 20106, the regulations in this part preempt any State...

32 CFR 637.17 - Police Intelligence.

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2010-07-01

... 32 National Defense 4 2010-07-01 2010-07-01 true Police Intelligence. 637.17 Section 637.17... CRIMINAL INVESTIGATIONS MILITARY POLICE INVESTIGATION Investigations § 637.17 Police Intelligence. (a) The purpose of gathering police intelligence is to identify individuals or groups of individuals in an effort...

32 CFR 705.17 - Participation guidelines.

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2010-07-01

... 32 National Defense 5 2010-07-01 2010-07-01 false Participation guidelines. 705.17 Section 705.17 National Defense Department of Defense (Continued) DEPARTMENT OF THE NAVY UNITED STATES NAVY REGULATIONS AND OFFICIAL RECORDS PUBLIC AFFAIRS REGULATIONS § 705.17 Participation guidelines. (a) The provisions...

38 CFR 17.504 - Disclosure methods.

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2010-07-01

... 38 Pensions, Bonuses, and Veterans' Relief 1 2010-07-01 2010-07-01 false Disclosure methods. 17.504 Section 17.504 Pensions, Bonuses, and Veterans' Relief DEPARTMENT OF VETERANS AFFAIRS MEDICAL Confidentiality of Healthcare Quality Assurance Review Records § 17.504 Disclosure methods. (a) Disclosure of...

The 17D-204 and 17DD yellow fever vaccines: an overview of major similarities and subtle differences.

Science.gov (United States)

Ferreira, Clarissa de Castro; Campi-Azevedo, Ana Carolina; Peruhype-Magalhāes, Vanessa; Costa-Pereira, Christiane; Albuquerque, Cleandro Pires de; Muniz, Luciana Feitosa; Yokoy de Souza, Talita; Oliveira, Ana Cristina Vanderley; Martins-Filho, Olindo Assis; da Mota, Licia Maria Henrique

2018-01-01

The yellow fever vaccine is a live attenuated virus vaccine that is considered one of the most efficient vaccines produced to date. The original 17D strain generated the substrains 17D-204 and 17DD, which are used for the current production of vaccines against yellow fever. The 17D-204 and 17DD substrains present subtle differences in their nucleotide compositions, which can potentially lead to variations in immunogenicity and reactogenicity. We will address the main changes in the immune responses induced by the 17D-204 and 17DD yellow fever vaccines and report similarities and differences between these vaccines in cellular and humoral immunity . This is a relevant issue in view of the re-emergence of yellow fever in Uganda in 2016 and in Brazil in the beginning of 2017. Areas covered: This article will be divided into 8 sections that will analyze the innate immune response, adaptive immune response, humoral response, production of cytokines, immunity in children, immunity in the elderly, gene expression and adverse reactions. Expert commentary: The 17D-204 and 17DD yellow fever vaccines present similar immunogenicity, with strong activation of the cellular and humoral immune responses. Additionally, both vaccines have similar adverse effects, which are mostly mild and thus are considered safe.

17DD and 17D-213/77 yellow fever substrains trigger a balanced cytokine profile in primary vaccinated children.

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Ana Carolina Campi-Azevedo

Full Text Available BACKGROUND: This study aimed to compare the cytokine-mediated immune response in children submitted to primary vaccination with the YF-17D-213/77 or YF-17DD yellow fever (YF substrains. METHODS: A non-probabilistic sample of eighty healthy primary vaccinated (PV children was selected on the basis of their previously known humoral immune response to the YF vaccines. The selected children were categorized according to their YF-neutralizing antibody titers (PRNT and referred to as seroconverters (PV-PRNT(+ or nonseroconverters (PV-PRNT(-. Following revaccination with the YF-17DD, the PV-PRNT(- children (YF-17D-213/77 and YF-17DD groups seroconverted and were referred as RV-PRNT(+. The cytokine-mediated immune response was investigated after short-term in vitro cultures of whole blood samples. The results are expressed as frequency of high cytokine producers, taking the global median of the cytokine index (YF-Ag/control as the cut-off. RESULTS: The YF-17D-213/77 and the YF-17DD substrains triggered a balanced overall inflammatory/regulatory cytokine pattern in PV-PRNT(+, with a slight predominance of IL-12 in YF-17DD vaccinees and a modest prevalence of IL-10 in YF-17D-213/77. Prominent frequency of neutrophil-derived TNF-α and neutrophils and monocyte-producing IL-12 were the major features of PV-PRNT(+ in the YF-17DD, whereas relevant inflammatory response, mediated by IL-12(+CD8(+ T cells, was the hallmark of the YF-17D-213/77 vaccinees. Both substrains were able to elicit particular but relevant inflammatory events, regardless of the anti-YF PRNT antibody levels. PV-PRNT(- children belonging to the YF-17DD arm presented gaps in the inflammatory cytokine signature, especially in terms of the innate immunity, whereas in the YF-17D-213/77 arm the most relevant gap was the deficiency of IL-12-producing CD8(+T cells. Revaccination with YF-17DD prompted a balanced cytokine profile in YF-17DD nonresponders and a robust inflammatory profile in YF-17D

Journal of Genetics | Indian Academy of Sciences

Indian Academy of Sciences (India)

Home; Journals; Journal of Genetics. Dimos Kapetis. Articles written in Journal of Genetics. Volume 91 Online resources 2012 pp e18-e23. Gene expression profiling and qRT-PCR expression of RRP1B, PCNT, KIF21A and ADRB2 in leucocytes of Down's syndrome subjects · Michele Salemi Concetta Barone Corrado ...

7 CFR 1738.17 - Civil rights.

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2010-01-01

... 7 Agriculture 11 2010-01-01 2010-01-01 false Civil rights. 1738.17 Section 1738.17 Agriculture Regulations of the Department of Agriculture (Continued) RURAL UTILITIES SERVICE, DEPARTMENT OF AGRICULTURE RURAL BROADBAND ACCESS LOANS AND LOAN GUARANTEES Loan Purposes and Basic Policies § 1738.17 Civil rights...

32 CFR 1602.17 - Military service.

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2010-07-01

... 32 National Defense 6 2010-07-01 2010-07-01 false Military service. 1602.17 Section 1602.17 National Defense Other Regulations Relating to National Defense SELECTIVE SERVICE SYSTEM DEFINITIONS § 1602.17 Military service. The term military service includes service in the Army, the Navy, the Air Force...

Development of top nozzle holddown spring for 17x17 next generation fuel assembly

International Nuclear Information System (INIS)

Lee, J. S.; Lee, S. H.; Kim, H. K.; Lee, J. N.; Jeon, K. R.

2002-01-01

Two conceptual holddown spring designs were developed for 17x17 Next Generation Fuel(NGF) top nozzle. One spring pack concept uses three 0.175 inch thick leaves. The other uses four 0.155 inch thick leaves. The room temperature elastic-plastic properties of each spring pack are calculated using the elastic-plastic model derived from classic beam theory and the exiting spring characteristics test. The stress analysis and spring characteristics of each spring pack are also analyzed using FEM(ANSYS 5.7) to verify the elastic-plastic model. The results of the elastic-plastic model have a good agreement to the results of finite element analysis. It is concluded that the 3-leaf 0.175 inch spring pack concept and 4-leaf 0.155 inch spring pack concept are both viable candidates for 17x17 NGF. A series of load-deflection tests will be used to verify the elastic-plastic model and finite element model

Cytochrome P450c17 (steroid 17α-hydroxylase/17,20 lyase): cloning of human adrenal and testis cDNAs indicates the same gene is expressed in both tissues

International Nuclear Information System (INIS)

Chung, B.; Picado-Leonard, J.; Haniu, M.; Bienkowski, M.; Hall, P.F.; Shively, J.E.; Miller, W.L.

1987-01-01

P450c17 is the single enzyme mediating both 17α-hydroxylase (steroid 17α-monooxygenase, EC 1.14.99.9) and 17,20 lyase activities in the synthesis of steroid hormones. It has been suggested that different P450c17 isozymes mediate these activities in the adrenal gland and testis. The authors sequenced 423 of the 509 amino acids (83%) of the porcine adrenal enzyme; based on this partial sequence, a 128-fold degenerate 17-mer was synthesized and used to screen a porcine adrenal cDNA library. This yielded a 380-base cloned cDNA, which in turn was used to isolate several human adrenal cDNAs. The longest of these, λ hac 17-2, is 1754 base pairs long and includes the full-length coding region, the complete 3'-untranslated region, and 41 bases of the 5'-untranslated region. This cDNA encodes a protein of 508 amino acids having a predicted molecular weight of 57,379.82. High-stringency screening of a human testicular cDNA library yielded a partial clone containing 1303 identical bases. RNA gel blots and nuclease S1-protection experiments confirm that the adrenal and testicular P450c17 mRNAs are indistinguishable. These data indicate that the testis possesses a P450c17 identical to that in the adrenal. The human amino acid sequence is 66.7% homologous to the corresponding regions of the porcine sequence, and the human cDNA and amino acid sequences are 80.1 and 70.3% homologous, respectively, to bovine adrenal P450c17 cDNA. Both comparisons indicate that a central region comprising amino acid residues 160-268 is hypervariable among these species of P450c17

PREFACE: 17th International Conference on Textures of Materials (ICOTOM 17)

Science.gov (United States)

Skrotzki, Werner; Oertel, Carl-Georg

2015-04-01

The 17th International Conference on Textures of Materials (ICOTOM 17) took place in Dresden, Germany, August 24-29, 2014. It belongs to the "triennial" series of ICOTOM meetings with a long tradition, starting in 1969 - Clausthal, 1971 - Cracow, 1973 - Pont-à-Mousson, 1975 - Cambridge, 1978 - Aachen, 1981 - Tokyo, 1984 - Noordwijkerhout, 1987 - Santa Fe, 1990 - Avignon, 1993 - Clausthal, 1996 - Xian, 1999 - Montreal, 2002 - Seoul, 2005 - Leuven, 2008 - Pittsburgh, 2011 - Mumbai, 2014 - Dresden. ICOTOM 17 was hosted by the Dresden University of Technology, Institute of Structural Physics. Following the tradition of the ICOTOM conferences, the main focus of ICOTOM-17 was to promote and strengthen the fundamental understanding of the basic processes that govern the formation of texture and its relation to the properties of polycrystalline materials. Nonetheless, it was the aim to forge links between basic research on model materials and applied research on engineering materials of technical importance. Thus, ICOTOM 17 provided a forum for the presentation and discussion of recent progress in research of texture and related anisotropy of mechanical and functional properties of all kinds of polycrystalline materials including natural materials like rocks. Particular attention was paid to recent advances in texture measurement and analysis as well as modeling of texture development for all kinds of processes like solidification, plastic deformation, recrystallization and grain growth, phase transformations, thin film deposition, etc. Hence, ICOTOM 17 was of great interest to materials scientists, engineers from many different areas and geoscientists. The topics covered by ICOTOM 17 were: 1. Mathematical, numerical and statistical methods of texture analysis 2. Deformation textures 3. Crystallization, recrystallization and growth textures 4. Transformation textures 5. Textures in functional materials 6. Textures in advanced materials 7. Textures in rocks 8. Texture

Targeting IL-17 AND IL-17D receptors of rheumatoid arthritis using phytocompounds: A Molecular Docking study

Science.gov (United States)

Thabitha, A.; Thoufic Ali, A. M. Mohamed; Shweta Kumari, Singh; Rakhi; Swami, Varsha; Mohana Priya, A.; Sajitha Lulu, S.

2017-11-01

Rheumatoid arthritis (RA) is a chronic autoimmune condition of the connective tissue in synovial joints, characterized by inflammation which can lead to bone and cartilage destruction. IL-17 and IL-17D cytokines produced by a number of cell types, primarily promote pro-inflammatory immune responses and negative regulator in fibroblast growth factor signalling. Thus, the promising therapeutic strategies focus on targeting these cytokines, which has led to the identification of effective inhibitors. However, several studies focused on identifying the anti-arthritic potential of natural compounds. Therefore, in the present study we undertook in silico investigations to decipher the anti-inflammatory prospective of phytocompounds by targeting IL-17 and IL-17D cytokines using Patch Dock algorithm. Additionally, IL-17 and IL-17D proteins structure were modelled and validated for molecular docking study. Further, phytocompounds based on anti-inflammatory property were subjected to Lipinski filter and ADMET properties indicated that all of these compounds showed desirable drug-like criteria. The outcome of this investigation sheds light on the anti-inflammatory mechanism of phytocompounds by targeting IL-17 and IL-D for effective treatment of RA.

Meta-analysis identifies seven susceptibility loci involved in the atopic march.

Science.gov (United States)

Marenholz, Ingo; Esparza-Gordillo, Jorge; Rüschendorf, Franz; Bauerfeind, Anja; Strachan, David P; Spycher, Ben D; Baurecht, Hansjörg; Margaritte-Jeannin, Patricia; Sääf, Annika; Kerkhof, Marjan; Ege, Markus; Baltic, Svetlana; Matheson, Melanie C; Li, Jin; Michel, Sven; Ang, Wei Q; McArdle, Wendy; Arnold, Andreas; Homuth, Georg; Demenais, Florence; Bouzigon, Emmanuelle; Söderhäll, Cilla; Pershagen, Göran; de Jongste, Johan C; Postma, Dirkje S; Braun-Fahrländer, Charlotte; Horak, Elisabeth; Ogorodova, Ludmila M; Puzyrev, Valery P; Bragina, Elena Yu; Hudson, Thomas J; Morin, Charles; Duffy, David L; Marks, Guy B; Robertson, Colin F; Montgomery, Grant W; Musk, Bill; Thompson, Philip J; Martin, Nicholas G; James, Alan; Sleiman, Patrick; Toskala, Elina; Rodriguez, Elke; Fölster-Holst, Regina; Franke, Andre; Lieb, Wolfgang; Gieger, Christian; Heinzmann, Andrea; Rietschel, Ernst; Keil, Thomas; Cichon, Sven; Nöthen, Markus M; Pennell, Craig E; Sly, Peter D; Schmidt, Carsten O; Matanovic, Anja; Schneider, Valentin; Heinig, Matthias; Hübner, Norbert; Holt, Patrick G; Lau, Susanne; Kabesch, Michael; Weidinger, Stefan; Hakonarson, Hakon; Ferreira, Manuel A R; Laprise, Catherine; Freidin, Maxim B; Genuneit, Jon; Koppelman, Gerard H; Melén, Erik; Dizier, Marie-Hélène; Henderson, A John; Lee, Young Ae

2015-11-06

Eczema often precedes the development of asthma in a disease course called the 'atopic march'. To unravel the genes underlying this characteristic pattern of allergic disease, we conduct a multi-stage genome-wide association study on infantile eczema followed by childhood asthma in 12 populations including 2,428 cases and 17,034 controls. Here we report two novel loci specific for the combined eczema plus asthma phenotype, which are associated with allergic disease for the first time; rs9357733 located in EFHC1 on chromosome 6p12.3 (OR 1.27; P=2.1 × 10(-8)) and rs993226 between TMTC2 and SLC6A15 on chromosome 12q21.3 (OR 1.58; P=5.3 × 10(-9)). Additional susceptibility loci identified at genome-wide significance are FLG (1q21.3), IL4/KIF3A (5q31.1), AP5B1/OVOL1 (11q13.1), C11orf30/LRRC32 (11q13.5) and IKZF3 (17q21). We show that predominantly eczema loci increase the risk for the atopic march. Our findings suggest that eczema may play an important role in the development of asthma after eczema.

Microstructure and Tensile Behavior of Al8Co17Cr17Cu8Fe17Ni33 (at.%) High-Entropy Alloy

Science.gov (United States)

Daoud, H. M.; Manzoni, A.; Völkl, R.; Wanderka, N.; Glatzel, U.

2013-12-01

Microstructure evolution and tensile behavior of the high-entropy alloy Al8Co17Cr17Cu8Fe17Ni33 (at.%) are investigated at room temperature and at 500°C in the as-cast state and under different heat-treatment conditions. Detailed microstructural characterizations are carried out using optical microscopy, scanning electron microscopy, and transmission electron microscopy. The equilibrium phase evolution as a function of temperature was calculated using the Thermo-Calc software (Thermo-Calc Software, Stockholm, Sweden) integrated with TTNi-7 database. The observed majority phase is a face-centered cubic solid solution for all tested specimens. Tensile ductility at room temperature and at elevated temperature is enhanced by heat treatment at 1150°C. An embrittlement phenomenon has been observed after a heat treatment at 700°C resulting in significant degradation in tensile properties.

Roles of human papillomavirus infection and stathmin in the pathogenesis of sinonasal inverted papilloma.

Science.gov (United States)

Lin, Hai; Lin, Dong; Xiong, Xi-Sheng

2016-02-01

The purpose of this study was to investigate roles of human papillomavirus (HPV) infection and stathmin in sinonasal inverted papilloma (SNIP). HPV DNA detection was performed by the fluorescence-based polymerase chain reaction (PCR) method. Stathmin protein expression was investigated by the immunohistochemistry method and mRNA expression of stathmin, Kif2a, and cyclin D1 were assessed by real-time PCR in SNIP and control subjects. The positive rate of HPV DNA detected in SNIP was about 53.6% (15 of 28). Recurrent cases showed a higher rate of HPV infection compared with initial cases and higher Krouse stage (T3 + T4) cases showed higher rate of HPV infection than lower Krouse stage (T1 + T2) cases. Stronger expression of stathmin, Kif2a, and cyclin D1 were observed in SNIP, especially HPV(+) SNIP. HPV infection was closely associated with recurrence and progression of SNIP. Stathmin is a valuable prognostic marker and could be considered as a therapeutic target in patients with SNIP. © 2015 Wiley Periodicals, Inc.

6 CFR 17.410 - Comparable facilities.

Science.gov (United States)

2010-01-01

... 6 Domestic Security 1 2010-01-01 2010-01-01 false Comparable facilities. 17.410 Section 17.410... the Basis of Sex in Education Programs or Activities Prohibited § 17.410 Comparable facilities. A recipient may provide separate toilet, locker room, and shower facilities on the basis of sex, but such...

7 CFR 7.17 - Dual office.

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2010-01-01

... 7 Agriculture 1 2010-01-01 2010-01-01 false Dual office. 7.17 Section 7.17 Agriculture Office of... STATE, COUNTY AND COMMUNITY COMMITTEES § 7.17 Dual office. (a) County committee membership. A member of... any other county office employee. (b) Community committee membership. A member of the community...

7 CFR 550.17 - Peer review.

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2010-01-01

... 7 Agriculture 6 2010-01-01 2010-01-01 false Peer review. 550.17 Section 550.17 Agriculture... § 550.17 Peer review. Upon request of the REE Agency, cooperators may be requested to provide documentation in support of peer review activities and cooperator personnel may be requested to participate in...

6 CFR 17.525 - Fringe benefits.

Science.gov (United States)

2010-01-01

... 6 Domestic Security 1 2010-01-01 2010-01-01 false Fringe benefits. 17.525 Section 17.525 Domestic... in Employment in Education Programs or Activities Prohibited § 17.525 Fringe benefits. (a) “Fringe benefits” defined. For purposes of these Title IX regulations, the term fringe benefits means any medical...

22 CFR 17.5 - Financial hardship.

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2010-04-01

...) Considerations. Some pertinent considerations in determining whether recovery would cause financial hardship are... 22 Foreign Relations 1 2010-04-01 2010-04-01 false Financial hardship. 17.5 Section 17.5 Foreign... SYSTEM (FSPS) § 17.5 Financial hardship. (a) Waiver of overpayment will not be allowed in any case prior...

17 CFR 240.17d-1 - Examination for compliance with applicable financial responsibility rules.

Science.gov (United States)

2010-04-01

... cooperation and coordination among self-regulatory organizations, and the development of a national market... with applicable financial responsibility rules. 240.17d-1 Section 240.17d-1 Commodity and Securities... financial responsibility rules. (a) Where a member of SIPC is a member of more than one self-regulatory...

14 CFR 73.17 - Controlling agency.

Science.gov (United States)

2010-01-01

... 14 Aeronautics and Space 2 2010-01-01 2010-01-01 false Controlling agency. 73.17 Section 73.17... SPECIAL USE AIRSPACE Restricted Areas § 73.17 Controlling agency. For the purposes of this part, the controlling agency is the FAA facility that may authorize transit through or flight within a restricted area...

43 CFR 17.10 - Judicial review.

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2010-10-01

... 43 Public Lands: Interior 1 2010-10-01 2010-10-01 false Judicial review. 17.10 Section 17.10... Origin § 17.10 Judicial review. Action taken pursuant to section 602 of the act is subject to judicial review as provided in section 603 of the act. [29 FR 16293, Dec. 4, 1964] ...

14 CFR 33.17 - Fire protection.

Science.gov (United States)

2010-01-01

... 14 Aeronautics and Space 1 2010-01-01 2010-01-01 false Fire protection. 33.17 Section 33.17... STANDARDS: AIRCRAFT ENGINES Design and Construction; General § 33.17 Fire protection. (a) The design and... protection unless damage by fire will not cause leakage or spillage of a hazardous quantity of flammable...

7 CFR 51.17 - Official sampling.

Science.gov (United States)

2010-01-01

... 7 Agriculture 2 2010-01-01 2010-01-01 false Official sampling. 51.17 Section 51.17 Agriculture... Inspection Service § 51.17 Official sampling. Samples may be officially drawn by any duly authorized... time and place of the sampling and the brands or other identifying marks of the containers from which...

22 CFR 142.17 - New construction.

Science.gov (United States)

2010-04-01

... 22 Foreign Relations 1 2010-04-01 2010-04-01 false New construction. 142.17 Section 142.17 Foreign... ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE Accessibility § 142.17 New construction. (a) Design and construction. Each facility or part of a facility constructed by, on behalf of, or for the use of a recipient...

Increased protein expression of LHCG receptor and 17a-hydroxylase/17,20-lyase in human polycystic ovaries

NARCIS (Netherlands)

Comim, F.V.; Teerds, K.J.; Hardy, K.; Franks, S.

2013-01-01

STUDY QUESTION Does the expression of LHCG receptor (LHCGR) protein and key enzymes in the androgen biosynthetic pathway differ in normal human versus polycystic ovarian tissue? SUMMARY ANSWER LHCGR and 17a-hydroxylase/17-20-lyase (CYP17A1) protein levels are increased in polycystic ovaries (PCOs).

Reflectometry on D17

Energy Technology Data Exchange (ETDEWEB)

Cubitt, R [Institut Max von Laue - Paul Langevin (ILL), 38 - Grenoble (France)

1997-04-01

As part of the package of instrument upgrades planned over the next few years, D17 is based on a straightened cold neutron-guide and converted into a dedicated and versatile reflectometer. In the meantime, in order for ILL to become as fully involved as possible in this growing area of activity, the current D17 has been optimised for reflectometry. Results of this project are presented. (author).

The Dynamics of Treg/Th17 and the Imbalance of Treg/Th17 in Clonorchis sinensis-Infected Mice

Science.gov (United States)

Hua, Hui; Li, Bo; Zhang, Bo; Yu, Qian; Li, Xiang-Yang; Liu, Ying; Pan, Wei; Liu, Xiang-Ye; Tang, Ren-Xian; Zheng, Kui-Yang

2015-01-01

Clonorchiasis, caused by the liver fluke Clonorchis sinensis, is a chronic parasitic infection regulated by T cell subsets. An imbalance of CD4+CD25+ Foxp3+regulatory T (Treg) and interleukin (IL)-17-secreting T cells (Th17) may control inflammation and play an important role in the pathogenesis of immune evasion. In the present study, we assessed the dynamics of Treg/Th17 and determined whether the Treg/Th17 ratio is altered in C. sinensis-infected mice. The results showed that the percentages of splenic Treg cells in CD4+ T cells were suppressed on day 14 post-infection (PI) but increased on day 56 PI, while Th17 cells were increased on day 56 PI compared with normal control (NC) mice. The Treg/Th17 ratio steadily increased from day 28 to day 56 PI. The hepatic levels of their specific transcription factors (Foxp3 for Treg and RORγt for Th17) were increased in C. sinensis-infected mice from day 14 to 56 PI, and significantly higher than those in NC mice. Meanwhile, serum levels of IL-2 and IL-17 were profoundly increased in C. sinensis-infected mice throughout the experiment; while the concentrations of IL-6 and transforming growth factor β1 (TGF-β1) peaked on day 14 PI, but then decreased on day 28 and 56 PI. Our results provide the first evidence of an increased Treg/Th17 ratio in C. sinensis-infected mice, suggesting that a Treg/Th17 imbalance may play a role in disease outcomes of clonorchiasis. PMID:26599407

Characterization of Bombyx mori nucleopolyhedrovirus Bm17.

Science.gov (United States)

Shen, Hongxing; Wang, Rudu; Han, Qinggong; Zhang, Wen; Nin, Bin; Zhou, Yang; Shao, Shihe; Yao, Qin; Chen, Keping; Liu, Xiaoyong

2013-10-01

Open reading frame17 (Bm17) of Bombyx mori nucleopolyhedrovirus is a highly conserved gene in lepidopteran nucleopolyhedroviruses, suggesting that it performs an important role in the virus life cycle whose function is unknown. In this report, we describe the characterization of Bm17. Reversed transcriptive-PCR (RT-PCR) and Western blot analysis demonstrated that Bm17 was expressed as a late gen. Immunofluorescence analysis by confocal microscopy showed that BM17 protein was localized on cytoplasm and nucleus of infected cells. These results show that BM17 was a late protein localized in cytoplasm and nucleus. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Initial investigation of glucose metabolism in mouse brain using enriched 17 O-glucose and dynamic 17 O-MRS.

Science.gov (United States)

Borowiak, Robert; Reichardt, Wilfried; Kurzhunov, Dmitry; Schuch, Christian; Leupold, Jochen; Krafft, Axel Joachim; Reisert, Marco; Lange, Thomas; Fischer, Elmar; Bock, Michael

2017-08-01

In this initial work, the in vivo degradation of 17 O-labeled glucose was studied during cellular glycolysis. To monitor cellular glucose metabolism, direct 17 O-magnetic resonance spectroscopy (MRS) was used in the mouse brain at 9.4 T. Non-localized spectra were acquired with a custom-built transmit/receive (Tx/Rx) two-turn surface coil and a free induction decay (FID) sequence with a short TR of 5.4 ms. The dynamics of labeled oxygen in the anomeric 1-OH and 6-CH 2 OH groups was detected using a Hankel-Lanczos singular value decomposition (HLSVD) algorithm for water suppression. Time-resolved 17 O-MRS (temporal resolution, 42/10.5 s) was performed in 10 anesthetized (1.25% isoflurane) mice after injection of a 2.2 M solution containing 2.5 mg/g body weight of differently labeled 17 O-glucose dissolved in 0.9% physiological saline. From a pharmacokinetic model fit of the H 2 17 O concentration-time course, a mean apparent cerebral metabolic rate of 17 O-labeled glucose in mouse brain of CMR Glc  = 0.07 ± 0.02 μmol/g/min was extracted, which is of the same order of magnitude as a literature value of 0.26 ± 0.06 μmol/g/min reported by 18 F-fluorodeoxyglucose ( 18 F-FDG) positron emission tomography (PET). In addition, we studied the chemical exchange kinetics of aqueous solutions of 17 O-labeled glucose at the C1 and C6 positions with dynamic 17 O-MRS. In conclusion, the results of the exchange and in vivo experiments demonstrate that the C6- 17 OH label in the 6-CH 2 OH group is transformed only glycolytically by the enzyme enolase into the metabolic end-product H 2 17 O, whereas C1- 17 OH ends up in water via direct hydrolysis as well as glycolysis. Therefore, dynamic 17 O-MRS of highly labeled 17 O-glucose could provide a valuable non-radioactive alternative to FDG PET in order to investigate glucose metabolism. Copyright © 2017 John Wiley & Sons, Ltd.

Proprotein Convertases Process Pmel17 during Secretion*

Science.gov (United States)

Leonhardt, Ralf M.; Vigneron, Nathalie; Rahner, Christoph; Cresswell, Peter

2011-01-01

Pmel17 is a melanocyte/melanoma-specific protein that traffics to melanosomes where it forms a fibrillar matrix on which melanin gets deposited. Before being cleaved into smaller fibrillogenic fragments the protein undergoes processing by proprotein convertases, a class of serine proteases that typically recognize the canonical motif RX(R/K)R↓. The current model of Pmel17 maturation states that this processing step occurs in melanosomes, but in light of recent reports this issue has become controversial. We therefore addressed this question by thoroughly assessing the processing kinetics of either wild-type Pmel17 or a secreted soluble Pmel17 derivative. Our results demonstrate clearly that processing of Pmel17 occurs during secretion and that it does not require entry of the protein into the endocytic system. Strikingly, processing proceeds even in the presence of the secretion inhibitor monensin, suggesting that Pmel17 is an exceptionally good substrate. In line with this, we find that newly synthesized surface Pmel17 is already quantitatively cleaved. Moreover, we demonstrate that Pmel17 function is independent of the sequence identity of its unconventional proprotein convertase-cleavage motif that lacks arginine in P4 position. The data alter the current view of Pmel17 maturation and suggest that the multistep processing of Pmel17 begins with an early cleavage during secretion that primes the protein for later functional processing. PMID:21247888

22 CFR 901.17 - Charged employee.

Science.gov (United States)

2010-04-01

... 22 Foreign Relations 2 2010-04-01 2010-04-01 true Charged employee. 901.17 Section 901.17 Foreign Relations FOREIGN SERVICE GRIEVANCE BOARD GENERAL Meanings of Terms As Used in This Chapter § 901.17 Charged employee. Charged employee means a member of the Senior Foreign Service or a member of the Service assigned...

46 CFR 78.17-25 - Sanitation.

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2010-10-01

... 46 Shipping 3 2010-10-01 2010-10-01 false Sanitation. 78.17-25 Section 78.17-25 Shipping COAST GUARD, DEPARTMENT OF HOMELAND SECURITY (CONTINUED) PASSENGER VESSELS OPERATIONS Tests, Drills, and Inspections § 78.17-25 Sanitation. (a) It shall be the duty of the master and chief engineer to see that the...

33 CFR 17.05-1 - Gifts.

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2010-07-01

... 33 Navigation and Navigable Waters 1 2010-07-01 2010-07-01 false Gifts. 17.05-1 Section 17.05-1 Navigation and Navigable Waters COAST GUARD, DEPARTMENT OF HOMELAND SECURITY GENERAL UNITED STATES COAST GUARD GENERAL GIFT FUND Administration § 17.05-1 Gifts. The gifts or bequests may be in money or...

31 CFR 17.110 - Self-evaluation.

Science.gov (United States)

2010-07-01

... 31 Money and Finance: Treasury 1 2010-07-01 2010-07-01 false Self-evaluation. 17.110 Section 17... § 17.110 Self-evaluation. (a) The agency shall, by two years after the effective date of this part... handicaps, to participate in the self-evaluation process. (c) The agency shall, until three years following...

Investigation of yeast genes possibly involved in mtDNA stability ...

African Journals Online (AJOL)

Phelim Isichei

maintenance was performed using our previous validated method of RNAi combined with ethidium bromide. ..... cytoskeleton-based segregation machinery. Mol. Biol. ... Axonal transport of mitochondria requires milton to recruit kinesin heavy.

Effects of HSP90 inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) on NEU/HER2 overexpressing mammary tumours in MMTV-NEU-NT mice monitored by Magnetic Resonance Spectroscopy.

Science.gov (United States)

Rodrigues, Loreta M; Chung, Yuen-Li; Al Saffar, Nada M S; Sharp, Swee Y; Jackson, Laura E; Banerji, Udai; Stubbs, Marion; Leach, Martin O; Griffiths, John R; Workman, Paul

2012-05-23

The importance of ERBB2/NEU/HER2 in the response of breast tumours to the heat shock protein 90 (HSP90) inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG; tanespimycin) has been demonstrated in the clinic. ERBB2 is an oncoprotein client that is highly dependent on HSP90. This and other oncogenic client proteins (e.g. B-RAF, C-RAF, ALK and CDK4) are depleted by 17-AAG in both animal tumours and patients. Here we investigate by Magnetic Resonance Spectroscopy (MRS) the metabolic response of 17-AAG in spontaneous, NEU/HER2 driven mammary tumours in transgenic MMTV-NEU-NT mice and in cells isolated and cultured from these tumours. Mammary tumours were monitored by 31P MRS in vivo and in tumour extracts, comparing control and 17-AAG treated mice. A cell line derived from NEU/HER2 mammary tumours was also cultured and the effect of 17-AAG was measured by 31P MRS in cell extracts. Molecular biomarkers were assessed by immunoblotting in extracts from cells and tumours. For comparison of tumour volume, metabolite concentrations and Western blot band intensities, two-tailed unpaired t-tests were used. The NEU/HER2 mammary tumours were very sensitive to 17-AAG and responded in a dose-dependent manner to 3 daily doses of 20, 40 and 80mg/kg of 17-AAG, all of which caused significant regression. At the higher doses, 31P MRS of tumour extracts showed significant decreases in phosphocholine (PC) and phosphoethanolamine (PE) whereas no significant changes were seen at the 20mg/kg dose. Extracts of isolated cells cultured from the mammary carcinomas showed a significant decrease in viable cell number and total PME after 17-AAG treatment. Western blots confirmed the expected action of 17-AAG in inducing HSP72 and significantly depleting HSP90 client proteins, including NEU/HER2 both in tumours and in isolated cells. The data demonstrate the high degree of sensitivity of this clinically relevant NEU/HER2-driven tumour model to HSP90 inhibition by 17-AAG, consistent with the

7 CFR 1209.17 - Promotion.

Science.gov (United States)

2010-01-01

... 7 Agriculture 10 2010-01-01 2010-01-01 false Promotion. 1209.17 Section 1209.17 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (MARKETING AGREEMENTS... CONSUMER INFORMATION ORDER Mushroom Promotion, Research, and Consumer Information Order Definitions § 1209...

Gallbladder Agenesis in 17 Dogs: 2006-2016.

Science.gov (United States)

Sato, K; Sakai, M; Hayakawa, S; Sakamoto, Y; Kagawa, Y; Kutara, K; Teshima, K; Asano, K; Watari, T

2018-01-01

Gallbladder agenesis (GBA) is extremely rare in dogs. To describe the history, clinical signs, diagnosis, treatment, and outcomes of dogs with GBA. Seventeen client-owned dogs with GBA. Medical records from 2006 through 2016 were retrospectively reviewed. Dogs were included when GBA was suspected on abdominal ultrasonography and confirmed by gross evaluation. Signalment, clinical signs, clinicopathological data, diagnostic imaging, histopathology, treatment, and outcome were recorded. Dogs were of 6 different breeds, and Chihuahuas (10 of 17) were most common. Median age at presentation was 1.9 (range, 0.7-7.4) years. Clinical signs included vomiting (5 of 17), anorexia (2 of 17), ascites (2 of 17), diarrhea (1 of 17), lethargy (1 of 17), and seizures (1 of 17). All dogs had increased serum activity of at least 1 liver enzyme, most commonly alanine aminotransferase (15 of 17). Fifteen dogs underwent computed tomography (CT) cholangiography; common bile duct (CBD) dilatation was confirmed in 12, without evidence of bile duct obstruction. Gross evaluation confirmed malformation of the liver lobes in 14 of 17 dogs and acquired portosystemic collaterals in 5 of 17. Ductal plate malformation was confirmed histologically in 16 of 17 dogs. During follow-up (range, 4-3,379 days), 16 of 17 dogs remained alive. Dogs with GBA exhibit clinicopathological signs of hepatobiliary injury and hepatic histopathological changes consistent with a ductal plate abnormality. Computed tomography cholangiography was superior to ultrasound examination in identifying accompanying nonobstructive CBD distention. Computed tomography cholangiography combined with laparoscopic liver biopsy is the preferable approach to characterize the full disease spectrum accompanying GBA in dogs. Copyright © 2018 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals, Inc. on behalf of the American College of Veterinary Internal Medicine.

Photoclickable dendritic molecular glue: noncovalent-to-covalent photochemical transformation of protein hybrids.

Science.gov (United States)

Uchida, Noriyuki; Okuro, Kou; Niitani, Yamato; Ling, Xiao; Ariga, Takayuki; Tomishige, Michio; Aida, Takuzo

2013-03-27

A water-soluble dendron with a fluorescein isothiocyanate (FITC) fluorescent label and bearing nine pendant guanidinium ion (Gu(+))/benzophenone (BP) pairs at its periphery (Glue(BP)-FITC) serves as a "photoclickable molecular glue". By multivalent salt-bridge formation between Gu(+) ions and oxyanions, Glue(BP)-FITC temporarily adheres to a kinesin/microtubule hybrid. Upon subsequent exposure to UV light, this noncovalent binding is made permanent via a cross-linking reaction mediated by carbon radicals derived from the photoexcited BP units. This temporal-to-permanent transformation by light occurs quickly and efficiently in this preorganized state, allowing the movements of microtubules on a kinesin-coated glass plate to be photochemically controlled. A fundamental difference between such temporal and permanent bindings was visualized by the use of "optical tweezers".

The Dynamics of Treg/Th17 and the Imbalance of Treg/Th17 in Clonorchis sinensis-Infected Mice.

Directory of Open Access Journals (Sweden)

Chao Yan

Full Text Available Clonorchiasis, caused by the liver fluke Clonorchis sinensis, is a chronic parasitic infection regulated by T cell subsets. An imbalance of CD4+CD25+ Foxp3+regulatory T (Treg and interleukin (IL-17-secreting T cells (Th17 may control inflammation and play an important role in the pathogenesis of immune evasion. In the present study, we assessed the dynamics of Treg/Th17 and determined whether the Treg/Th17 ratio is altered in C. sinensis-infected mice. The results showed that the percentages of splenic Treg cells in CD4+ T cells were suppressed on day 14 post-infection (PI but increased on day 56 PI, while Th17 cells were increased on day 56 PI compared with normal control (NC mice. The Treg/Th17 ratio steadily increased from day 28 to day 56 PI. The hepatic levels of their specific transcription factors (Foxp3 for Treg and RORγt for Th17 were increased in C. sinensis-infected mice from day 14 to 56 PI, and significantly higher than those in NC mice. Meanwhile, serum levels of IL-2 and IL-17 were profoundly increased in C. sinensis-infected mice throughout the experiment; while the concentrations of IL-6 and transforming growth factor β1 (TGF-β1 peaked on day 14 PI, but then decreased on day 28 and 56 PI. Our results provide the first evidence of an increased Treg/Th17 ratio in C. sinensis-infected mice, suggesting that a Treg/Th17 imbalance may play a role in disease outcomes of clonorchiasis.

Thrombopoietin/MPL participates in initiating and maintaining RUNX1-ETO acute myeloid leukemia via PI3K/AKT signaling

NARCIS (Netherlands)

J.A. Pulikkan (John); D. Madera (Dmitri); L. Xue (Liting); P. Bradley (Paul); S.F. Landrette (Sean Francis); Y.-H. Kuo (Ya-Huei); S. Abbas (Saman); L.J. Zhu (Lihua Julie); P.J.M. Valk (Peter); L.H. Castilla (Lucio)

2012-01-01

textabstractOncogenic mutations in components of cytokine signaling pathways elicit ligand-independent activation of downstream signaling, enhancing proliferation and survival in acute myeloid leukemia (AML). The myeloproliferative leukemia virus oncogene, MPL, a homodimeric receptor activated by

Magnetic sublattices in Np{sub 2}Co{sub 17} and Np{sub 2}Ni{sub 17}

Energy Technology Data Exchange (ETDEWEB)

Colineau, E., E-mail: eric.colineau@ec.europa.eu; Hen, A. [Institute for Transuranium Elements (ITU), European Commission, Joint Research Centre (JRC) (Germany); Sanchez, J.-P. [CEA, INAC-SPSMS (France); Griveau, J.-C.; Magnani, N.; Eloirdi, R. [Institute for Transuranium Elements (ITU), European Commission, Joint Research Centre (JRC) (Germany); Halevy, I. [Ben Gurion University, Nuclear Engineering Department (Israel); Gaczyński, P. [Institute for Transuranium Elements (ITU), European Commission, Joint Research Centre (JRC) (Germany); Orion, I. [Ben Gurion University, Nuclear Engineering Department (Israel); Shick, A. B. [Institute of Physics, ASCR (Czech Republic); Caciuffo, R. [Institute for Transuranium Elements (ITU), European Commission, Joint Research Centre (JRC) (Germany)

2016-12-15

Rare-earth-based compounds R{sub 2}T{sub 17} (R=Rare earth; T=Transition metal) have been extensively studied and developed for applications as permanent magnets. The actinide-based analogues, however, are much less documented and we report here about the magnetic properties of Np{sub 2}Co{sub 17} and Np{sub 2}Ni{sub 17}, as inferred from {sup 237}Np Mössbauer spectroscopy, the best resonance in actinides, and specific heat.

Gclust Server: 94819 [Gclust Server

Lifescience Database Archive (English)

Full Text Available ted protein required for both karyogamy and mitotic spindle organization, interacts stably and specifically ...ve annotation Kinesin-associated protein required for both karyogamy and mitotic spindle organization, inter

7 CFR 924.17 - Container.

Science.gov (United States)

2010-01-01

... 7 Agriculture 8 2010-01-01 2010-01-01 false Container. 924.17 Section 924.17 Agriculture Regulations of the Department of Agriculture (Continued) AGRICULTURAL MARKETING SERVICE (Marketing Agreements... Container. Container means a box, bag, crate, lug, basket, carton, package, or any other type of receptacle...

31 CFR 223.17 - Revocation.

Science.gov (United States)

2010-07-01

... 31 Money and Finance: Treasury 2 2010-07-01 2010-07-01 false Revocation. 223.17 Section 223.17 Money and Finance: Treasury Regulations Relating to Money and Finance (Continued) FISCAL SERVICE, DEPARTMENT OF THE TREASURY FINANCIAL MANAGEMENT SERVICE SURETY COMPANIES DOING BUSINESS WITH THE UNITED...

50 CFR 17.104 - Prohibitions.

Science.gov (United States)

2010-10-01

... PLANTS (CONTINUED) ENDANGERED AND THREATENED WILDLIFE AND PLANTS (CONTINUED) Manatee Protection Areas § 17.104 Prohibitions. Except as provided in § 17.105, (a) Manatee sanctuary. It is unlawful for any person to engage in any waterborne activity within a manatee sanctuary. (b) Manatee refuge. It is...

High IL-17E and low IL-17C dermal expression identifies a fibrosis-specific motif common to morphea and systemic sclerosis.

Directory of Open Access Journals (Sweden)

Paola Adele Lonati

Full Text Available BACKGROUND: High interleukin (IL-17A levels are characteristically found in the skin of systemic sclerosis (SSc individuals. Our aim was to investigate whether the dermal expression of IL-17A and related IL-17 family members (i.e. IL-17C, IL-17E and IL-17F could distinguish fibrotic from healthy skin and could show similarities in SSc and morphea, two disorders with presumed distinct pathogenesis, but characterized by skin fibrosis. METHODS: Biopsies were obtained from the involved skin of 14 SSc, 5 morphea and 8 healthy donors (HD undergoing plastic surgery. Immunohistochemistry/immunofluorescence techniques were coupled to a semi-automated imaging quantification approach to determine the presence of the IL-17 family members in the skin. The in vitro effects induced by the IL-17 family members on fibroblasts from normal and SSc individuals were assessed by ELISA and RIA. RESULTS: Positive cells for each of the IL-17 isoforms investigated were present in the dermis of all the individuals tested, though with variable frequencies. SSc individuals had increased frequency of IL-17A+ (p = 0.0237 and decreased frequency of IL-17F+ (p = 0.0127 and IL-17C+ cells (p = 0.0008 when compared to HD. Similarly, morphea individuals had less frequent IL-17C+ cells (p = 0.0186 in their skin but showed similar number of IL-17A+ and IL-17F+ cells when compared to HD. Finally, IL-17E+ cells were more numerous in morphea (p = 0.0109 and tended to be more frequent in SSc than in HD. Fibroblast production of IL-6, MMP-1 and MCP-1 was enhanced in a dose-dependent manner in the presence of IL-17E and IL-17F, but not in the presence of IL-17C. None of the cytokine tested had significant effect on type I collagen production. Of interest, in SSc the frequency of both IL-17A and IL-17F positive cells increased with disease duration. CONCLUSIONS: The frequency of IL-17A and IL-17F distinguish SSc to morphea individuals while dermal expression of IL-17C (low and IL-17E (high

Th-17 regulatory cytokines IL-21, IL-23, and IL-6 enhance neutrophil production of IL-17 cytokines during asthma.

Science.gov (United States)

Halwani, Rabih; Sultana, Asma; Vazquez-Tello, Alejandro; Jamhawi, Amer; Al-Masri, Abeer A; Al-Muhsen, Saleh

2017-11-01

In a subset of severe asthma patients, chronic airway inflammation is associated with infiltration of neutrophils, Th-17 cells and elevated expression of Th-17-derived cytokines (e.g., interleukin [IL]-17, IL-21, IL-22). Peripheral neutrophils from allergic asthmatics are known to express higher IL-17 cytokine levels than those from healthy subjects, but the regulatory mechanisms involved are not well understood. We hypothesize that Th-17 regulatory cytokines could modulate IL-17 expression in neutrophils. Peripheral blood neutrophils isolated from asthmatics were stimulated with IL-21, IL-23, and IL-6 cytokines and their ability to produce IL-17A and IL-17F was determined relative to healthy controls. Signal transducer and activator of transcription 3 (STAT3) phosphorylation levels were measured in stimulated neutrophil using flow cytometry. The requirement for STAT3 phosphorylation was determined by blocking its activation using a specific chemical inhibitor. Stimulating asthmatic neutrophils with IL-21, 23, and 6 enhanced the production of IL-17A and IL-17F at significantly higher levels comparatively to healthy controls. Stimulating neutrophils with IL-21, IL-23, and IL-6 cytokines enhanced STAT3 phosphorylation, in all cases. Interestingly, inhibiting STAT3 phosphorylation using a specific chemical inhibitor dramatically blocked the ability of neutrophils to produce IL-17, demonstrating that STAT3 activation is the major factor mediating IL-17 gene expression. These findings suggest that neutrophil infiltration in lungs of severe asthmatics may represent an important source of pro-inflammatory IL-17A and -F cytokines, a production enhanced by Th-17 regulatory cytokines, and thus providing a feedback mechanism that sustains inflammation. Our results suggest that STAT3 pathway could be a potential target for regulating neutrophilic inflammation during severe asthma.

Systemic Th17/IL-17A response appears prior to hippocampal neurodegeneration in rats exposed to low doses of ozone.

Science.gov (United States)

Solleiro-Villavicencio, H; Rivas-Arancibia, S

2017-06-03

Exposure to low doses of O 3 leads to a state of oxidative stress. Some studies show that oxidative stress can modulate both the CNS and systemic inflammation, which are important factors in the development of Alzheimer disease (AD). This study aims to evaluate changes in the frequency of Th17-like cells (CD3 + CD4 + IL-17A + ), the concentration of IL-17A in peripheral blood, and hippocampal immunoreactivity to IL-17A in rats exposed to low doses of O 3 . One hundred eight male Wistar rats were randomly assigned to 6 groups (n=18) receiving the following treatments: control (O 3 free) or O 3 exposure (0.25ppm, 4hours daily) over 7, 15, 30, 60, and 90 days. Twelve animals from each group were decapitated and a peripheral blood sample was taken to isolate plasma and mononuclear cells. Plasma IL-17A was quantified using LUMINEX, while Th17-like cells were counted using flow cytometry. The remaining 6 rats were deeply anaesthetised and underwent transcardial perfusion for immunohistological study of the hippocampus. Results show that exposure to O 3 over 7 days resulted in a significant increase in the frequency of Th17-like cells and levels of IL-17A in peripheral blood. However, levels of Th17/IL-17A in peripheral blood were lower at day 15 of exposure. We also observed increased IL-17A in the hippocampus beginning at 30 days of exposure. These results indicate that O 3 induces a short-term, systemic Th17-like/IL-17A effect and an increase of IL-17A in the hippocampal tissue during the chronic neurodegenerative process. Copyright © 2017 Sociedad Española de Neurología. Publicado por Elsevier España, S.L.U. All rights reserved.

IL17/IL17RA as a Novel Signaling Axis Driving Mesenchymal Stem Cell Therapeutic Function in Experimental Autoimmune Encephalomyelitis

Directory of Open Access Journals (Sweden)

Mónica Kurte

2018-04-01

Full Text Available The therapeutic effect of mesenchymal stem cells (MSCs in multiple sclerosis (MS and the experimental autoimmune encephalomyelitis (EAE model has been well described. This effect is, in part, mediated through the inhibition of IL17-producing cells and the generation of regulatory T cells. While proinflammatory cytokines such as IFNγ, TNFα, and IL1β have been shown to enhance MSCs immunosuppressive function, the role of IL17 remains poorly elucidated. The aim of this study was, therefore, to investigate the role of the IL17/IL17R pathway on MSCs immunoregulatory effects focusing on Th17 cell generation in vitro and on Th17-mediated EAE pathogenesis in vivo. In vitro, we showed that the immunosuppressive effect of MSCs on Th17 cell proliferation and differentiation is partially dependent on IL17RA expression. This was associated with a reduced expression level of MSCs immunosuppressive mediators such as VCAM1, ICAM1, and PD-L1 in IL17RA−/− MSCs as compared to wild-type (WT MSCs. In the EAE model, we demonstrated that while WT MSCs significantly reduced the clinical scores of the disease, IL17RA−/− MSCs injected mice exhibited a clinical worsening of the disease. The disability of IL17RA−/− MSCs to reduce the progression of the disease paralleled the inability of these cells to reduce the frequency of Th17 cells in the draining lymph node of the mice as compared to WT MSCs. Moreover, we showed that the therapeutic effect of MSCs was correlated with the generation of classical Treg bearing the CD4+CD25+Foxp3+ signature in an IL17RA-dependent manner. Our findings reveal a novel role of IL17RA on MSCs immunosuppressive and therapeutic potential in EAE and suggest that the modulation of IL17RA in MSCs could represent a novel method to enhance their therapeutic effect in MS.

First-forbidden mirror β-decays in A = 17 mass region and the role of proton halo in 17F

International Nuclear Information System (INIS)

Michel, N.; Okolowicz, J.; Ploszajczak, M.; Okolowicz, J.; Nowacki, F.; Ploszajczak, M.

2001-01-01

The first-forbidden β-decay of 17 Ne into the 'halo' state J π 1/2 + 1 of 17 F presents one of the largest measured asymmetries for mirror β-decay feeding bound final states. This asymmetry is studied in the framework of the Shell Model Embedded in the Continuum (SMEC). The spatial extent of single particle orbits calculated in SMEC is constrained by the proton capture cross-section 16 O(p,γ) 17 F. This allows to estimate the mirror symmetry breaking in 17 F and 17 O nuclei. (authors)

36 CFR 331.17 - Minerals.

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2010-07-01

... 36 Parks, Forests, and Public Property 3 2010-07-01 2010-07-01 false Minerals. 331.17 Section 331..., KENTUCKY AND INDIANA § 331.17 Minerals. All activities in connection with prospecting, exploration, development, mining or other removal or the processing of mineral resources and all uses reasonably incident...

31 CFR 17.140 - Employment.

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2010-07-01

... 31 Money and Finance: Treasury 1 2010-07-01 2010-07-01 false Employment. 17.140 Section 17.140... Employment. No qualified individual with handicaps shall, on the basis of handicap, be subjected to discrimination in employment under any program or activity conducted by the Department. The definitions...

48 CFR 17.206 - Evaluation.

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2010-10-01

... 48 Federal Acquisition Regulations System 1 2010-10-01 2010-10-01 false Evaluation. 17.206 Section... CONTRACT TYPES SPECIAL CONTRACTING METHODS Options 17.206 Evaluation. (a) In awarding the basic contract... officer need not evaluate offers for any option quantities when it is determined that evaluation would not...

23 CFR 658.17 - Weight.

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2010-04-01

... 23 Highways 1 2010-04-01 2010-04-01 false Weight. 658.17 Section 658.17 Highways FEDERAL HIGHWAY ADMINISTRATION, DEPARTMENT OF TRANSPORTATION ENGINEERING AND TRAFFIC OPERATIONS TRUCK SIZE AND WEIGHT, ROUTE... agency transit passenger bus, is excluded from the axle weight limits in paragraphs (c) through (e) of...

Pressure loss tests for DR-BEP of fullsize 17 x 17 PWR fuel assembly

International Nuclear Information System (INIS)

Chung, Moon Ki; Chun, Se Young; Chang, Seok Kyu; Won, Soon Youn; Cho, Young Rho; Kim, Bok Deuk; Min, Kyoung Ho

1993-01-01

This report describes the conditions, procedure and results in the pressure loss tests carried out for a double grid type debris resistance bottom end piece (DR-BEP) designed by KAERI. In this test, the pressure loss coefficients of the full size 17 x 17 PWR simulated fuel assembly with DR-BET and with standard-BEP were measured respectively, and the pressure loss coefficients of DR-BEP were compared with the coefficients of STD-BET. The test conditions fall within the ranges of loop pressure from 5.2 to 45 bar, loop temperature from 27 to 221 deg C and Reynolds number in fuel bundle from 2.17 x 10 4 to 3.85 x 10 5 . (Author) 5 refs., 18 figs., 5 tabs

31 CFR 17.102 - Application.

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2010-07-01

... 31 Money and Finance: Treasury 1 2010-07-01 2010-07-01 false Application. 17.102 Section 17.102 Money and Finance: Treasury Office of the Secretary of the Treasury ENFORCEMENT OF NONDISCRIMINATION ON... Application. This part applies to all programs or activities conducted by the agency, except for programs or...

21 CFR 17.32 - Motions.

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2010-04-01

... 21 Food and Drugs 1 2010-04-01 2010-04-01 false Motions. 17.32 Section 17.32 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL CIVIL MONEY PENALTIES... shall be filed with the Division of Dockets Management (HFA-305), Food and Drug Administration, 5630...

21 CFR 17.9 - Answer.

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2010-04-01

... 21 Food and Drugs 1 2010-04-01 2010-04-01 false Answer. 17.9 Section 17.9 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL CIVIL MONEY PENALTIES HEARINGS... Dockets Management (HFA-305), Food and Drug Administration, 5630 Fishers Lane, rm. 1061, Rockville, MD...

Molecular cloning and characterization of duck interleukin-17

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Interleukin-17 (IL-17) belonging to the Th17 family is a proinflammatory cytokine produced by activated T cells. A 1034-bp cDNA encoding duck IL-17 (duIL-17) was cloned from ConA-activated splenic lymphocytes of ducks. The encoded protein, predicted to consisted of 169 amino acids, displayed a molec...

Fra SK 4b til MH 17

DEFF Research Database (Denmark)

Stræde, Therkel

2015-01-01

Om den nazistiske besættelsespolitik i Sydøstukraine og Sonderkommando 4b's udryddelse af jøder på den egn, hvor Malaysian Airlines MH 17 styrtede ned den 17. juli 2014......Om den nazistiske besættelsespolitik i Sydøstukraine og Sonderkommando 4b's udryddelse af jøder på den egn, hvor Malaysian Airlines MH 17 styrtede ned den 17. juli 2014...

Spin reorientation and magnetic anisotropy in Y2Co17-xCr x (x 1.17-3.0) compounds

International Nuclear Information System (INIS)

Fuquan, B.; Tegus, O.; Dagula, W.; Brueck, E.; Boer, F.R. de; Buschow, K.H.J.

2005-01-01

Spin reorientation transitions and magnetic anisotropy in Y 2 Co 17-x Cr x (x = 1.17-3.0) compounds have been investigated by means of X-ray diffraction and magnetization measurements. The powder X-ray diffraction patterns show that most samples crystallize as a single phase with the rhombohedral Th 2 Zn 17 -type structure. However, in the compound Y 2 Co 14 Cr 3 the Th 2 Zn 17 phase coexist with the hexagonal Th 2 Ni 17 -type phase. The lattice parameters a and c hardly change and the unit cell volume V increases slightly with increasing Cr content. The X-ray diffraction patterns of the aligned powder of the samples have confirmed that at room temperature the compound with x = 1.17 has planar anisotropy, but the compounds with x = 1.76, 2.34 and 3.00 have uniaxial anisotropy. Spin reorientation phenomena occur in all of the compounds. With increasing Cr content, the Curie temperature, the spin reorientation temperature, the spontaneous magnetization, and the anisotropy constant K 2 of the Y 2 Co 17-x Cr x (x = 1.17-3.0) compounds decrease strongly while the anisotropy constant K 1 increases in the range of x from 1.17 to 2.34 and then decreases in the range of x from 2.34 to 3.00

Biophysics of filament length regulation by molecular motors

International Nuclear Information System (INIS)

Kuan, Hui-Shun; Betterton, M D

2013-01-01

Regulating physical size is an essential problem that biological organisms must solve from the subcellular to the organismal scales, but it is not well understood what physical principles and mechanisms organisms use to sense and regulate their size. Any biophysical size-regulation scheme operates in a noisy environment and must be robust to other cellular dynamics and fluctuations. This work develops theory of filament length regulation inspired by recent experiments on kinesin-8 motor proteins, which move with directional bias on microtubule filaments and alter microtubule dynamics. Purified kinesin-8 motors can depolymerize chemically-stabilized microtubules. In the length-dependent depolymerization model, the rate of depolymerization tends to increase with filament length, because long filaments accumulate more motors at their tips and therefore shorten more quickly. When balanced with a constant filament growth rate, this mechanism can lead to a fixed polymer length. However, the mechanism by which kinesin-8 motors affect the length of dynamic microtubules in cells is less clear. We study the more biologically realistic problem of microtubule dynamic instability modulated by a motor-dependent increase in the filament catastrophe frequency. This leads to a significant decrease in the mean filament length and a narrowing of the filament length distribution. The results improve our understanding of the biophysics of length regulation in cells. (paper)

T-helper 17 and interleukin-17-producing lymphoid tissue inducer-like cells make different contributions to colitis in mice.

Science.gov (United States)

Ono, Yuichi; Kanai, Takanori; Sujino, Tomohisa; Nemoto, Yasuhiro; Kanai, Yasumasa; Mikami, Yohei; Hayashi, Atsushi; Matsumoto, Atsuhiro; Takaishi, Hiromasa; Ogata, Haruhiko; Matsuoka, Katsuyoshi; Hisamatsu, Tadakazu; Watanabe, Mamoru; Hibi, Toshifumi

2012-11-01

T helper (Th) 17 cells that express the retinoid-related orphan receptor (ROR) γt contribute to the development of colitis in mice, yet are found in normal and inflamed intestine. We investigated their development and functions in intestines of mice. We analyzed intestinal Th17 cells in healthy and inflamed intestinal tissues of mice. We analyzed expression of lymphotoxin (LT)α by Th17 cells and lymphoid tissue inducer-like cells. LTα(-/-) and RORγt(-/-) mice had significantly lower percentages of naturally occurring Th17 cells in the small intestine than wild-type mice. Numbers of CD3(-)CD4(+/-)interleukin-7Rα(+)c-kit(+)CCR6(+)NKp46(-) lymphoid tissue inducer-like cells that produce interleukin-17A were increased in LTα(-/-) and LTα(-/-) × recombination activating gene (RAG)-2(-/-) mice, compared with wild-type mice, but were absent from RORγt(-/-) mice. Parabiosis of wild-type and LTα(-/-) mice and bone marrow transplant experiments revealed that LTα-dependent gut-associated lymphoid tissue structures are required for generation of naturally occurring Th17 cells. However, when wild-type or LTα(-/-) CD4(+)CD45RB(high) T cells were transferred to RAG-2(-/-) or LTα(-/-)×RAG-2(-/-) mice, all groups, irrespective of the presence or absence of LTα on the donor or recipient cells, developed colitis and generated Th1, Th17, and Th17/Th1 cells. RAG-2(-/-) mice that received a second round of transplantation, with colitogenic but not naturally occurring Th17 cells, developed intestinal inflammation. The presence of naturally occurring Th17 cells in the colons of mice inhibited development of colitis after transfer of CD4(+)CD45RB(high) T cells and increased the numbers of Foxp3(+) cells derived from CD4(+)CD45RB(high) T cells. Gut-associated lymphoid tissue structures are required to generate naturally occurring Th17 cells that have regulatory activities in normal intestines of mice, but not for colitogenic Th17 and Th17/Th1 cells during inflammation

12 CFR 602.17 - Policy.

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2010-01-01

... 12 Banks and Banking 6 2010-01-01 2010-01-01 false Policy. 602.17 Section 602.17 Banks and Banking... produce documents. This subpart does not affect access to documents under the FOIA or the Privacy Act. See... this subpart remain our property. Any employee having information or privileged documents may disclose...

21 CFR 17.37 - Witnesses.

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2010-04-01

... 21 Food and Drugs 1 2010-04-01 2010-04-01 false Witnesses. 17.37 Section 17.37 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES GENERAL CIVIL MONEY PENALTIES... pay for his or her travel to the hearing. The sponsoring party is responsible for producing the...

Crystal structure of a plant albumin from Cicer arietinum (chickpea) possessing hemopexin fold and hemagglutination activity.

Science.gov (United States)

Sharma, Urvashi; Katre, Uma V; Suresh, C G

2015-05-01

Crystal structure of a reported PA2 albumin from Cicer arietinum shows that it belongs to hemopexin fold family, has four beta-propeller motifs and possesses hemagglutination activity, making it different from known legume lectins. A plant albumin (PA2) from Cicer arietinum, presumably a lectin (CAL) owing to its hemagglutination activity which is inhibited by complex sugars as well as glycoproteins such as fetuin, desialylated fetuin and fibrinogen. The three-dimensional structure of this homodimeric protein has been determined using X-ray crystallography at 2.2 Å in two crystal forms: orthorhombic (P21212) and trigonal (P3). The structure determined using molecular replacement method and refined in orthorhombic crystal form reached R-factors R free 22.6 % and R work 18.2 % and in trigonal form had 22.3 and 17.9 % in the resolution range of 20.0-2.2 and 35.3-2.2 Å, respectively. Interestingly, unlike the known legume lectin fold, the structure of this homodimeric hemagglutinin belonged to hemopexin fold that consisted of four-bladed β-propeller architecture. Each subunit has a central cavity forming a channel, inside of which is lined with hydrophobic residues. The channel also bears binding sites for ligands such as calcium, sodium and chloride ions, iodine atom in the case of iodine derivative and water molecules. However, none of these ligands seem important for the sugar recognition. No monosaccharide sugar specificity could be detected using hemagglutination inhibition. Chemical modification studies identified a potential sugar-binding site per subunit molecule. Comparison of C-alpha atom positions in subunit structures showed that the deviations between the two crystal forms were more with respect to blades I and IV. Differences also existed between subunits in two forms in terms of type and site of ligand binding.

Study of RNA interference inhibiting rat ovarian androgen biosynthesis by depressing 17alpha-hydroxylase/17, 20-lyase activity in vivo

Directory of Open Access Journals (Sweden)

Yang Xing

2009-07-01

Full Text Available Abstract Background 17alpha-hydroxylase/17, 20-lyase encoded by CYP17 is the key enzyme in androgen biosynthesis pathway. Previous studies demonstrated the accentuation of the enzyme in patients with polycystic ovary syndrome (PCOS was the most important mechanism of androgen excess. We chose CYP17 as the therapeutic target, trying to suppress the activity of 17alpha-hydroxylase/17, 20-lyase and inhibit androgen biosynthesis by silencing the expression of CYP17 in the rat ovary. Methods Three CYP17-targeting and one negative control oligonucleotides were designed and used in the present study. The silence efficiency of lentivirus shRNA was assessed by qRT-PCR, Western blotting and hormone assay. After subcapsular injection of lentivirus shRNA in rat ovary, the delivery efficiency was evaluated by GFP fluorescence and qPCR. Total RNA was extracted from rat ovary for CYP17 mRNA determination and rat serum was collected for hormone measurement. Results In total, three CYP17-targeting lentivirus shRNAs were synthesized. The results showed that all of them had a silencing effect on CYP17 mRNA and protein. Moreover, androstenedione secreted by rat theca interstitial cells (TIC in the RNAi group declined significantly compared with that in the control group. Two weeks after rat ovarian subcapsular injection of chosen CYP17 shRNA, the GFP fluorescence of frozen ovarian sections could be seen clearly under fluorescence microscope. It also showed that the GFP DNA level increased significantly, and its relative expression level was 7.42 times higher than that in the control group. Simultaneously, shRNA treatment significantly decreased CYP17 mRNA and protein levels at 61% and 54%, respectively. Hormone assay showed that all the levels of androstenedione, 17-hydroxyprogesterone and testosterone declined to a certain degree, but progesterone levels declined significantly. Conclusion The present study proves for the first time that ovarian androgen

Purification, crystallization, X-ray diffraction analysis and phasing of an engineered single-chain PvuII restriction endonuclease

International Nuclear Information System (INIS)

Meramveliotaki, Chrysi; Kotsifaki, Dina; Androulaki, Maria; Hountas, Athanasios; Eliopoulos, Elias; Kokkinidis, Michael

2007-01-01

PvuII is the first type II restriction endonuclease to be converted from its wild-type homodimeric form into an enzymatically active single-chain variant. The enzyme was crystallized and phasing was successfully performed by molecular replacement. The restriction endonuclease PvuII from Proteus vulgaris has been converted from its wild-type homodimeric form into the enzymatically active single-chain variant scPvuII by tandemly joining the two subunits through the peptide linker Gly-Ser-Gly-Gly. scPvuII, which is suitable for the development of programmed restriction endonucleases for highly specific DNA cleavage, was purified and crystallized. The crystals diffract to a resolution of 2.35 Å and belong to space group P4 2 , with unit-cell parameters a = b = 101.92, c = 100.28 Å and two molecules per asymmetric unit. Phasing was successfully performed by molecular replacement

15 CFR 785.17 - Settlement.

Science.gov (United States)

2010-01-01

... 15 Commerce and Foreign Trade 2 2010-01-01 2010-01-01 false Settlement. 785.17 Section 785.17... Settlement. (a) Settlements before issuance of a NOVA. When the parties have agreed to a settlement of the case prior to issuance of a NOVA, a settlement proposal consisting of a settlement agreement and order...

The IL-17 and Th17 cell immune response in cervical cancer : angels or demons : it depends on the context

NARCIS (Netherlands)

Punt, Birgitte Simone

2015-01-01

This thesis provides novel insights into the role of IL-17 and Th17 cells in cervical cancer. While IL-17 was shown to be predominantly produced by innate myeloid cells such as neutrophils and correlated with poor survival, Th17 cells were generally a small cell population correlated with improved

Effect of 17-allylamino-17-demethoxygeldanamycin (17-AAG) on Akt protein expression is more effective in head and neck cancer cell lineages that retain PTEN protein expression.

Science.gov (United States)

Pontes, Flávia Sirotheau C; Pontes, Hélder A R; de Souza, Lucas L; de Jesus, Adriana S; Joaquim, Andrea M C; Miyahara, Ligia A N; Fonseca, Felipe P; Pinto Junior, Décio S

2018-03-01

The aim of this study was to evaluate the expression of Akt, PTEN, Mdm2 and p53 proteins in three different head and neck squamous cell carcinoma (HNSCC) cell lines (HN6, HN19 and HN30), all of them treated with epidermal growth factor (EGF) and 17-allylamino-17-demethoxygeldanamycin (17-AAG), an inhibitor of Hsp90 protein. Immunofluorescence and western blot were performed in order to analyze the location and quantification, respectively, of proteins under the action 17-AAG and EGF. Treatment with EGF resulted in increased levels of Akt, PTEN and p53 in all cell lineages. The expression of Mdm2 was constant in HN30 and HN6 lineages, while in HN19 showed slightly decreased expression. Under the action 17-AAG, in HN6 and HN19, the expression of PTEN and p53 proteins was suppressed, while Akt and Mdm2 expression was reduced. Finally, in the HN30 cell lineage were absolute absence of expression of Akt, Mdm2 and p53 and decreased expression of PTEN. These data allow us to speculate on the particular utility of 17-AAG for HNSCC treatment through the inhibition of Akt protein expression, especially in the cases that retain the expression of PTEN protein. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Dicty_cDB: FC-AI17 [Dicty_cDB

Lifescience Database Archive (English)

Full Text Available FC (Link to library) FC-AI17 (Link to dictyBase) - - - Contig-U15690-1 FC-AI17P (Li...nk to Original site) FC-AI17F 587 FC-AI17Z 626 FC-AI17P 1212 - - Show FC-AI17 Library FC (Link to library) Clone ID FC-AI...nal site URL http://dictycdb.biol.tsukuba.ac.jp/CSM/FC/FC-AI/FC-AI17Q.Seq.d/ Representative seq. ID FC-AI...17P (Link to Original site) Representative DNA sequence >FC-AI17 (FC-AI17Q) /CSM/FC/FC-AI/FC-AI...slated Amino Acid sequence ANIATVGDFLKADTVVPKMIITYNKRKQGTDYLKAVIGPILSNVIKQELNLELKPNLVYA AIISEQEIRTGEKSTLDRNV

In vitro study comparing the efficacy of the water-soluble HSP90 inhibitors, 17-AEPGA and 17-DMAG, with that of the non‑water-soluble HSP90 inhibitor, 17-AAG, in breast cancer cell lines.

Science.gov (United States)

Ghadban, Tarik; Jessen, André; Reeh, Matthias; Dibbern, Judith L; Mahner, Sven; Mueller, Volkmar; Wellner, Ulrich F; Güngör, Cenap; Izbicki, Jakob R; Vashist, Yogesh K

2016-10-01

Heat shock protein (HSP)90 has emerged as an important target in cancer therapeutics. Diverse HSP90 inhibitors are under evaluation. The aim of the present study was to investigate the growth inhibitory effects of the newly developed water-soluble HSP90 inhibitors, 17-[2-(Pyrrolidin-1-yl)ethyl]amino-17-demethoxygeldanamycin (17-AEPGA) and 17-dimethylaminoethylamino-17-demethoxygeldanamycin (17-DMAG), compared to that of the non-water-soluble HSP90 inhibitor, 17-allylamino-17-demethoxygeldanamycin (17-AAG). The anti-proliferative effects of the 3 drugs on the human breast cancer cell lines, MCF-7, SKBR-3 and MDA-MB-231, were examined in vitro. In addition, tumor progression factors, including human epidermal growth factor receptor 2 (HER2), epidermal growth factor receptor 1 (EGFR1) and insulin-like growth factor type 1 receptor (IGF1R), as well as apoptotic markers were analysed. We found a time- and dose-dependent effect in all the tested cell lines. The effects of 17-AEPGA and 17-DMAG were equal or superior to those of 17-AAG. The 50% growth inhibition concentration was AAG.

22 CFR 226.17 - Certifications and representations.

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2010-04-01

... 22 Foreign Relations 1 2010-04-01 2010-04-01 false Certifications and representations. 226.17 Section 226.17 Foreign Relations AGENCY FOR INTERNATIONAL DEVELOPMENT ADMINISTRATION OF ASSISTANCE AWARDS TO U.S. NON-GOVERNMENTAL ORGANIZATIONS Pre-award Requirements § 226.17 Certifications and...

21 CFR 17.1 - Scope.

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2010-04-01

... 21 Food and Drugs 1 2010-04-01 2010-04-01 false Scope. 17.1 Section 17.1 Food and Drugs FOOD AND... Food, Drug, and Cosmetic Act (the act) authorizing civil money penalties for certain violations of the... trial data bank and section 303(f)(4) of the act authorizing civil money penalties for certain...

The structure of Lactococcus lactis thioredoxin reductase reveals molecular features of photo-oxidative damage

DEFF Research Database (Denmark)

Skjoldager, Nicklas; Bang, Maria Blanner; Rykær, Martin

2017-01-01

The NADPH-dependent homodimeric flavoenzyme thioredoxin reductase (TrxR) provides reducing equivalents to thioredoxin, a key regulator of various cellular redox processes. Crystal structures of photo-inactivated thioredoxin reductase (TrxR) from the Gram-positive bacterium Lactococcus lactis have...

5 CFR 1320.17 - Information collection budget.

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2010-01-01

... 5 Administrative Personnel 3 2010-01-01 2010-01-01 false Information collection budget. 1320.17 Section 1320.17 Administrative Personnel OFFICE OF MANAGEMENT AND BUDGET OMB DIRECTIVES CONTROLLING PAPERWORK BURDENS ON THE PUBLIC § 1320.17 Information collection budget. Each agency's Senior Official, or...

32 CFR 631.17 - Marine Corps policy.

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2010-07-01

... 32 National Defense 4 2010-07-01 2010-07-01 true Marine Corps policy. 631.17 Section 631.17... CRIMINAL INVESTIGATIONS ARMED FORCES DISCIPLINARY CONTROL BOARDS AND OFF-INSTALLATION LIAISON AND OPERATIONS Off-Installation Operations (Military Patrols and Investigative Activities) and Policy § 631.17...

7 CFR 15b.17 - Discrimination prohibited.

Science.gov (United States)

2010-01-01

... 7 Agriculture 1 2010-01-01 2010-01-01 false Discrimination prohibited. 15b.17 Section 15b.17... ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE Accessibility § 15b.17 Discrimination prohibited. No... to discrimination under any program or activity receiving assistance from this Department. ...

40 CFR 73.14-73.17 - [Reserved

Science.gov (United States)

2010-07-01

... 40 Protection of Environment 16 2010-07-01 2010-07-01 false [Reserved] 73.14-73.17 Section 73.14-73.17 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) AIR PROGRAMS (CONTINUED) SULFUR DIOXIDE ALLOWANCE SYSTEM Allowance Allocations §§ 73.14-73.17 [Reserved] ...

Estrogen Deficiency Promotes Cerebral Aneurysm Rupture by Upregulation of Th17 Cells and Interleukin-17A Which Downregulates E-Cadherin.

Science.gov (United States)

Hoh, Brian L; Rojas, Kelley; Lin, Li; Fazal, Hanain Z; Hourani, Siham; Nowicki, Kamil W; Schneider, Matheus B; Hosaka, Koji

2018-04-13

Estrogen deficiency is associated with the development of cerebral aneurysms; however, the mechanism remains unknown. We explored the pathway of cerebral aneurysm development by investigating the potential link between estrogen deficiency and inflammatory factors. First, we established the role of interleukin-17 (IL-17)A. We performed a cytokine screen demonstrating that IL-17A is significantly expressed in mouse and human aneurysms ( P =0.03). Likewise, IL-17A inhibition was shown to prevent aneurysm formation by 42% ( P =0.02) and rupture by 34% ( P <0.05). Second, we found that estrogen deficiency upregulates T helper 17 cells and IL-17A and promotes aneurysm rupture. Estrogen-deficient mice had more ruptures than control mice (47% versus 7%; P =0.04). Estradiol supplementation or IL-17A inhibition decreased the number of ruptures in estrogen-deficient mice (estradiol 6% versus 37%; P =0.04; IL-17A inhibition 18% versus 47%; P =0.018). Third, we found that IL-17A-blockade protects against aneurysm formation and rupture by increased E-cadherin expression. IL-17-inhibited mice had increased E-cadherin expression ( P =0.003). E-cadherin inhibition reversed the protective effect of IL-17A inhibition and increased the rate of aneurysm formation (65% versus 28%; P =0.04) and rupture (12% versus 0%; P =0.22). However, E-cadherin inhibition alone does not significantly increase aneurysm formation in normal mice or in estrogen-deficient mice. In cell migration assays, E-cadherin inhibition promoted macrophage infiltration across endothelial cells ( P <0.05), which may be the mechanism for the estrogen deficiency/IL-17/E-cadherin aneurysm pathway. Our data suggest that estrogen deficiency promotes cerebral aneurysm rupture by upregulating IL-17A, which downregulates E-cadherin, encouraging macrophage infiltration in the aneurysm vessel wall. © 2018 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley.

Anti-retroviral therapy fails to restore the severe Th-17: Tc-17 imbalance observed in peripheral blood during simian immunodeficiency virus infection.

Science.gov (United States)

Kader, M; Bixler, S; Piatak, M; Lifson, J; Mattapallil, J J

2009-10-01

Human immuno deficiency virus and simian immunodeficiency virus infections are characterized by a severe loss of Th-17 cells (IL-17(+)CD4(+) T cells) that has been associated with disease progression and systemic dissemination of bacterial infections. Anti-retroviral therapy (ART) has led to repopulation of CD4(+) T cells in peripheral tissues with little sustainable repopulation in mucosal tissues. Given the central importance of Th-17 cells in mucosal homeostasis, it is not known if the failure of ART to permanently repopulate mucosal tissues is associated with a failure to restore Th-17 cells that are lost during infection. Dynamics of alpha4(+)beta7(hi) CD4(+) T cells in peripheral blood of SIV infected rhesus macaques were evaluated and compared to animals that were treated with ART. The frequency of Th-17 and Tc-17 cells was determined following infection and after therapy. Relative expression of IL-21, IL-23, and TGFbeta was determined using Taqman PCR. Treatment of SIV infected rhesus macaques with anti-retroviral therapy was associated with a substantial repopulation of mucosal homing alpha4(+)beta7(hi)CD4(+) T cells in peripheral blood. This repopulation, however, was not accompanied by a restoration of Th-17 responses. Interestingly, SIV infection was associated with an increase in Tc-17 responses (IL-17(+)CD8(+) T cells) suggesting to a skewing in the ratio of Th-17: Tc-17 cells from a predominantly Th-17 phenotype to a predominantly Tc-17 phenotype. Surprisingly, Tc-17 responses remained high during the course of therapy suggesting that ART failed to correct the imbalance in Th-17 : Tc-17 responses induced following SIV infection. ART was associated with substantial repopulation of alpha4(+)beta7(hi) CD4(+) T cells in peripheral blood with little or no rebound of Th-17 cells. On the other hand, repopulation of alpha4(+)beta7(hi) CD4(+) T cells was accompanied by persistence of high levels of Tc-17 cells in peripheral blood. The dysregulation of Th-17

The excretory-secretory products of Echinococcus granulosus protoscoleces directly regulate the differentiation of B10, B17 and Th17 cells.

Science.gov (United States)

Pan, Wei; Hao, Wen-Ting; Shen, Yu-Juan; Li, Xiang-Yang; Wang, Yan-Juan; Sun, Fen-Fen; Yin, Jian-Hai; Zhang, Jing; Tang, Ren-Xian; Cao, Jian-Ping; Zheng, Kui-Yang

2017-07-21

Excretory-secretory products (ESPs) released by helminths are well-known to regulate T cell responses in the host. However, their direct influence in the differentiation of naïve T cells, and especially B cells, remains largely unknown. This study investigated the effects of Echinococcus granulosus protoscoleces ESPs (EgPSC-ESPs) on the differentiation of IL-10-producing B cells (B10), IL-17A-producing B cells (B17) and Th17 cells. BALB/c mice injected with EgPSC were used to evaluate the in vivo profiles of B10, B17 and Th17 cells. In vitro purified CD19 + B and naïve CD4 + T cells were cultured in the presence of native, heat-inactivated or periodate-treated EgPSC-ESPs, and the differentiation of these cell subsets were compared. In contrast to the control group, infected mice showed higher frequencies of B10, B17 and Th17 cells, and higher levels of IL-10 and IL-17A in the sera. Interestingly, B17 cells were first identified to express CD19 + CD1d high . In vitro, B cells cultured with native ESPs exhibited a higher percentage of B10 cells but lower percentage of B17 and Th17 cells compared to the PBS group. Moreover, the relative expression of IL-10 and IL-17A mRNA were consistent with the altered frequencies. However, ESPs subjected to heat-inactivation or periodate treatment exhibited an inverse effect on the induction of these cell subsets. Our findings indicate that ESPs released by EgPSC can directly regulate the differentiation of B10, B17 and Th17 cells, which appear to be heat-labile and carbohydrate-dependent.

44 CFR 17.620 - Effect of violation.

Science.gov (United States)

2010-10-01

... 44 Emergency Management and Assistance 1 2010-10-01 2010-10-01 false Effect of violation. 17.620 Section 17.620 Emergency Management and Assistance FEDERAL EMERGENCY MANAGEMENT AGENCY, DEPARTMENT OF HOMELAND SECURITY GENERAL GOVERNMENTWIDE REQUIREMENTS FOR DRUG-FREE WORKPLACE (GRANTS) § 17.620 Effect of...

Increased protein expression of LHCG receptor and 17α-hydroxylase/17-20-lyase in human polycystic ovaries.

Science.gov (United States)

Comim, F V; Teerds, K; Hardy, K; Franks, S

2013-11-01

Does the expression of LHCG receptor (LHCGR) protein and key enzymes in the androgen biosynthetic pathway differ in normal human versus polycystic ovarian tissue? LHCGR and 17α-hydroxylase/17-20-lyase (CYP17A1) protein levels are increased in polycystic ovaries (PCOs). The predominant source of excess androgen secretion in women with polycystic ovary syndrome (PCOS) is ovarian theca cells but few studies have directly assessed the presence and abundance of protein for key molecules involved in androgen production by theca, including LHCGR and the rate-limiting enzyme in androgen production, CYP17A1. This is a laboratory-based, cross-sectional study comparing protein expression of key molecules in the androgen biosynthetic pathway in archived ovarian tissue from women with normal ovaries (n = 10) with those with PCOs (n = 16). A quantitative morphometric study was performed using sections of archived human ovaries (n = 26) previously characterized as normal or polycystic. The distribution and abundance of LHCGR, CYP17A1, 3β-hydroxysteroid dehydrogenase type 2 (3βHSDII) and 17β-hydroxysteroid dehydrogenase type 5 (17βHSD5) proteins were evaluated by immunohistochemistry and quantified. A higher proportion of theca cells from anovulatory PCO expressed LHCGR protein when compared with control ovaries (P = 0.01). A significant increase in the intensity of immunostaining for CYP17A1 was identified in antral follicles in sections of PCO compared with ovaries from normal women (P = 0.04). As the study used formalin-fixed ovarian tissue sections, it was not possible to carry out studies 'in vitro' using the same ovarian tissues in order to also demonstrate increased functional activity of LHCGR and CYP17A1. The data are in keeping with the results of previous studies in isolated theca cells and support the notion of an intrinsic abnormality of theca cell androgen production in women with PCOS. The research was supported by a Programme Grant, G0802782, from the Medical

Interleukin-17 receptor A (IL-17RA) as a central regulator of the protective immune response against Giardia.

Science.gov (United States)

Paerewijck, Oonagh; Maertens, Brecht; Dreesen, Leentje; Van Meulder, Frederik; Peelaers, Iris; Ratman, Dariusz; Li, Robert W; Lubberts, Erik; De Bosscher, Karolien; Geldhof, Peter

2017-08-17

The protozoan parasite Giardia is a highly prevalent intestinal pathogen with a wide host range. Data obtained in mice, cattle and humans revealed the importance of IL-17A in the development of a protective immune response against Giardia. The aim of this study was to further unravel the protective effector mechanisms triggered by IL-17A following G. muris infection in mice, by an RNA-sequencing approach. C57BL/6 WT and C57BL/6 IL-17RA KO mice were orally infected with G. muris cysts. Three weeks post infection, intestinal tissue samples were collected for RNA-sequencing, with samples from uninfected C57BL/6 WT and C57BL/6 IL-17RA KO animals serving as negative controls. Differential expression analysis showed that G. muris infection evoked the transcriptional upregulation of a wide array of genes, mainly in animals with competent IL-17RA signaling. IL-17RA signaling induced the production of various antimicrobial peptides, such as angiogenin 4 and α- and β-defensins and regulated complement activation through mannose-binding lectin 2. The expression of the receptor that regulates the secretion of IgA into the intestinal lumen, the polymeric immunoglobulin receptor, was also dependent on IL-17RA signaling. Interestingly, the transcriptome data showed for the first time the involvement of the circadian clock in the host response following Giardia infection.

17-AAG, an Hsp90 inhibitor, causes kinetochore defects: a novel mechanism by which 17-AAG inhibits cell proliferation.

Science.gov (United States)

Niikura, Y; Ohta, S; Vandenbeldt, K J; Abdulle, R; McEwen, B F; Kitagawa, K

2006-07-13

The Hsp90 inhibitor 17-allylaminogeldanamycin (17-AAG), which is currently in clinical trials, is thought to exert antitumor activity by simultaneously targeting several oncogenic signaling pathways. Here we report a novel mechanism by which 17-AAG inhibits cell proliferation, and we provide the first evidence that HSP90 is required for the assembly of kinetochore protein complexes in humans. 17-AAG caused delocalization of several kinetochore proteins including CENP-I and CENP-H but excluding CENP-B and CENP-C. Consistently, 17-AAG induced a mitotic arrest that depends on the spindle checkpoint and induced misalignment of chromosomes and aneuploidy. We found that HSP90 associates with SGT1 (suppressor of G2 allele of skp1; SUGT1) in human cells and that depletion of SGT1 sensitizes HeLa cells to 17-AAG. Overexpression of SGT1 restored the localization of specific kinetochore proteins and chromosome alignment in cells treated with 17-AAG. Biochemical and genetic results suggest that HSP90, through its interaction with SGT1 (SUGT1), is required for kinetochore assembly. Furthermore, time-course experiments revealed that transient treatment with 17-AAG between late S and G2/M phases causes substantial delocalization of CENP-H and CENP-I, a finding that strongly suggests that HSP90 participates in kinetochore assembly in a cell cycle-dependent manner.

Cyanobacterial photosystem II at 2.9-A resolution and the role of quinones, lipids, channels and chloride

NARCIS (Netherlands)

Guskov, Albert; Kern, Jan; Gabdulkhakov, Azat; Broser, Matthias; Zouni, Athina; Saenger, Wolfram

Photosystem II (PSII) is a large homodimeric protein-cofactor complex located in the photosynthetic thylakoid membrane that acts as light-driven water:plastoquinone oxidoreductase. The crystal structure of PSII from Thermosynechococcus elongatus at 2.9-A resolution allowed the unambiguous assignment

38 CFR 17.180 - Delegation of authority.

Science.gov (United States)

2010-07-01

... 38 Pensions, Bonuses, and Veterans' Relief 1 2010-07-01 2010-07-01 false Delegation of authority. 17.180 Section 17.180 Pensions, Bonuses, and Veterans' Relief DEPARTMENT OF VETERANS AFFAIRS MEDICAL Veterans Canteen Service § 17.180 Delegation of authority. In connection with the Veterans Canteen Service...

37 CFR 385.17 - Effect of rates.

Science.gov (United States)

2010-07-01

... 37 Patents, Trademarks, and Copyrights 1 2010-07-01 2010-07-01 false Effect of rates. 385.17 Section 385.17 Patents, Trademarks, and Copyrights COPYRIGHT ROYALTY BOARD, LIBRARY OF CONGRESS RATES AND... Digital Phonorecord Deliveries and Limited Downloads § 385.17 Effect of rates. In any future proceedings...

Measuring device and method for dimples height differences of 17 x 17 grid

International Nuclear Information System (INIS)

Xu Yilan; Zheng Zhihui; Yan Liwei; Wang Xihe

2001-01-01

There are 264 cell for fastening fuel rods in the grid of 17 x 17 fuel assembly of PWR. The height differences of top and bottom dimples in a grid is an important quality characteristic of the grid. The report deals with measuring machine and method for dimples height differences of the grid. The device has two measuring probes. The Parallel Leaf Spring is used for transmitting the little displacement between two probes. The uncertainty of the device is σ≤4 μm. The measuring method is shown to be practicable

17 CFR 240.17a-6 - Right of national securities exchange, national securities association, registered clearing...

Science.gov (United States)

2010-04-01

... 17 Commodity and Securities Exchanges 3 2010-04-01 2010-04-01 false Right of national securities exchange, national securities association, registered clearing agency or the Municipal Securities... and Reports of Certain Stabilizing Activities § 240.17a-6 Right of national securities exchange...

Anti-IL-17 Antibody Improves Hepatic Steatosis by Suppressing Interleukin-17-Related Fatty Acid Synthesis and Metabolism

Directory of Open Access Journals (Sweden)

Weidong Shi

2013-01-01

Full Text Available To investigate the relationship between interleukin-17 and proteins involved in fatty acid metabolism with respect to alcoholic liver disease, male ICR mice were randomized into five groups: control, alcoholic liver disease (ALD at 4 weeks, 8 weeks, and 12 weeks, and anti-IL-17 antibody treated ALD. A proteomic approach was adopted to investigate changes in liver proteins between control and ALD groups. The proteomic analysis was performed by two-dimensional difference gel electrophoresis. Spots of interest were subsequently subjected to nanospray ionization tandem mass spectrometry (MS/MS for protein identification. Additionally, expression levels of selected proteins were confirmed by western blot. Transcriptional levels of some selected proteins were determined by RT-PCR. Expression levels of 95 protein spots changed significantly (ratio >1.5, P<0.05 during the development of ALD. Sterol regulatory element-binding protein-lc (SREBP-1c, carbohydrate response element binding protein (ChREBP, enoyl-coenzyme A hydratase (ECHS1, and peroxisome proliferator-activated receptor alpha (PPAR-α were identified by MS/MS among the proteins shown to vary the most; increased IL-17 elevated the transcription of SREBP-1c and ChREBP but suppressed ECHS1 and PPAR-α. The interleukin-17 signaling pathway is involved in ALD development; anti-IL-17 antibody improved hepatic steatosis by suppressing interleukin-17-related fatty acid metabolism.

50 CFR 17.41 - Special rules-birds.

Science.gov (United States)

2010-10-01

... 50 Wildlife and Fisheries 2 2010-10-01 2010-10-01 false Special rules-birds. 17.41 Section 17.41 Wildlife and Fisheries UNITED STATES FISH AND WILDLIFE SERVICE, DEPARTMENT OF THE INTERIOR (CONTINUED... rules—birds. (a) Bald eagles (Haliaeetus leucocephalus) wherever listed as threatened under § 17.11(h...

27 CFR 17.133 - Food product formulas.

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2010-04-01

... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Food product formulas. 17.133 Section 17.133 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU... PRODUCTS Formulas and Samples Approval of Formulas § 17.133 Food product formulas. Formulas for nonbeverage...

43 CFR 3.17 - Preservation of collection.

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2010-10-01

... 43 Public Lands: Interior 1 2010-10-01 2010-10-01 false Preservation of collection. 3.17 Section 3.17 Public Lands: Interior Office of the Secretary of the Interior PRESERVATION OF AMERICAN ANTIQUITIES § 3.17 Preservation of collection. Every collection made under the authority of the act and of...

14 CFR 13.17 - Seizure of aircraft.

Science.gov (United States)

2010-01-01

... 14 Aeronautics and Space 1 2010-01-01 2010-01-01 false Seizure of aircraft. 13.17 Section 13.17... INVESTIGATIVE AND ENFORCEMENT PROCEDURES Legal Enforcement Actions § 13.17 Seizure of aircraft. (a) Under... officer, or a Federal Aviation Administration safety inspector, authorized in an order of seizure issued...

6 CFR 13.17 - Rights of parties.

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2010-01-01

... 6 Domestic Security 1 2010-01-01 2010-01-01 false Rights of parties. 13.17 Section 13.17 Domestic Security DEPARTMENT OF HOMELAND SECURITY, OFFICE OF THE SECRETARY PROGRAM FRAUD CIVIL REMEDIES § 13.17 Rights of parties. Except as otherwise limited by this part, all parties may: (a) Be accompanied...

46 CFR 61.20-17 - Examination intervals.

Science.gov (United States)

2010-10-01

... 46 Shipping 2 2010-10-01 2010-10-01 false Examination intervals. 61.20-17 Section 61.20-17... INSPECTIONS Periodic Tests of Machinery and Equipment § 61.20-17 Examination intervals. (a) A lubricant that... examination interval. (b) Except as provided in paragraphs (c) through (f) of this section, each tailshaft on...

7 CFR 1221.17 - Net market value.

Science.gov (United States)

2010-01-01

... 7 Agriculture 10 2010-01-01 2010-01-01 false Net market value. 1221.17 Section 1221.17 Agriculture... INFORMATION ORDER Sorghum Promotion, Research, and Information Order Definitions § 1221.17 Net market value. Net market value means: (a) Except as provided in paragraph (b)and (c) of this section, the value...

11 CFR 110.17 - Price index increase.

Science.gov (United States)

2010-01-01

... 11 Federal Elections 1 2010-01-01 2010-01-01 false Price index increase. 110.17 Section 110.17... PROHIBITIONS § 110.17 Price index increase. (a) Price index increases for party committee expenditure... 11 CFR 109.32 and 110.8 shall be increased by the percent difference between the price index, as...