NF-κB p50 promotes tumor cell invasion through negative regulation of invasion suppressor gene CRMP-1 in human lung adenocarcinoma cells

International Nuclear Information System (INIS)

Gao Ming; Yeh, P.Y.; Lu, Y.-S.; Chang, W.C.; Kuo, M.-L.; Cheng, A.-L.

2008-01-01

Lung adenocarcinoma Cl1-5 cells were selected from parental Cl1-0 cells based on their high metastatic potential. In a previous study, CRMP-1, an invasion suppressor gene, was shown to be suppressed in Cl1-5 cells. However, the regulation of CRMP-1 expression has not been explored. In this study, we showed nuclear factor-κB controls CRMP-1 expression. The electromobility shift assay showed that while Cl1-0 cells exhibited low NF-κB activity in response to TNF-α, an abundance of basal and TNF-α-induced NF-κB-DNA complex was detected in Cl1-5 cells. Supershift-coupled EMSA and Western blotting of nuclear proteins, however, revealed p50 protein, but not classic p65/p50 heterodimer in the complex. ChIP and EMSA demonstrated that p50 binds to a κB site residing between -1753 and -1743 of the CRMP-1 promoter region. Transfection of antisense p50 gene into Cl1-5 cells increased the CRMP-1 protein level and decreased the invasive activity of Cl1-5 cells

Expression of RKIP, E-cadherin and NF-kB p65 in esophageal squamous cell carcinoma and their correlations.

Science.gov (United States)

Ping, Fu-Min; Liu, Gui-Jing; Liu, Zhi-Jun; Li, Hai-Bin; Zhai, Jian-Wen; Li, Shu-Xia; Liu, Yue-Mei; Li, Bao-Wei; Wei, Hong

2015-01-01

To detect the expression of RKIP, E-cadherin and NF-kB p65 in esophageal squamous cell carcinoma (ESCC) and study their correlations. Steptavidin-peroxidase (S-P) method was employed to detect the expressions of RKIP, E-cadherin and NF-kB p65 in ESCC tissues from 77 cases and paracancerous tissues from 77 cases. The correlations between their expressions and clinicopathological indices and between the expressions of these proteins themselves were analyzed. The expressions of RKIP and E-cadherin in ESCC tissues were obviously lower than those in the paracancerous tissues (PkB p65 in ESCC tissues was correlated with clinical staging, lymph node metastasis and tumor differentiation (PkB p65 in ESCC tissues (PkB p65.

Application of pH regulator 65 to wet processing of textiles%pH值控制剂65在印染加工中的应用

Institute of Scientific and Technical Information of China (English)

王力民; 李春光; 苏长智; 张万伟; 秦新波

2012-01-01

pH值控制剂65系有机酯和有机羧酸盐复配而成,它能够完全中和纤维内部的碱剂而不损伤织物强力,将布面的pH值控制在7.0～7.5.pH值控制剂65可解决敏感色织物、增白织物的pH值控制以及织物泛黄等问题,且与各种功能性整理剂相容性好.%pH regulator 65 is a complex of organic ester and organic carboxylate. The pH regulator 65 can completely neutralize the base in the internal fiber and stablize the pH value of fabric surface at 7～7.5 without strength loss. It can solve the problems of pH controlling and fabric yellowing in processing of sensitive and whitening fabric, and has good compatibility with all kinds of functional finishing agents.

Targeting NF-κB RelA/p65 phosphorylation overcomes RITA resistance.

Science.gov (United States)

Bu, Yiwen; Cai, Guoshuai; Shen, Yi; Huang, Chenfei; Zeng, Xi; Cao, Yu; Cai, Chuan; Wang, Yuhong; Huang, Dan; Liao, Duan-Fang; Cao, Deliang

2016-12-28

Inactivation of p53 occurs frequently in various cancers. RITA is a promising anticancer small molecule that dissociates p53-MDM2 interaction, reactivates p53 and induces exclusive apoptosis in cancer cells, but acquired RITA resistance remains a major drawback. This study found that the site-differential phosphorylation of nuclear factor-κB (NF-κB) RelA/p65 creates a barcode for RITA chemosensitivity in cancer cells. In naïve MCF7 and HCT116 cells where RITA triggered vast apoptosis, phosphorylation of RelA/p65 increased at Ser536, but decreased at Ser276 and Ser468; oppositely, in RITA-resistant cells, RelA/p65 phosphorylation decreased at Ser536, but increased at Ser276 and Ser468. A phosphomimetic mutation at Ser536 (p65/S536D) or silencing of endogenous RelA/p65 resensitized the RITA-resistant cells to RITA while the phosphomimetic mutant at Ser276 (p65/S276D) led to RITA resistance of naïve cells. In mouse xenografts, intratumoral delivery of the phosphomimetic p65/S536D mutant increased the antitumor activity of RITA. Furthermore, in the RITA-resistant cells ATP-binding cassette transporter ABCC6 was upregulated, and silencing of ABCC6 expression in these cells restored RITA sensitivity. In the naïve cells, ABCC6 delivery led to RITA resistance and blockage of p65/S536D mutant-induced RITA sensitivity. Taken together, these data suggest that the site-differential phosphorylation of RelA/p65 modulates RITA sensitivity in cancer cells, which may provide an avenue to manipulate RITA resistance. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

SIRT1 overexpression decreases cisplatin-induced acetylation of NF-κB p65 subunit and cytotoxicity in renal proximal tubule cells

International Nuclear Information System (INIS)

Jung, Yu Jin; Lee, Jung Eun; Lee, Ae Sin; Kang, Kyung Pyo; Lee, Sik; Park, Sung Kwang; Lee, Sang Yong; Han, Myung Kwan; Kim, Duk Hoon; Kim, Won

2012-01-01

Highlights: ► Cisplatin increases acetylation of NF-κB p65 subunit in HK2 cells. ► SIRT1 overexpression decreases cisplatin-induced p65 acetylation and -cytotoxicity. ► Resveratrol decreased cisplatin-induced cell viability through deacetylation of p65. -- Abstract: As the increased acetylation of p65 is linked to nuclear factor-κB (NF-κB) activation, the regulation of p65 acetylation can be a potential target for the treatment of inflammatory injury. Cisplatin-induced nephrotoxicity is an important issue in chemotherapy of cancer patients. SIRT1, nicotinamide adenine dinucleotide (NAD + )-dependent protein deacetylase, has been implicated in a variety of cellular processes such as inflammatory injury and the control of multidrug resistance in cancer. However, there is no report on the effect of SIRT1 overexpression on cisplatin-induced acetylation of p65 subunit of NF-κB and cell injury. To investigate the effect of SIRT1 in on cisplatin-induced acetylation of p65 subunit of NF-κB and cell injury, HK2 cells were exposed with SIRT1 overexpression, LacZ adenovirus or dominant negative adenovirus after treatment with cisplatin. While protein expression of SIRT1 was decreased by cisplatin treatment compared with control buffer treatment, acetylation of NF-κB p65 subunit was significantly increased after treatment with cisplatin. Overexpression of SIRT1 ameliorated the increased acetylation of p65 of NF-κB during cisplatin treatment and cisplatin-induced cytotoxicity. Further, treatment of cisplatin-treated HK2 cells with resveratrol, a SIRT1 activator, also decreased acetylation of NF-κB p65 subunit and cisplatin-induced increase of the cell viability in HK2 cells. Our findings suggests that the regulation of acetylation of p65 of NF-κB through SIRT1 can be a possible target to attenuate cisplatin-induced renal cell damage.

Nuclear NF-κB p65 in peripheral blood mononuclear cells correlates with urinary MCP-1, RANTES and the severity of type 2 diabetic nephropathy.

Directory of Open Access Journals (Sweden)

Bin Yi

Full Text Available AIMS: To investigate if nuclear NF-κB p65 expression in ex vivo isolated peripheral blood mononuclear cells correlates with urinary MCP-1 or RANTES and the severity of type 2 diabetic nephropathy. METHODS: According to their urinary albumin-to-creatinine ratio (uACR, 107 patients with type 2 diabetes (eGFR >60 ml/min were divided into normal albuminuria group (DN0 group, 38 cases, microalbuminuria group (DN1 group, 38 cases, and macroalbuminuria group (DN2 group, 31 cases, compared with matched healthy normal control group (NC group, 30 cases. Nuclear NF-κB p65 protein expression levels in peripheral blood mononuclear cells were detected by western blotting. Real-time quantitative polymerase chain reaction was used to detect NF-κB p65 mRNA expression and ELISA assay was used to detect the levels of urinary MCP-1 and RANTES. RESULTS: Nuclear NF-κB p65 protein and NF-κB p65 mRNA expression levels in peripheral blood mononuclear cells, urinary MCP-1/Cr and RANTES/Cr were all significantly higher in all diabetes groups as compared with NC group. In particular, the increase of nuclear NF-κB p65 protein and NF-κB p65 mRNA expressions, urinary MCP-1/Cr and RANTES/Cr all correlated with the severity of type 2 diabetic nephropathy as indicated by the increase in uACR. Pearson correlation analysis indicated that both urinary MCP-1/Cr and RANTES/Cr were positively correlated with nuclear NF-κB p65 protein or NF-κB p65 mRNA levels. Stepwise multiple regression analysis showed that nuclear NF-κB p65 protein or NF-κB p65 mRNA was an independent variable for urinary MCP-1/Cr, and MCP-1/Cr and RANTES/Cr were two independent variables for uACR. CONCLUSION: Our research demonstrates that nuclear NF-κB p65 protein and mRNA expressions in ex vivo isolated peripheral blood mononuclear cells well correlate with urinary MCP-1/Cr, RANTES/Cr and the severity of type 2 diabetic nephropathy.

Eukaryotic Initiation Factor 4H Is under Transcriptional Control of p65/NF-κB

Science.gov (United States)

Fiume, Giuseppe; Rossi, Annalisa; de Laurentiis, Annamaria; Falcone, Cristina; Pisano, Antonio; Vecchio, Eleonora; Pontoriero, Marilena; Scala, Iris; Scialdone, Annarita; Masci, Francesca Fasanella; Mimmi, Selena; Palmieri, Camillo; Scala, Giuseppe; Quinto, Ileana

2013-01-01

Protein synthesis is mainly regulated at the initiation step, allowing the fast, reversible and spatial control of gene expression. Initiation of protein synthesis requires at least 13 translation initiation factors to assemble the 80S ribosomal initiation complex. Loss of translation control may result in cell malignant transformation. Here, we asked whether translational initiation factors could be regulated by NF-κB transcription factor, a major regulator of genes involved in cell proliferation, survival, and inflammatory response. We show that the p65 subunit of NF-κB activates the transcription of eIF4H gene, which is the regulatory subunit of eIF4A, the most relevant RNA helicase in translation initiation. The p65-dependent transcriptional activation of eIF4H increased the eIF4H protein content augmenting the rate of global protein synthesis. In this context, our results provide novel insights into protein synthesis regulation in response to NF-κB activation signalling, suggesting a transcription-translation coupled mechanism of control. PMID:23776612

Tumour suppressor protein p53 regulates the stress activated bilirubin oxidase cytochrome P450 2A6

Energy Technology Data Exchange (ETDEWEB)

Hu, Hao, E-mail: hao.hu1@uqconnect.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Yu, Ting, E-mail: t.yu2@uq.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Arpiainen, Satu, E-mail: Satu.Juhila@orion.fi [Institute of Biomedicine, Department of Pharmacology and Toxicology and Medical Research Center Oulu, Oulu University Hospital and University of Oulu, Oulu (Finland); Lang, Matti A., E-mail: m.lang@uq.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Hakkola, Jukka, E-mail: Jukka.hakkola@oulu.fi [Institute of Biomedicine, Department of Pharmacology and Toxicology and Medical Research Center Oulu, Oulu University Hospital and University of Oulu, Oulu (Finland); Abu-Bakar, A' edah, E-mail: a.abubakar@uq.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia)

2015-11-15

Human cytochrome P450 (CYP) 2A6 enzyme has been proposed to play a role in cellular defence against chemical-induced oxidative stress. The encoding gene is regulated by various stress activated transcription factors. This paper demonstrates that p53 is a novel transcriptional regulator of the gene. Sequence analysis of the CYP2A6 promoter revealed six putative p53 binding sites in a 3 kb proximate promoter region. The site closest to transcription start site (TSS) is highly homologous with the p53 consensus sequence. Transfection with various stepwise deletions of CYP2A6-5′-Luc constructs – down to − 160 bp from the TSS – showed p53 responsiveness in p53 overexpressed C3A cells. However, a further deletion from − 160 to − 74 bp, including the putative p53 binding site, totally abolished the p53 responsiveness. Electrophoretic mobility shift assay with a probe containing the putative binding site showed specific binding of p53. A point mutation at the binding site abolished both the binding and responsiveness of the recombinant gene to p53. Up-regulation of the endogenous p53 with benzo[α]pyrene – a well-known p53 activator – increased the expression of the p53 responsive positive control and the CYP2A6-5′-Luc construct containing the intact p53 binding site but not the mutated CYP2A6-5′-Luc construct. Finally, inducibility of the native CYP2A6 gene by benzo[α]pyrene was demonstrated by dose-dependent increases in CYP2A6 mRNA and protein levels along with increased p53 levels in the nucleus. Collectively, the results indicate that p53 protein is a regulator of the CYP2A6 gene in C3A cells and further support the putative cytoprotective role of CYP2A6. - Highlights: • CYP2A6 is an immediate target gene of p53. • Six putative p53REs located on 3 kb proximate CYP2A6 promoter region. • The region − 160 bp from TSS is highly homologous with the p53 consensus sequence. • P53 specifically bind to the p53RE on the − 160 bp region. • HNF4�

SIRT1 overexpression decreases cisplatin-induced acetylation of NF-{kappa}B p65 subunit and cytotoxicity in renal proximal tubule cells

Energy Technology Data Exchange (ETDEWEB)

Jung, Yu Jin; Lee, Jung Eun; Lee, Ae Sin [Department of Internal Medicine, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Kang, Kyung Pyo; Lee, Sik; Park, Sung Kwang [Department of Internal Medicine, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Institute for Medical Sciences, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Lee, Sang Yong [Department of Diagnostic Radiology, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Institute for Medical Sciences, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Han, Myung Kwan [Department of Microbiology, Institute for Medical Sciences, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Kim, Duk Hoon [Division of Forensic Medicine, National Forensic Service, Seoul (Korea, Republic of); Kim, Won, E-mail: kwon@jbnu.ac.kr [Department of Internal Medicine, Chonbuk National University Medical School, Jeonju (Korea, Republic of); Institute for Medical Sciences, Chonbuk National University Medical School, Jeonju (Korea, Republic of)

2012-03-09

Highlights: Black-Right-Pointing-Pointer Cisplatin increases acetylation of NF-{kappa}B p65 subunit in HK2 cells. Black-Right-Pointing-Pointer SIRT1 overexpression decreases cisplatin-induced p65 acetylation and -cytotoxicity. Black-Right-Pointing-Pointer Resveratrol decreased cisplatin-induced cell viability through deacetylation of p65. -- Abstract: As the increased acetylation of p65 is linked to nuclear factor-{kappa}B (NF-{kappa}B) activation, the regulation of p65 acetylation can be a potential target for the treatment of inflammatory injury. Cisplatin-induced nephrotoxicity is an important issue in chemotherapy of cancer patients. SIRT1, nicotinamide adenine dinucleotide (NAD{sup +})-dependent protein deacetylase, has been implicated in a variety of cellular processes such as inflammatory injury and the control of multidrug resistance in cancer. However, there is no report on the effect of SIRT1 overexpression on cisplatin-induced acetylation of p65 subunit of NF-{kappa}B and cell injury. To investigate the effect of SIRT1 in on cisplatin-induced acetylation of p65 subunit of NF-{kappa}B and cell injury, HK2 cells were exposed with SIRT1 overexpression, LacZ adenovirus or dominant negative adenovirus after treatment with cisplatin. While protein expression of SIRT1 was decreased by cisplatin treatment compared with control buffer treatment, acetylation of NF-{kappa}B p65 subunit was significantly increased after treatment with cisplatin. Overexpression of SIRT1 ameliorated the increased acetylation of p65 of NF-{kappa}B during cisplatin treatment and cisplatin-induced cytotoxicity. Further, treatment of cisplatin-treated HK2 cells with resveratrol, a SIRT1 activator, also decreased acetylation of NF-{kappa}B p65 subunit and cisplatin-induced increase of the cell viability in HK2 cells. Our findings suggests that the regulation of acetylation of p65 of NF-{kappa}B through SIRT1 can be a possible target to attenuate cisplatin-induced renal cell damage.

Atomic structure of the sweet-tasting protein thaumatin I at pH 8.0 reveals the large disulfide-rich region in domain II to be sensitive to a pH change.

OpenAIRE

Masuda, Tetsuya; Ohta, Keisuke; Mikami, Bunzo; Kitabatake, Naofumi; Tani, Fumito

2012-01-01

Thaumatin, an intensely sweet-tasting plant protein, elicits a sweet taste at 50 nM. Although the sweetness remains when thaumatin is heated at 80 °C for 4h under acid conditions, it rapidly declines when heating at a pH above 6.5. To clarify the structural difference at high pH, the atomic structure of a recombinant thaumatin I at pH 8.0 was determined at a resolution of 1.0Å. Comparison to the crystal structure of thaumatin at pH 7.3 and 7.0 revealed the root-mean square deviation value of ...

Citrullination of NF-κB p65 promotes its nuclear localization and TLR-induced expression of IL-1β and TNFα.

Science.gov (United States)

Sun, Bo; Dwivedi, Nishant; Bechtel, Tyler J; Paulsen, Janet L; Muth, Aaron; Bawadekar, Mandar; Li, Gang; Thompson, Paul R; Shelef, Miriam A; Schiffer, Celia A; Weerapana, Eranthie; Ho, I-Cheng

2017-06-09

Many citrullinated proteins are known autoantigens in rheumatoid arthritis, a disease mediated by inflammatory cytokines, such as tumor necrosis factor-α (TNFα). Citrullinated proteins are generated by converting peptidylarginine to peptidylcitrulline, a process catalyzed by the peptidylarginine deiminases (PADs), including PAD1 to PAD4 and PAD6. Several major risk factors for rheumatoid arthritis are associated with heightened citrullination. However, the physiological role of citrullination in immune cells is poorly understood. We report that suppression of PAD activity attenuates Toll-like receptor-induced expression of interleukin-1β (IL-1β) and TNFα by neutrophils in vivo and in vitro but not their global transcription activity. Mechanistically, PAD4 directly citrullinates nuclear factor κB (NF-κB) p65 and enhances the interaction of p65 with importin α3, which brings p65 into the nucleus. The citrullination-enhanced interaction of p65 with importin α3 and its nuclear translocation and transcriptional activity can be attributed to citrullination of four arginine residues located in the Rel homology domain of p65. Furthermore, a rheumatoid arthritis-prone variant of PAD4, carrying three missense mutations, is more efficient in interacting with p65 and enhancing NF-κB activity. Together, these data not only demonstrate a critical role of citrullination in an NF-κB-dependent expression of IL-1β and TNFα but also provide a molecular mechanism by which heightened citrullination propagates inflammation in rheumatoid arthritis. Accordingly, attenuating p65-mediated production of IL-1β and TNFα by blocking the citrullination of p65 has great therapeutic potential in rheumatoid arthritis. Copyright © 2017, American Association for the Advancement of Science.

The novel fusion proteins, GnRH-p53 and GnRHIII-p53, expression and their anti-tumor effect.

Directory of Open Access Journals (Sweden)

Peiyuan Jia

Full Text Available p53, one of the most well studied tumor suppressor factor, is responsible to a variety of damage owing to the induction of apoptosis and cell cycle arrest in the tumor cells. More than 50% of human tumors contain mutation or deletion of p53. Gonadotrophin-releasing hormone (GnRH, as the ligand of Gonadotrophin-releasing hormone receptor (GnRH-R, was used to deliver p53 into tumor cells. The p53 fusion proteins GnRH-p53 and GnRH iii-p53 were expressed and their targeted anti-tumor effects were determined. GnRH mediates its fusion proteins transformation into cancer cells. The intracellular delivery of p53 fusion proteins exerted the inhibition of the growth of H1299 cells in vitro and the reduction of tumor volume in vivo. Their anti-tumor effect was functioned by the apoptosis and cell cycle arrest induced by p53. Hence, the fusion protein could be a novel protein drug for anti-tumor therapy.

Expression and significance of HMGB1, TLR4 and NF-κB p65 in human epidermal tumors

International Nuclear Information System (INIS)

Weng, Hui; Deng, Yunhua; Xie, Yuyan; Liu, Hongbo; Gong, Feili

2013-01-01

High mobility group protein box 1 (HMGB1) is a DNA binding protein located in nucleus. It is released into extracellular fluid where it acts as a novel proinflammatory cytokine which interacts with Toll like receptor 4 (TLR4) to activate nuclear factor-κB (NF-κB). This sequence of events is involved in tumor growth and progression. However, the effects of HMGB1, TLR4 and NF-κB on epidermal tumors remain unclear. Human epidermal tumor specimens were obtained from 96 patients. Immunohistochemistry was used to detect expression of HMGB1, TLR4 and NF-κB p65 in human epidermal tumor and normal skin specimens. Western blot analysis was used to detect the expression of NF-κB p65 in epithelial cell nuclei in human epidermal tumor and normal tissues. Immunohistochemistry and western blot analysis indicated a progressive but statistically significant increase in p65 expression in epithelial nuclei in benign seborrheic keratosis (SK), precancerous lesions (PCL), low malignancy basal cell carcinoma (BCC) and high malignancy squamous cell carcinoma (SCC) (P <0.01). The level of extracellular HMGB1 in SK was significantly higher than in normal skin (NS) (P <0.01), and was higher than in SCC but without statistical significance. The level of TLR4 on epithelial membranes of SCC cells was significantly higher than in SK, PCL, BCC and NS (P <0.01). There was a significant positive correlation between p65 expression in the epithelial nuclei and TLR4 expression on the epithelial cell membranes (r = 0.3212, P <0.01). These findings indicate that inflammation is intensified in parallel with increasing malignancy. They also indicate that the TLR4 signaling pathway, rather than HMGB1, may be the principal mediator of inflammation in high-grade malignant epidermal tumors. Combined detection of p65 in the epithelial nuclei and TLR4 on the epithelial membranes may assist the accurate diagnosis of malignant epidermal tumors

Modulation of P1798 lymphosarcoma proliferation by protein phosphorylation

International Nuclear Information System (INIS)

Michnoff, C.A.H.

1983-01-01

The role of protein kinases in modulating cell proliferation was examined. Studies characterized the regulation of cell proliferation by adenosine 3':5'-monophosphate-dependent protein kinase (cA-Pk). Calcium/calmodulin-dependent myosin light chain kinase (MLCK) was isolated and examined as a potential substrate regulated by cA-PK in the rapidly proliferating P1798 lymphosarcoma. Modulation of cell proliferation by cA-PK was characterized by quantitating cell division by [methyl- 3 H] thymidine ([ 3 H]-dT) incorporation into DNA, cAMP accumulations, and activation of cA-PK using P1798 lymphosarcoma cells. Epinephrine and prostaglandin E 1 (PGE 1 ) were demonstrated to suppress [ 3 H]-dT incorporation into DNA, to stimulate cAMP accumulation, and to activate cA-PK with dose-dependency. Calcium/calmodulin-dependent MLCK was partially purified from P1798 lymphosarcoma. P1798 MLCK phosphorylated myosin regulatory light chains (P-LC) from thymus, cardiac and skeletal muscles. One mol [ 32 Pi] was transferred into one mol cardiac or skeletal P-LC by P1798 MLCK. Apparent Km values of 65 μM and 51 μM were determined for ATP and cardiac P-LC, respectively. The apparent molecular weight of P1798 MLCK was 135,000. P1798 MLCK was phosphorylated by cA-PK. Phosphorylated MLCK showed a 41% decrease in calcium-dependent activity. Two additional protein kinases from P1798 lymphosarcoma phosphorylated cardiac and skeletal light chains

DNM3, p65 and p53 from exosomes represent potential clinical diagnosis markers for glioblastoma multiforme

Science.gov (United States)

Yang, Jian-kai; Song, Jian; Huo, Hao-ran; Zhao, Yin-long; Zhang, Guang-yu; Zhao, Zong-mao; Sun, Guo-zhu; Jiao, Bao-hua

2017-01-01

Background: Glioblastoma multiforme (GBM) is the most aggressive and deadly primary brain cancer that arises from astrocytes and classified as grade IV. Recently, exosomes have been reported as an essential mediator in diverse cancer carcinogenesis and metastasis. However, their role in GBM is still unclear. In this study, we aimed to investigate whether blood exosomes can be potential clinical diagnostic markers for GBM. Methods: We used a xenograft orthotopic mouse model to detect the differentially expressed genes in the brain and blood exosomes of original/recurrent GBM. Results: We found that recurrent GBM had stronger growth capacity and lethality than original GBM in the mouse model. A gene microarray of original tumors and blood exosomes from GBM orthotopic xenografts results showed that DNM3, p65 and CD117 expressions increased, whereas PTEN and p53 expressions decreased in both original tumors and blood exosomes. In the recurrent GBM tumor model, DNM3 and p65 showed increased expressions, whereas ST14 and p53 showed decreased expressions in tumor and blood exosomes of the recurrent GBM mouse model. Conclusion: In summary, we found that DNM3, p65 and p53 had a similar trend in brain and blood exosomes both for original and recurrent GBM, and could serve as potential clinical diagnostic markers for GBM. PMID:29449895

(5'-32P)-8-azidoguanosine-3',5'-monophosphate. I. Synthesis and properties. II. Interaction with E. coli proteins

International Nuclear Information System (INIS)

Owens, J.R.

1983-01-01

Under certain conditions of nutritional deprivation, microorganisms produce the magic spot nucleotides guanosine-3'-diphosphate-5'-triphosphate(pppGpp) and the tetraphosphate ppGpp. The latter is known to be a pleiotypic effector, i.e. it inhibits (and sometimes stimulates) many biological processes including transcription, translation, and metabolic pathways. It is unknown whether pppGpp, ppGp, pGpp, and pGp, other members of this family of guanosine-3',5'-phosphates, also have regulatory properties. To begin to investigate this question, a radioactive photoaffinity analog of pGp was prepared: (5' 32 P)pN 3 Gp. The interaction of this photoprobe with E. coli sonicates and a purified protein (RNA polymerase) was examined. At physiological salt concentrations two proteins (RNA polymerase) was examined. At physiological salt concentrations two proteins of 86,000 and 65,000 daltons (p86 and p65) were primarily photolabeled. Competition studies with guanosine and adenosine nucleotides indicated (5 32 P)pN 3 Gp was labeling a ppGpp binding site on p86, and a pGp (or GMP) site on p65. ATP phosphorylation of p86 increased photoincorporation, while it decreased labeling of p65. The data also provide evidence of a different type of regulatory mechanism, i.e. phosphorylation modulates binding of an allosteric effector (ppGpp) to a protein(enzyme). Both ATP and GTP were found to phosphorylate the same proteins, although GTP was the preferred substrate in some cases

Valproic acid exposure decreases Cbp/p300 protein expression and histone acetyltransferase activity in P19 cells

Energy Technology Data Exchange (ETDEWEB)

Lamparter, Christina L. [Department of Biomedical and Molecular Sciences, Graduate Program in Pharmacology and Toxicology, Queen' s University, Kingston, Ontario K7L 3N6 (Canada); Winn, Louise M., E-mail: winnl@queensu.ca [Department of Biomedical and Molecular Sciences, Graduate Program in Pharmacology and Toxicology, Queen' s University, Kingston, Ontario K7L 3N6 (Canada); School of Environmental Studies, Queen' s University, Kingston, Ontario K7L 3N6 (Canada)

2016-09-01

The teratogenicity of the antiepileptic drug valproic acid (VPA) is well established and its inhibition of histone deacetylases (HDAC) is proposed as an initiating factor. Recently, VPA-mediated HDAC inhibition was demonstrated to involve transcriptional downregulation of histone acetyltransferases (HATs), which was proposed to compensate for the increased acetylation resulting from HDAC inhibition. Cbp and p300 are HATs required for embryonic development and deficiencies in either are associated with congenital malformations and embryolethality. The objective of the present study was to characterize Cbp/p300 following VPA exposure in P19 cells. Consistent with previous studies, exposure to 5 mM VPA over 24 h induced a moderate decrease in Cbp/p300 mRNA, which preceded a strong decrease in total cellular protein mediated by ubiquitin-proteasome degradation. Nuclear Cbp/p300 protein was also decreased following VPA exposure, although to a lesser extent. Total cellular and nuclear p300 HAT activity was reduced proportionately to p300 protein levels, however while total cellular HAT activity also decreased, nuclear HAT activity was unaffected. Using the Cbp/p300 HAT inhibitor C646, we demonstrated that HAT inhibition similarly affected many of the same endpoints as VPA, including increased reactive oxygen species and caspase-3 cleavage, the latter of which could be attenuated by pre-treatment with the antioxidant catalase. C646 exposure also decreased NF-κB/p65 protein, which was not due to reduced mRNA and was not attenuated with catalase pre-treatment. This study provides support for an adaptive HAT response following VPA exposure and suggests that reduced Cbp/p300 HAT activity could contribute to VPA-mediated alterations. - Highlights: • VPA exposure in vitro downregulates Cbp/p300 mRNA and induces protein degradation. • Cbp/p300 histone acetyltransferase activity is similarly reduced with VPA exposure. • Inhibition of Cbp/p300 acetyltransferase activity

Normal P50 gating in children with autism, yet attenuated P50 amplitude in the Asperger subcategory

DEFF Research Database (Denmark)

Madsen, Gitte; Bilenberg, Niels; Jepsen, Jens Richardt Møllegaard

2015-01-01

Autism spectrum disorders (ASD) and schizophrenia are separate disorders, but there is evidence of conversion or comorbid overlap. The objective of this paper was to explore whether deficits in sensory gating, as seen in some schizophrenia patients, can also be found in a group of ASD children...... compared to neurotypically developed children. An additional aim was to investigate the possibility of subdividing our ASD sample based on these gating deficits. In a case-control design, we assessed gating of the P50 and N100 amplitude in 31 ASD children and 39 healthy matched controls (8-12 years......) and screened for differences between groups and within the ASD group. We did not find disturbances in auditory P50 and N100 filtering in the group of ASD children as a whole, nor did we find abnormal P50 and N100 amplitudes. However, the P50 amplitude to the conditioning stimulus was significantly reduced...

Baicalin attenuates focal cerebral ischemic reperfusion injury through inhibition of nuclear factor κB p65 activation

International Nuclear Information System (INIS)

Xue, Xia; Qu, Xian-Jun; Yang, Ying; Sheng, Xie-Huang; Cheng, Fang; Jiang, E-Nang; Wang, Jian-hua; Bu, Wen; Liu, Zhao-Ping

2010-01-01

Research highlights: → Permanent NF-κB p65 activation contributes to the infarction after ischemia-reperfusion injury in rats. → Baicalin can markedly inhibit the nuclear NF-κB p65 expression and m RNA levels after ischemia-reperfusion injury in rats. → Baicalin decreased the cerebral infarction area via inhibiting the activation of nuclear NF-κB p65. -- Abstract: Baicalin is a flavonoid compound purified from plant Scutellaria baicalensis Georgi. We aimed to evaluate the neuroprotective effects of baicalin against cerebral ischemic reperfusion injury. Male Wistar rats were subjected to middle cerebral artery occlusion (MCAO) for 2 h followed by reperfusion for 24 h. Baicalin at doses of 50, 100 and 200 mg/kg was intravenously injected after ischemia onset. Twenty-four hours after reperfusion, the neurological deficit was scored and infarct volume was measured. Hematoxylin and eosin (HE) staining was performed to analyze the histopathological changes of cortex and hippocampus neurons. We examined the levels of NF-κB p65 in ischemic cortexes by Western blot analysis and RT-PCR assay. The results showed that the neurological deficit scores were significantly decreased from 2.0 ± 0.7 to 1.2 ± 0.4 and the volume of infarction was reduced by 25% after baicalin injection. Histopathological examination showed that the increase of neurons with pycnotic shape and condensed nuclear in cortex and hippocampus were not observed in baicalin treated animals. Further examination showed that NF-κB p65 in cortex was increased after ischemia reperfusion injury, indicating the molecular mechanism of ischemia reperfusion injury. The level of NF-κB p65 was decreased by 73% after baicalin treatment. These results suggest that baicalin might be useful as a potential neuroprotective agent in stroke therapy. The neuroprotective effects of baicalin may relate to inhibition of NF-κB p65.

Normal P50 Gating in Children with Autism, Yet Attenuated P50 Amplitude in the Asperger Subcategory.

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Madsen, Gitte Falcher; Bilenberg, Niels; Jepsen, Jens Richardt; Glenthøj, Birte; Cantio, Cathriona; Oranje, Bob

2015-08-01

Autism spectrum disorders (ASD) and schizophrenia are separate disorders, but there is evidence of conversion or comorbid overlap. The objective of this paper was to explore whether deficits in sensory gating, as seen in some schizophrenia patients, can also be found in a group of ASD children compared to neurotypically developed children. An additional aim was to investigate the possibility of subdividing our ASD sample based on these gating deficits. In a case-control design, we assessed gating of the P50 and N100 amplitude in 31 ASD children and 39 healthy matched controls (8-12 years) and screened for differences between groups and within the ASD group. We did not find disturbances in auditory P50 and N100 filtering in the group of ASD children as a whole, nor did we find abnormal P50 and N100 amplitudes. However, the P50 amplitude to the conditioning stimulus was significantly reduced in the Asperger subgroup compared to healthy controls. In contrast to what is usually reported for patients with schizophrenia, we found no evidence for sensory gating deficits in our group of ASD children taken as a whole. However, reduced P50 amplitude to conditioning stimuli was found in the Asperger group, which is similar to what has been described in some studies in schizophrenia patients. There was a positive correlation between the P50 amplitude of the conditioning stimuli and anxiety score in the pervasive developmental disorder not otherwise specified group, which indicates a relation between anxiety and sensory registration in this group. © 2015 International Society for Autism Research, Wiley Periodicals, Inc.

File list: InP.PSC.50.AllAg.mESC_derived_neural_cells [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.PSC.50.AllAg.mESC_derived_neural_cells mm9 Input control Pluripotent stem cell mESC derived neural...65,SRX1214070,SRX810564,SRX101693,SRX604259,SRX018672,SRX276677 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.PSC.50.AllAg.mESC_derived_neural_cells.bed ...

Exposure to coplanar PCBs induces endothelial cell inflammation through epigenetic regulation of NF-κB subunit p65

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Liu, Dandan; Perkins, Jordan T. [Superfund Research Center, University of Kentucky, Lexington, KY 40536 (United States); Department of Animal and Food Sciences, College of Agriculture, Food and Environment, University of Kentucky, Lexington, KY 40536 (United States); Petriello, Michael C. [Superfund Research Center, University of Kentucky, Lexington, KY 40536 (United States); Graduate Center for Toxicology and Cancer Biology, College of Medicine, University of Kentucky, Lexington, KY 40536 (United States); Hennig, Bernhard, E-mail: bhennig@uky.edu [Superfund Research Center, University of Kentucky, Lexington, KY 40536 (United States); Department of Animal and Food Sciences, College of Agriculture, Food and Environment, University of Kentucky, Lexington, KY 40536 (United States)

2015-12-15

Epigenetic modifications of DNA and histones alter cellular phenotypes without changing genetic codes. Alterations of epigenetic marks can be induced by exposure to environmental pollutants and may contribute to associated disease risks. Here we test the hypothesis that endothelial cell dysfunction induced by exposure to polychlorinated biphenyls (PCBs) is mediated in part though histone modifications. In this study, human vascular endothelial cells were exposed to physiologically relevant concentrations of several PCBs congeners (e.g., PCBs 77, 118, 126 and 153) followed by quantification of inflammatory gene expression and changes of histone methylation. Only exposure to coplanar PCBs 77 and 126 induced the expression of histone H3K9 trimethyl demethylase jumonji domain-containing protein 2B (JMJD2B) and nuclear factor-kappa B (NF-κB) subunit p65, activated NF-κB signaling as evidenced by nuclear translocation of p65, and up-regulated p65 target inflammatory genes, such as interleukin (IL)-6, C-reactive protein (CRP), intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and IL-1α/β. The increased accumulation of JMJD2B in the p65 promoter led to a depletion of H3K9me3 repression mark, which accounts for the observed up-regulation of p65 and associated inflammatory genes. JMJD2B gene knockdown confirmed a critical role for this histone demethylase in mediating PCB-induced inflammation of the vascular endothelium. Finally, it was determined, via chemical inhibition, that PCB-induced up-regulation of JMJD2B was estrogen receptor-alpha (ER-α) dependent. These data suggest that coplanar PCBs may exert endothelial cell toxicity through changes in histone modifications. - Highlights: • Coplanar PCBs significantly induced histone demethylase JMJD2B expression. • Coplanar PCBs activated NF-κB through p65 up-regulation and nuclear translocation. • Histone H3K4 and K9 modifications were mediated by ER-α/JMJD2B/MLL2 complex. �

Overexpression of p65 attenuates celecoxib-induced cell death in MDA-MB-231 human breast cancer cell line

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Wang Ling

2013-02-01

Full Text Available Abstract Background Celecoxib is a selective cyclooxygenase (COX-2 inhibitor that has been reported to reduce the risk of breast cancer. In our previous study, celecoxib induced apoptosis and caused cell cycle arrest at the G0/G1 phase in the breast cancer cell line MDA-MB-231, and its effects were mediated by downregulation of NF-κB signaling. The NF-κB p65/RelA subunit may play a role in cell death through the activation of anti-apoptotic target genes including the inhibitor of apoptosis (IAP and Bcl-2 families, and inhibition of protein kinase B/Akt. The aim of the present study was to investigate p65 as the potential target of celecoxib treatment and determine whether p65 overexpression can override the inhibitory effect of celecoxib on NF-κB activity and affect cell survival. Methods The effects of p65 overexpression on celecoxib-inhibited NF-κB transcriptional activity were examined by western blotting, electrophoretic mobility shift assay (EMSA and luciferase reporter gene assay. Cell viability and cell death were evaluated by the 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazoliumbromide (MTT assay, and the levels of cleaved poly(ADP-ribose polymerase (PARP and caspase. Anti-apoptotic NF-κB target genes and cell cycle regulators were examined by western blotting to screen for the expression of target genes under direct regulation by p65. Results Overexpression of p65 increased NF-κB transcriptional activity and interfered with celecoxib-mediated apoptosis as assessed by MTT assay and caspase-3, caspase-9, and PARP expressions. Exogenously overexpressed p65 upregulated NF-κB-responsive genes, including anti-apoptotic genes such as survivin and XIAP, and the cell cycle regulatory gene cyclin D1. However, p65 overexpression did not affect celecoxib-induced p-Akt inactivation, suggesting that celecoxib might have separate molecular mechanisms for regulating Akt signaling independently of its inhibition of NF-κB transcriptional

Characterization of tumour virus proteins. I. radioimmunoassay of the P27 protein of avian viruses

International Nuclear Information System (INIS)

Higuchi, T.

1977-01-01

The major structural protein of avian oncornaviruses, a core component of about 27000 daltons, has been measured by radioimmunoassay. The purified protein was labelled with 125 Iodine by chloramine-T method. The immune serum titer was defined as the highest serum dilution able to precipitate 50% of the labelled antigon present in the system. Standard competition curve was constructed in order to determine the equivalents of protein, in a system with limiting antibody concentration. In the experimental conditions used, 0.14 ng of AMV-P27 inhibited 50% of 125 I-AMV-P27 (1.0 ng) precipitation. The 125 I-AMV-P27 vs anti-AMV-P27 system was used to study the competition of normal cells, purified virus suspension, productive cells and supernatant fluids. Most of the chicken ombryo fibroblast showed expression of this viral component. The phenomena of cell transformation, the increase in total protein, and the expression of P27 were studied in rapid transformation of CEF by RSV-SR sub(A) [pt

Klotho ameliorates cyclosporine A-induced nephropathy via PDLIM2/NF-kB p65 signaling pathway.

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Jin, Meihua; Lv, Pengfei; Chen, Guanyu; Wang, Peng; Zuo, Zhongfu; Ren, Lili; Bi, Jing; Yang, Chul-Woo; Mei, Xifan; Han, Donghe

2017-04-29

Klotho, an antiaging protein, can extend the lifespan and modulate cellular responses to inflammation and oxidative stress which can ameliorate chronic kidney diseases (CKD). To investigate the molecular mechanism of Klotho on inflammation in cyclosporine A (CsA) induced nephropathy, the mice were transfected with adenovirus mediated Klotho gene and treated with cyclosporine A (CsA; 30 mg/kg/day) for 4 weeks. Also, primary human renal proximal tubule epithelial cells (RPTECs) were treated with soluble Klotho protein and LPS. The results showed that Ad-klotho significantly reduced serum creatinine (Scr) and blood urea nitrogen (BUN) caused by CsA, and significantly increased creatinine clearance. Tubule interstitial fibrosis score (TIF), renal 8-OHdG excretion, macrophage infiltration and MCP-1 were decreased after Ad-klotho gene transfer. In addition, the overexpression of Klotho led to increase in the expression of PDLIM2, decreased in the amount of NF-kB p65, and inhibited the production of inflammatory cytokines (TNFα, IL-6, IL-12) and iNOS. Accordingly, in vitro results showed, Klotho enhanced PDLIM2 expression and reduced NF-kB p65 expression, while PDLIM2 siRNA could block the inhibitory effects of Klotho on expression of NF-kB p65. Secretion of inflammatory cytokines was also inhibited by Klotho treatment, and PDLIM2 siRNA hindered regulatory effects of Klotho on the cytokines. Real-time PCR and Luciferase assay showed that Klotho markedly increased expression of PDLIM2 mRNA and PDLIM2 reporter activity in a dose-dependent manner. These findings suggest that Klotho can modulate inflammation via PDLIM2/NF-kB p65 pathway in CsA-induced nephropathy. Copyright © 2017 Elsevier Inc. All rights reserved.

Normal p50 gating in unmedicated schizophrenia outpatients

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Arnfred, Sidse M; Chen, Andrew C.N.; Glenthøj, Birte Y

2003-01-01

The hypothesis of a sensory gating defect in schizophrenia has been supported by studies demonstrating deficient auditory P50 gating in patients. P50 gating is the relative attenuation of P50 amplitude in the auditory evoked potential following the second auditory stimulus of a stimulus pair....

Dimethyl Fumarate Inhibits the Nuclear Factor κB Pathway in Breast Cancer Cells by Covalent Modification of p65 Protein.

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Kastrati, Irida; Siklos, Marton I; Calderon-Gierszal, Esther L; El-Shennawy, Lamiaa; Georgieva, Gergana; Thayer, Emily N; Thatcher, Gregory R J; Frasor, Jonna

2016-02-12

In breast tumors, activation of the nuclear factor κB (NFκB) pathway promotes survival, migration, invasion, angiogenesis, stem cell-like properties, and resistance to therapy--all phenotypes of aggressive disease where therapy options remain limited. Adding an anti-inflammatory/anti-NFκB agent to breast cancer treatment would be beneficial, but no such drug is approved as either a monotherapy or adjuvant therapy. To address this need, we examined whether dimethyl fumarate (DMF), an anti-inflammatory drug already in clinical use for multiple sclerosis, can inhibit the NFκB pathway. We found that DMF effectively blocks NFκB activity in multiple breast cancer cell lines and abrogates NFκB-dependent mammosphere formation, indicating that DMF has anti-cancer stem cell properties. In addition, DMF inhibits cell proliferation and significantly impairs xenograft tumor growth. Mechanistically, DMF prevents p65 nuclear translocation and attenuates its DNA binding activity but has no effect on upstream proteins in the NFκB pathway. Dimethyl succinate, the inactive analog of DMF that lacks the electrophilic double bond of fumarate, is unable to inhibit NFκB activity. Also, the cell-permeable thiol N-acetyl l-cysteine, reverses DMF inhibition of the NFκB pathway, supporting the notion that the electrophile, DMF, acts via covalent modification. To determine whether DMF interacts directly with p65, we synthesized and used a novel chemical probe of DMF by incorporating an alkyne functionality and found that DMF covalently modifies p65, with cysteine 38 being essential for the activity of DMF. These results establish DMF as an NFκB inhibitor with anti-tumor activity that may add therapeutic value in the treatment of aggressive breast cancers. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Analysis of recombinant proteins by isoelectric focusing in immobilized pH gradients

NARCIS (Netherlands)

Bischoff, Rainer; Roecklin, D.; Roitsch, C.

1992-01-01

Isoelectric focusing in immobilized pH gradients (IEF-IPG) was used to analyze three different recombinant proteins. Recombinant leech hirudin (65 amino acids, three disulfide bonds) expressed in Saccharomyces cerevisiae as a secreted protein and purified by anion-exchange and reversed-phase

Airborne spectrophotometry of P/Halley from 20 to 65 microns

Science.gov (United States)

Glaccum, W.; Moseley, S. H.; Campins, H.; Loewenstein, R. F.

1986-01-01

Simultaneous 20 to 65 microns spectrometry and 100 microns photometry of P/Halley obtained on board the Kuiper Airborne Observatory (KAO) in 1985 Dec. and 1986 April are discussed. Spectra with resolution 30 to 50 were obtained with the NASA/Goddard 24 channel grating spectrometer. Measurements were made on the nucleus as well as 5 points along and perpendicular to the Sun-tail direction. The observations reveal the absence of any strong spectral features. The color temperature of the dust varies over time scales as short as 2 days, but is higher than that expected for a rapidly rotating blackbody at the same distance from the Sun. The color temperature does not vary within 1 arcmin of the nucleus, but the coma is brighter on the sunward side than on the antisunward side.

PRMT5-Mediated Methylation of NF-κB p65 at Arg174 Is Required for Endothelial CXCL11 Gene Induction in Response to TNF-α and IFN-γ Costimulation.

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Daniel P Harris

Full Text Available Inflammatory agonists differentially activate gene expression of the chemokine family of proteins in endothelial cells (EC. TNF is a weak inducer of the chemokine CXCL11, while TNF and IFN-γ costimulation results in potent CXCL11 induction. The molecular mechanisms underlying TNF plus IFN-γ-mediated CXCL11 induction are not fully understood. We have previously reported that the protein arginine methyltransferase PRMT5 catalyzes symmetrical dimethylation of the NF-κB subunit p65 in EC at multiple arginine residues. Methylation of Arg30 and Arg35 on p65 is critical for TNF induction of CXCL10 in EC. Here we show that PRMT5-mediated methylation of p65 at Arg174 is required for induction of CXCL11 when EC are costimulated with TNF and IFN-γ. Knockdown of PRMT5 by RNAi reduced CXCL11 mRNA and protein levels in costimulated cells. Reconstitution of p65 Arg174Ala or Arg174Lys mutants into EC that were depleted of endogenous p65 blunted TNF plus IFN-γ-mediated CXCL11 induction. Mass spectrometric analyses showed that p65 Arg174 arginine methylation is enhanced by TNF plus IFN-γ costimulation, and is catalyzed by PRMT5. Chromatin immunoprecipitation assays (ChIP demonstrated that PRMT5 is necessary for p65 association with the CXCL11 promoter in response to TNF plus IFN-γ. Further, reconstitution of p65 Arg174Lys mutant in EC abrogated this p65 association with the CXCL11 promoter. Finally, ChIP and Re-ChIP assays revealed that symmetrical dimethylarginine-containing proteins complexed with the CXCL11 promoter were diminished in p65 Arg174Lys-reconstituted EC stimulated with TNF and IFN-γ. In total, these results indicate that PRMT5-mediated p65 methylation at Arg174 is essential for TNF plus IFN-γ-mediated CXCL11 gene induction. We therefore suggest that the use of recently developed small molecule inhibitors of PRMT5 may present a therapeutic approach to moderating chronic inflammatory pathologies.

Expression of 65-kDa oncofetal protein in experimental hepatoma after antivancer therapy

International Nuclear Information System (INIS)

Mirowski, M.; Rozalski, M.; Krajewska, U.; Wierbicki, R.; Hanausek, M.

1997-01-01

We have tested the expression of 65-kDa oncofetal protein (p65) after combined treatment with menadione and methotrexate in hamsters transplanted with Kirkman-Robbins hepatoma. The treatment of tumor-bearing animals with these compounds significantly inhibited both the tumor development and the expression of p65. This inhibition in tumor tissue was calculated from densitograms of Western blots. The inhibition of p65 was also confirmed in the serum of hepatoma bearing animals by using solid-phase radioimmunoassay (RIA) to quantify the specificity of polyclonal antibodies to fetal p65 molecules. Additionally, p65 was shown to localize both in cytoplasm an in the nuclear extracts prepared from hepatoma tissue. (author)

Synergistic activation of the CMV promoter by NF-κB P50 and PKG

International Nuclear Information System (INIS)

He Bin; Weber, Georg F.

2004-01-01

Several DNA binding NF-κB subunits are substrates for cGMP-dependent kinase (PKG) and their transactivation from cognate sites is induced by phosphorylation. This includes p50, which does not have a transcriptional activation domain and therefore needs to bind to other proteins to mediate gene expression. Here, we describe the synergistic transactivation by p50 and PKG from the CMV promoter. This is caused not only by phosphorylation of p50, leading to increased DNA binding, but also by PKG-dependent activation of CRE sites in the promoter. One of the CRE sites is located directly adjacent to a NF-κB site and is essential for p50-mediated induction of transcription. According to the binding of CREB to p50 in pull-down assays and according to the inhibition of p50-dependent transactivation by dominant-negative CREB, this reflects the formation of a transcription factor complex containing CREB and p50. The nuclear translocation of NF-κB is insufficient to distinguish among the multitude of promoters that harbor cognate recognition sites. The phosphorylation of multiple transcription factors by an upstream kinase, such as PKG, can lead to the formation of transcription factor complexes and differential transactivation from a subset of NF-κB sites. These interactions may be relevant for the activation of viral gene expression

MicroRNAs control transcription factor NF-kB (p65) expression in human ovarian cells.

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Sirotkin, Alexander V; Alexa, Richard; Kišová, Gabriela; Harrath, Abdel Halim; Alwasel, Saleh; Ovcharenko, Dmitriy; Mlynček, Miloš

2015-05-01

MicroRNAs (miRNAs) are known to influence ovarian cell proliferation, apoptosis and hormone release, but it remains unknown whether miRNAs affect ovarian functions via transcription factors. We examined the effect of miRNAs on nuclear factor-κappaB (NF-kB) (p65) expression in human ovarian luteinized granulosa cells. We transfected cultured primary human ovarian luteinized granulosa cells with 80 different constructs encoding human pre-miRNAs and then evaluated NF-kB (p65) expression (percentage of cells containing p65) by immunocytochemistry. We found that 21 of the constructs stimulated NF-kB (p65) expression and 18 of the constructs inhibited NF-kB (p65) expression. This is the first direct demonstration that miRNAs affect NF-kB (p65) expression and the first genome-scale miRNA screen to identify upregulation and downregulation of NF-kB accumulation by miRNAs in the ovary. Novel miRNAs that affect the NF-kB signalling pathway could be useful for the control of NF-kB-dependent reproductive processes and the treatment of NF-kB-dependent reproductive disorders.

Degradation kinetics of fisetin and quercetin in solutions affected by medium pH, temperature and co-existed proteins

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Wang Jing

2016-01-01

Full Text Available Impacts of medium pH, temperature and coexisted proteins on the degradation of two flavonoids fisetin and quercetin were assessed by spectroscopic method in the present study. Based on the measured degradation rate constants (k, fisetin was more stable than quercetin in all cases. Increasing medium pH from 6.0 to 7.5 at 37°C enhanced respective k values of fisetin and quercetin from 8.30x10−3 and 2.81x10−2 to 0.202 and 0.375 h-1 (P<0.05. In comparison with their degradation at 37°C, fisetin and quercetin showed larger k values at higher temperature (0.124 and 0.245 h−1 at 50°C, or 0.490 and 1.42 h−1 at 65°C. Four protein products in medium could stabilize the two flavonoids (P<0.05, as these proteins at 0.10 g L-1 decreased respective k values of fisetin and quercetin to 2.28x10−2-2.98x10−2 and 4.37´10−2-5.97x10−2 h−1. Hydrophobic interaction between the proteins and the two flavonoids was evidenced responsible for the stabilization, as sodium dodecyl sulfate could destroy the stabilization significantly (P<0.05. Casein and soybean protein provided greater stabilization than whey protein isolate. It is thus concluded that higher temperature and alkaline pH can enhance flavonoid loss, whereas coexisted proteins as flavonoid stabilizers can inhibit flavonoid degradation.

Effect of Huperzine A on Aβ-induced p65 of astrocyte in vitro.

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Xie, Lushuang; Jiang, Cen; Wang, Zhang; Yi, Xiaohong; Gong, Yuanyuan; Chen, Yunhui; Fu, Yan

2016-12-01

Alzheimer's disease (AD) is the most common cause of dementia. Its pathology often accompanies inflammatory action, and astrocytes play important roles in such procedure. Rela(p65) is one of significant message factors in NF-κB pathway which has been reported high expression in astrocyte treated by Aβ. HupA, an alkaloid isolated from Chinese herb Huperzia serrata, has been widely used to treat AD and observations reflected that it improves memory and cognitive capacity of AD patients. To reveal its molecular mechanisms on p65, we cultured astrocytes, built Aβ-induced AD model, treated astrocytes with HupA at different concentrations, assayed cell viability with MTT, and detected p65 expression by immunohistochemistry and PCR. Our results revealed that treatment with 10 μM Aβ1-42 for 24 h induced a significant increase of NF-κB in astrocytes; HupA significantly down-regulated p65 expression induced by Aβ in astrocytes. This study infers that HupA can regulate NF-κB pathway to treat AD.

A high-yield co-expression system for the purification of an intact drs2p-cdc50p lipid flippase complex, critically dependent on and stabilized by phosphatidylinositol-4-phosphate

DEFF Research Database (Denmark)

Azouaoui, Hassina; Montigny, Cédric; Ash, Miriam-Rose

2014-01-01

, the Drs2p-Cdc50p complex. After recovery of yeast membranes expressing both proteins, efficient purification was achieved in a single step by affinity chromatography on streptavidin beads, yielding ∼1-2 mg purified Drs2p-Cdc50p complex per liter of culture. Importantly, the procedure enabled us to recover...... was critically dependent on the simultaneous presence of PI4P and PS. We also identified a prominent role for PI4P in stabilization of the Drs2p-Cdc50p complex towards temperature- or C12E8-induced irreversible inactivation. These results indicate that the Drs2p-Cdc50p complex remains functional after affinity...... purification and that PI4P as a cofactor tightly controls its stability and catalytic activity. This work offers appealing perspectives for detailed structural and functional characterization of the Drs2p-Cdc50p lipid transport mechanism....

Inhibition of transcription factor NF-kappaB signaling proteins IKKbeta and p65 through specific cysteine residues by epoxyquinone A monomer: correlation with its anti-cancer cell growth activity.

Science.gov (United States)

Liang, Mei-Chih; Bardhan, Sujata; Pace, Emily A; Rosman, Diana; Beutler, John A; Porco, John A; Gilmore, Thomas D

2006-02-28

Transcription factor NF-kappaB is constitutively active in many human chronic inflammatory diseases and cancers. Epoxyquinone A monomer (EqM), a synthetic derivative of the natural product epoxyquinol A, has previously been shown to be a potent inhibitor of tumor necrosis factor-alpha (TNF-alpha)-induced activation of NF-kappaB, but the mechanism by which EqM inhibits NF-kappaB activation was not known. In this report, we show that EqM blocks activation of NF-kappaB by inhibiting two molecular targets: IkappaB kinase IKKbeta and NF-kappaB subunit p65. EqM inhibits TNF-alpha-induced IkappaBalpha phosphorylation and degradation by targeting IKKbeta, and an alanine substitution for Cys179 in the activation loop of IKKbeta makes it resistant to EqM-mediated inhibition. EqM also directly inhibits DNA binding by p65, but not p50; moreover, replacement of Cys38 in p65 with Ser abolishes EqM-mediated inhibition of DNA binding. Pretreatment of cells with reducing agent dithiothreitol dose-dependently reduces EqM-mediated inhibition of NF-kappaB, further suggesting that EqM directly modifies the thiol group of Cys residues in protein targets. Modifications of the exocyclic alkene of EqM substantially reduce EqM's ability to inhibit NF-kappaB activation. In the human SUDHL-4 lymphoma cell line, EqM inhibits both proliferation and NF-kappaB DNA binding, and activates caspase-3 activity. EqM also effectively inhibits the growth of human leukemia, kidney, and colon cancer cell lines in the NCI's tumor cell panel. Among six colon cancer cell lines, those with low amounts of constitutive NF-kappaB DNA-binding activity are generally more sensitive to growth inhibition by EqM. Taken together, these results suggest that EqM inhibits growth and induces cell death in tumor cells through a mechanism that involves inhibition of NF-kappaB activity at multiple steps in the signaling pathway.

Bcl-2 protein expression is associated with p27 and p53 protein expressions and MIB-1 counts in breast cancer

International Nuclear Information System (INIS)

Tsutsui, Shinichi; Yasuda, Kazuhiro; Suzuki, Kosuke; Takeuchi, Hideya; Nishizaki, Takashi; Higashi, Hidefumi; Era, Shoichi

2006-01-01

Recent experimental studies have shown that Bcl-2, which has been established as a key player in the control of apoptosis, plays a role in regulating the cell cycle and proliferation. The aim of this study was to investigate the relationship between Bcl-2 and p27 protein expression, p53 protein expression and the proliferation activity as defined by the MIB-1 counts. The prognostic implication of Bcl-2 protein expression in relation to p27 and p53 protein expressions and MIB-1 counts for breast cancer was also evaluated. The immunohistochemical expression of Bcl-2 protein was evaluated in a series of 249 invasive ductal carcinomas of the breast, in which p27 and p53 protein expressions and MIB-1 counts had been determined previously. The Bcl-2 protein expression was found to be decreased in 105 (42%) cases. A decreased Bcl-2 protein expression was significantly correlated with a nuclear grade of III, a negative estrogen receptor, a decreased p27 protein expression, a positive p53 protein expression, positive MIB-1 counts and a positive HER2 protein expression. The incidence of a nuclear grade of III and positive MIB-1 counts increased as the number of abnormal findings of Bcl-2, p27 and p53 protein expressions increased. A univariate analysis indicated a decreased Bcl-2 protein expression to be significantly (p = 0.0089) associated with a worse disease free survival (DFS), while a multivariate analysis indicated the lymph node status and MIB-1 counts to be independently significant prognostic factors for the DFS. The Bcl-2 protein expression has a close correlation with p27 and p53 protein expressions and the proliferation activity determined by MIB-1 counts in invasive ductal carcinoma of the breast. The prognostic value of Bcl-2 as well as p27 and p53 protein expressions was dependent on the proliferation activity in breast cancer

Bcl-2 protein expression is associated with p27 and p53 protein expressions and MIB-1 counts in breast cancer

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Nishizaki Takashi

2006-07-01

Full Text Available Abstract Background Recent experimental studies have shown that Bcl-2, which has been established as a key player in the control of apoptosis, plays a role in regulating the cell cycle and proliferation. The aim of this study was to investigate the relationship between Bcl-2 and p27 protein expression, p53 protein expression and the proliferation activity as defined by the MIB-1 counts. The prognostic implication of Bcl-2 protein expression in relation to p27 and p53 protein expressions and MIB-1 counts for breast cancer was also evaluated. Methods The immunohistochemical expression of Bcl-2 protein was evaluated in a series of 249 invasive ductal carcinomas of the breast, in which p27 and p53 protein expressions and MIB-1 counts had been determined previously. Results The Bcl-2 protein expression was found to be decreased in 105 (42% cases. A decreased Bcl-2 protein expression was significantly correlated with a nuclear grade of III, a negative estrogen receptor, a decreased p27 protein expression, a positive p53 protein expression, positive MIB-1 counts and a positive HER2 protein expression. The incidence of a nuclear grade of III and positive MIB-1 counts increased as the number of abnormal findings of Bcl-2, p27 and p53 protein expressions increased. A univariate analysis indicated a decreased Bcl-2 protein expression to be significantly (p = 0.0089 associated with a worse disease free survival (DFS, while a multivariate analysis indicated the lymph node status and MIB-1 counts to be independently significant prognostic factors for the DFS. Conclusion The Bcl-2 protein expression has a close correlation with p27 and p53 protein expressions and the proliferation activity determined by MIB-1 counts in invasive ductal carcinoma of the breast. The prognostic value of Bcl-2 as well as p27 and p53 protein expressions was dependent on the proliferation activity in breast cancer.

Neonatal High Bone Mass With First Mutation of the NF-κB Complex: Heterozygous De Novo Missense (p.Asp512Ser) RELA (Rela/p65).

Science.gov (United States)

Frederiksen, Anja L; Larsen, Martin J; Brusgaard, Klaus; Novack, Deborah V; Knudsen, Peter Juel Thiis; Schrøder, Henrik Daa; Qiu, Weimin; Eckhardt, Christina; McAlister, William H; Kassem, Moustapha; Mumm, Steven; Frost, Morten; Whyte, Michael P

2016-01-01

Heritable disorders that feature high bone mass (HBM) are rare. The etiology is typically a mutation(s) within a gene that regulates the differentiation and function of osteoblasts (OBs) or osteoclasts (OCs). Nevertheless, the molecular basis is unknown for approximately one-fifth of such entities. NF-κB signaling is a key regulator of bone remodeling and acts by enhancing OC survival while impairing OB maturation and function. The NF-κB transcription complex comprises five subunits. In mice, deletion of the p50 and p52 subunits together causes osteopetrosis (OPT). In humans, however, mutations within the genes that encode the NF-κB complex, including the Rela/p65 subunit, have not been reported. We describe a neonate who died suddenly and unexpectedly and was found at postmortem to have HBM documented radiographically and by skeletal histopathology. Serum was not available for study. Radiographic changes resembled malignant OPT, but histopathological investigation showed morphologically normal OCs and evidence of intact bone resorption excluding OPT. Furthermore, mutation analysis was negative for eight genes associated with OPT or HBM. Instead, accelerated bone formation appeared to account for the HBM. Subsequently, trio-based whole exome sequencing revealed a heterozygous de novo missense mutation (c.1534_1535delinsAG, p.Asp512Ser) in exon 11 of RELA encoding Rela/p65. The mutation was then verified using bidirectional Sanger sequencing. Lipopolysaccharide stimulation of patient fibroblasts elicited impaired NF-κB responses compared with healthy control fibroblasts. Five unrelated patients with unexplained HBM did not show a RELA defect. Ours is apparently the first report of a mutation within the NF-κB complex in humans. The missense change is associated with neonatal osteosclerosis from in utero increased OB function rather than failed OC action. These findings demonstrate the importance of the Rela/p65 subunit within the NF-κB pathway for human

Increased p50/p50 NF-κB Activation in Human Papillomavirus Type 6- or Type 11-Induced Laryngeal Papilloma Tissue

Science.gov (United States)

Vancurova, Ivana; Wu, Rong; Miskolci, Veronika; Sun, Shishinn

2002-01-01

We have observed elevated NF-κB DNA-binding activity in nuclear extracts from human papillomavirus type 6- and 11-infected laryngeal papilloma tissues. The predominant DNA-binding species is the p50/p50 homodimer. The elevated NF-κB activity could be correlated with a reduced level of cytoplasmic IκBβ and could be associated with the overexpression of p21CIP1/WAF1 in papilloma cells. Increased NF-κB activity and cytoplasmic accumulation of p21CIP1/WAF1 might counteract death-promoting effects elicited by overexpressed PTEN and reduced activation of Akt and STAT3 previously noted in these tissues. PMID:11773428

Turnover-dependent inactivation of the nitrogenase MoFe-protein at high pH.

Science.gov (United States)

Yang, Kun-Yun; Haynes, Chad A; Spatzal, Thomas; Rees, Douglas C; Howard, James B

2014-01-21

Proton uptake accompanies the reduction of all known substrates by nitrogenase. As a consequence, a higher pH should limit the availability of protons as a substrate essential for turnover, thereby increasing the proportion of more highly reduced forms of the enzyme for further study. The utility of the high-pH approach would appear to be problematic in view of the observation reported by Pham and Burgess [(1993) Biochemistry 32, 13725-13731] that the MoFe-protein undergoes irreversible protein denaturation above pH 8.65. In contrast, we found by both enzyme activity and crystallographic analyses that the MoFe-protein is stable when incubated at pH 9.5. We did observe, however, that at higher pHs and under turnover conditions, the MoFe-protein is slowly inactivated. While a normal, albeit low, level of substrate reduction occurs under these conditions, the MoFe-protein undergoes a complex transformation; initially, the enzyme is reversibly inhibited for substrate reduction at pH 9.5, yet in a second, slower process, the MoFe-protein becomes irreversibly inactivated as measured by substrate reduction activity at the optimal pH of 7.8. The final inactivated MoFe-protein has an increased hydrodynamic radius compared to that of the native MoFe-protein, yet it has a full complement of iron and molybdenum. Significantly, the modified MoFe-protein retains the ability to specifically interact with its nitrogenase partner, the Fe-protein, as judged by the support of ATP hydrolysis and by formation of a tight complex with the Fe-protein in the presence of ATP and aluminum fluoride. The turnover-dependent inactivation coupled to conformational change suggests a mechanism-based transformation that may provide a new probe of nitrogenase catalysis.

Cross-talk between PKA-Cβ and p65 mediates synergistic induction of PDE4B by roflumilast and NTHi

Science.gov (United States)

Susuki-Miyata, Seiko; Miyata, Masanori; Lee, Byung-Cheol; Xu, Haidong; Kai, Hirofumi; Yan, Chen; Li, Jian-Dong

2015-01-01

Phosphodiesterase 4B (PDE4B) plays a key role in regulating inflammation. Roflumilast, a phosphodiesterase (PDE)4-selective inhibitor, has recently been approved for treating severe chronic obstructive pulmonary disease (COPD) patients with exacerbation. However, there is also clinical evidence suggesting the development of tachyphylaxis or tolerance on repeated dosing of roflumilast and the possible contribution of PDE4B up-regulation, which could be counterproductive for suppressing inflammation. Thus, understanding how PDE4B is up-regulated in the context of the complex pathogenesis and medications of COPD may help improve the efficacy and possibly ameliorate the tolerance of roflumilast. Here we show that roflumilast synergizes with nontypeable Haemophilus influenzae (NTHi), a major bacterial cause of COPD exacerbation, to up-regulate PDE4B2 expression in human airway epithelial cells in vitro and in vivo. Up-regulated PDE4B2 contributes to the induction of certain important chemokines in both enzymatic activity-dependent and activity-independent manners. We also found that protein kinase A catalytic subunit β (PKA-Cβ) and nuclear factor-κB (NF-κB) p65 subunit were required for the synergistic induction of PDE4B2. PKA-Cβ phosphorylates p65 in a cAMP-dependent manner. Moreover, Ser276 of p65 is critical for mediating the PKA-Cβ–induced p65 phosphorylation and the synergistic induction of PDE4B2. Collectively, our data unveil a previously unidentified mechanism underlying synergistic up-regulation of PDE4B2 via a cross-talk between PKA-Cβ and p65 and may help develop new therapeutic strategies to improve the efficacy of PDE4 inhibitor. PMID:25831493

African Heath Sciences Vol 7 No 4.p65

African Journals Online (AJOL)

FOMCS2

no evidence of significant associative interactions between Hib-CRM197 and MenC-CRM197 in saline-based buffers, pH 7.2. African ... aggregation of vaccine saccharide or protein components. 17, 18 ... chromatography and for analysis of the generated data. .... protein would be expected to elute after the main MenC-.

Piperlongumine selectively suppresses ABC-DLBCL through inhibition of NF-κB p65 subunit nuclear import

Energy Technology Data Exchange (ETDEWEB)

Niu, Mingshan [Blood Diseases Institute, Xuzhou Medical College, Xuzhou, Jiangsu (China); Jiangsu Key Laboratory of Bone Marrow Stem Cell, Xuzhou Medical College, Xuzhou, Jiangsu (China); Department of Hematology, Affiliated Hospital of Xuzhou Medical College, Xuzhou, Jiangsu (China); Shen, Yangling; Xu, Xiaoyu [Blood Diseases Institute, Xuzhou Medical College, Xuzhou, Jiangsu (China); Jiangsu Key Laboratory of Bone Marrow Stem Cell, Xuzhou Medical College, Xuzhou, Jiangsu (China); Yao, Yao; Fu, Chunling [Blood Diseases Institute, Xuzhou Medical College, Xuzhou, Jiangsu (China); Jiangsu Key Laboratory of Bone Marrow Stem Cell, Xuzhou Medical College, Xuzhou, Jiangsu (China); Department of Hematology, Affiliated Hospital of Xuzhou Medical College, Xuzhou, Jiangsu (China); Yan, Zhiling [Department of Hematology, Affiliated Hospital of Xuzhou Medical College, Xuzhou, Jiangsu (China); Wu, Qingyun [Blood Diseases Institute, Xuzhou Medical College, Xuzhou, Jiangsu (China); Jiangsu Key Laboratory of Bone Marrow Stem Cell, Xuzhou Medical College, Xuzhou, Jiangsu (China); Department of Hematology, Affiliated Hospital of Xuzhou Medical College, Xuzhou, Jiangsu (China); Cao, Jiang; Sang, Wei [Department of Hematology, Affiliated Hospital of Xuzhou Medical College, Xuzhou, Jiangsu (China); Zeng, Lingyu [Blood Diseases Institute, Xuzhou Medical College, Xuzhou, Jiangsu (China); Jiangsu Key Laboratory of Bone Marrow Stem Cell, Xuzhou Medical College, Xuzhou, Jiangsu (China); Department of Hematology, Affiliated Hospital of Xuzhou Medical College, Xuzhou, Jiangsu (China); Li, Zhenyu [Department of Hematology, Affiliated Hospital of Xuzhou Medical College, Xuzhou, Jiangsu (China); Liu, Xuejiao, E-mail: liuxuejiao0923@126.com [Insititute of Nervous System Diseases, Xuzhou Medical College, Xuzhou, Jiangsu (China); and others

2015-07-10

Constitutive NF-κB activation is required for survival of activated B cell-like subtype of diffuse large B cell lymphoma (ABC-DLBCL). However, current NF-κB targeting strategies lack cancer cell specificity. Here, we identified a novel inhibitor, piperlongumine, features direct binding to NF-κB p65 subunit and suppression of p65 nuclear import. This was accompanied by NF-κB reporter activity suppression and NF-κB target gene downregulation. Moreover, mutation of Cys{sup 38} to Ser in p65 abolished this effect of piperlongumine on inhibition of p65 nuclear import. Furthermore, we show that piperlongumine selectively inhibited proliferation and induced apoptosis of ABC-DLBCL cells. Most notably, it has been reported that piperlongumine did not affect normal cells even at high doses and was nontoxic to animals. Hence, our current study provides new insight into piperlongumine's mechanism of action and novel approach to ABC-DLBCL target therapy. - Highlights: • Current NF-κB targeting strategies lack cancer cell specificity. • Piperlongumine inhibits NF-κB p65 subunit nuclear import via directly binding to p65. • Piperlongumine selectively inhibits proliferation of ABC-DLBCL cells. • This study provides a novel approach to ABC-DLBCL target therapy.

Piperlongumine selectively suppresses ABC-DLBCL through inhibition of NF-κB p65 subunit nuclear import

International Nuclear Information System (INIS)

Niu, Mingshan; Shen, Yangling; Xu, Xiaoyu; Yao, Yao; Fu, Chunling; Yan, Zhiling; Wu, Qingyun; Cao, Jiang; Sang, Wei; Zeng, Lingyu; Li, Zhenyu; Liu, Xuejiao

2015-01-01

Constitutive NF-κB activation is required for survival of activated B cell-like subtype of diffuse large B cell lymphoma (ABC-DLBCL). However, current NF-κB targeting strategies lack cancer cell specificity. Here, we identified a novel inhibitor, piperlongumine, features direct binding to NF-κB p65 subunit and suppression of p65 nuclear import. This was accompanied by NF-κB reporter activity suppression and NF-κB target gene downregulation. Moreover, mutation of Cys 38 to Ser in p65 abolished this effect of piperlongumine on inhibition of p65 nuclear import. Furthermore, we show that piperlongumine selectively inhibited proliferation and induced apoptosis of ABC-DLBCL cells. Most notably, it has been reported that piperlongumine did not affect normal cells even at high doses and was nontoxic to animals. Hence, our current study provides new insight into piperlongumine's mechanism of action and novel approach to ABC-DLBCL target therapy. - Highlights: • Current NF-κB targeting strategies lack cancer cell specificity. • Piperlongumine inhibits NF-κB p65 subunit nuclear import via directly binding to p65. • Piperlongumine selectively inhibits proliferation of ABC-DLBCL cells. • This study provides a novel approach to ABC-DLBCL target therapy

Magnetocaloric effects in Mn1.35Fe0.65P1−xSix compounds

International Nuclear Information System (INIS)

Geng Yao-Xiang; Tegus O; Bi Li-Ge

2012-01-01

The structural and magnetocaloric properties of Mn 1.35 Fe 0.65 P 1−x Si x compounds are investigated. The Si-substituted compounds, Mn 1.35 Fe 0.65 P 1−x Si x with x = 0.52, 0.54, 0.55, 0.56, and 0.57, are prepared by high-energy ball milling and the solid-state reaction. The X-ray diffraction shows that the compounds crystallize into the Fe 2 P-type hexagonal structure with space group P6-bar2m. The magnetic measurements show that the Curie temperature of the compound increases from 253 K for x = 0.52 to 296 K for x = 0.56. The isothermal magnetic-entropy change of the Mn 1.35 Fe 0.65 P 1−x Si x compound decreases with the Si content increasing. The maximal value of the magnetic-entropy change is about 7.0 J/kg·K in the Mn 1.35 Fe 0.65 P 0.48 Si 0.52 compound with a field change of 1.5 T. The compound quenched in water possesses a larger magnetic entropy change and a smaller thermal hysteresis than the non-quenched samples. The thermal hysteresis of the compound is less than 3.5 K. The maximum adiabatic temperature change is about 1.4 K in the Mn 1.35 Fe 0.65 P 0.45 Si 0.55 compound with a field change of 1.48 T. (condensed matter: electronic structure, electrical, magnetic, and optical properties)

The subcellular distribution of the human ribosomal "stalk" components: P1, P2 and P0 proteins

DEFF Research Database (Denmark)

Tchórzewski, Marek; Krokowski, Dawid; Rzeski, Wojciech

2003-01-01

The ribosomal "stalk" structure is a distinct lateral protuberance located on the large ribosomal subunit in prokaryotic, as well as in eukaryotic cells. In eukaryotes, this ribosomal structure is composed of the acidic ribosomal P proteins, forming two hetero-dimers (P1/P2) attached...

Insulin receptor substrate-3, interacting with Bcl-3, enhances p50 NF-{kappa}B activity

Energy Technology Data Exchange (ETDEWEB)

Kabuta, Tomohiro [Departments of Animal Sciences and Applied Biological Chemistry, Graduate School of Agriculture and Life Sciences, The University of Tokyo, Tokyo 113-8657 (Japan); Department of Degenerative Neurological Diseases, National Institute of Neuroscience, National Center of Neurology and Psychiatry, Kodaira, Tokyo 187-8502 (Japan); Hakuno, Fumihiko; Cho, Yoshitake; Yamanaka, Daisuke; Chida, Kazuhiro [Departments of Animal Sciences and Applied Biological Chemistry, Graduate School of Agriculture and Life Sciences, The University of Tokyo, Tokyo 113-8657 (Japan); Asano, Tomoichiro [Graduate School of Biomedical Science, Hiroshima University, Hiroshima 734-8551 (Japan); Wada, Keiji [Department of Degenerative Neurological Diseases, National Institute of Neuroscience, National Center of Neurology and Psychiatry, Kodaira, Tokyo 187-8502 (Japan); Takahashi, Shin-Ichiro, E-mail: atkshin@mail.ecc.u-tokyo.ac.jp [Departments of Animal Sciences and Applied Biological Chemistry, Graduate School of Agriculture and Life Sciences, The University of Tokyo, Tokyo 113-8657 (Japan)

2010-04-09

The insulin receptor substrate (IRS) proteins are major substrates of both insulin receptor and insulin-like growth factor (IGF)-I receptor tyrosine kinases. Previously, we reported that IRS-3 is localized to both cytosol and nucleus, and possesses transcriptional activity. In the present study, we identified Bcl-3 as a novel binding protein to IRS-3. Bcl-3 is a nuclear protein, which forms a complex with the homodimer of p50 NF-{kappa}B, leading to enhancement of transcription through p50 NF-{kappa}B. We found that Bcl-3 interacts with the pleckstrin homology domain and the phosphotyrosine binding domain of IRS-3, and that IRS-3 interacts with the ankyrin repeat domain of Bcl-3. In addition, IRS-3 augmented the binding activity of p50 to the NF-{kappa}B DNA binding site, as well as the tumor necrosis factor (TNF)-{alpha}-induced transcriptional activity of NF-{kappa}B. Lastly, IRS-3 enhanced NF-{kappa}B-dependent anti-apoptotic gene induction and consequently inhibited TNF-{alpha}-induced cell death. This series of results proposes a novel function for IRS-3 as a transcriptional regulator in TNF-{alpha} signaling, distinct from its function as a substrate of insulin/IGF receptor kinases.

P50/P90 Analysis for Solar Energy Systems Using the System Advisor Model: Preprint

Energy Technology Data Exchange (ETDEWEB)

Dobos, A. P.; Gilman, P.; Kasberg, M.

2012-06-01

To secure competitive financing for a solar energy generation project, the economic risk associated with interannual solar resource variability must be quantified. One way to quantify this risk is to calculate exceedance probabilities representing the amount of energy expected to be produced by a plant. Many years of solar radiation and metereological data are required to determine these values, often called P50 or P90 values for the level of certainty they represent. This paper describes the two methods implemented in the National Renewable Energy Laboratory's System Advisor Model (SAM) to calculate P50 and P90 exceedance probabilities for solar energy projects. The methodology and supporting data sets are applicable to photovoltaic, solar water heating, and concentrating solar power (CSP) systems.

Suppressor of cytokine signaling 1 (SOCS1) limits NFkappaB signaling by decreasing p65 stability within the cell nucleus.

Science.gov (United States)

Strebovsky, Julia; Walker, Patrick; Lang, Roland; Dalpke, Alexander H

2011-03-01

Suppressor of cytokine signaling (SOCS) proteins are inhibitors of cytoplasmic Janus kinases (Jak) and signal transducer and activator of transcription (STAT) signaling pathways. Previously the authors surprisingly observed that SOCS1 translocated into the nucleus, which was because of the presence of a nuclear localization sequence. This report now hypothesizes that SOCS1 mediates specific functions within the nuclear compartment because it is instantly transported into the nucleus, as shown by photoactivation and live cell imaging in human HEK293 cells. The NFκB component p65 is identified as an interaction partner for SOCS1 but not for other members of the SOCS family. SOCS1 bound to p65 only within the nucleus. By means of its SOCS box domain, SOCS1 operated as a ubiquitin ligase, leading to polyubiquitination and proteasomal degradation of nuclear p65. Thus, SOCS1 limited prolonged p65 signaling and terminated expression of NFκB inducible genes. Using mutants that lack either nuclear translocation or a functional SOCS box, this report identifies genes that are regulated in a manner dependent on the nuclear availability of SOCS1. Data show that beyond its receptor-proximal function in Jak/STAT signaling, SOCS1 also regulates the duration of NFκB signaling within the cell nucleus, thus exerting a heretofore unrecognized function.

Immunochemical characterization of temperature-regulated production of enterocin L50 (EntL50A and EntL50B), enterocin P, and enterocin Q by Enterococcus faecium L50.

Science.gov (United States)

Criado, Raquel; Gutiérrez, Jorge; Martín, María; Herranz, Carmen; Hernández, Pablo E; Cintas, Luis M

2006-12-01

Polyclonal antibodies with specificity for enterocin L50A (EntL50A), enterocin L50B (EntL50B), and enterocin Q (EntQ) produced by Enterococcus faecium L50 have been generated by immunization of rabbits with chemically synthesized peptides derived from the C terminus of EntL50A (LR1) and EntL50B (LR2) and from the complete enterocin Q (EntQ) conjugated to the carrier protein keyhole limpet hemocyanin (KLH). The sensitivity and specificity of these antibodies were evaluated by a noncompetitive indirect enzyme-linked immunosorbent assay (NCI-ELISA) and a competitive indirect ELISA (CI-ELISA). The NCI-ELISA was valuable for detecting anti-EntL50A-, anti-EntL50B-, and anti-EntQ-specific antibodies in the sera of the LR1-KLH-, LR2-KLH-, and EntQ-KLH-immunized animals, respectively. Moreover, these antibodies and those specific for enterocin P (EntP) obtained in a previous work (J. Gutiérrez, R. Criado, R. Citti, M. Martín, C. Herranz, M. F. Fernández, L. M. Cintas, and P. E. Hernández, J. Agric. Food Chem. 52:2247-2255, 2004) were used in an NCI-ELISA to detect and quantify the production of EntL50A, EntL50B, EntP, and EntQ by the multiple-bacteriocin producer E. faecium L50 grown at different temperatures (16 to 47 degrees C). Our results show that temperature has a strong influence on bacteriocin production by this strain. EntL50A and EntL50B are synthesized at 16 to 32 degrees C, but production becomes negligible when the growth temperature is above 37 degrees C, whereas EntP and EntQ are synthesized at temperatures ranging from 16 to 47 degrees C. Maximum EntL50A and EntL50B production was detected at 25 degrees C, while EntP and EntQ are maximally produced at 37 and 47 degrees C, respectively. The loss of plasmid pCIZ1 (50 kb) and/or pCIZ2 (7.4 kb), encoding EntL50A and EntL50B as well as EntQ, respectively, resulted in a significant increase in production and stability of the chromosomally encoded EntP.

Acidic tumor microenvironment and pH-sensing G protein-coupled receptors.

Science.gov (United States)

Justus, Calvin R; Dong, Lixue; Yang, Li V

2013-12-05

The tumor microenvironment is acidic due to glycolytic cancer cell metabolism, hypoxia, and deficient blood perfusion. It is proposed that acidosis in the tumor microenvironment is an important stress factor and selection force for cancer cell somatic evolution. Acidic pH has pleiotropic effects on the proliferation, migration, invasion, metastasis, and therapeutic response of cancer cells and the function of immune cells, vascular cells, and other stromal cells. However, the molecular mechanisms by which cancer cells and stromal cells sense and respond to acidic pH in the tumor microenvironment are poorly understood. In this article the role of a family of pH-sensing G protein-coupled receptors (GPCRs) in tumor biology is reviewed. Recent studies show that the pH-sensing GPCRs, including GPR4, GPR65 (TDAG8), GPR68 (OGR1), and GPR132 (G2A), regulate cancer cell metastasis and proliferation, immune cell function, inflammation, and blood vessel formation. Activation of the proton-sensing GPCRs by acidosis transduces multiple downstream G protein signaling pathways. Since GPCRs are major drug targets, small molecule modulators of the pH-sensing GPCRs are being actively developed and evaluated. Research on the pH-sensing GPCRs will continue to provide important insights into the molecular interaction between tumor and its acidic microenvironment and may identify new targets for cancer therapy and chemoprevention.

Prion protein (PrP) gene-knockout cell lines: insight into functions of the PrP

OpenAIRE

Sakudo, Akikazu; Onodera, Takashi

2015-01-01

Elucidation of prion protein (PrP) functions is crucial to fully understand prion diseases. A major approach to studying PrP functions is the use of PrP gene-knockout (Prnp ?/?) mice. So far, six types of Prnp ?/? mice have been generated, demonstrating the promiscuous functions of PrP. Recently, other PrP family members, such as Doppel and Shadoo, have been found. However, information obtained from comparative studies of structural and functional analyses of these PrP family proteins do not ...

SIMPL enhancement of tumor necrosis factor-α dependent p65-MED1 complex formation is required for mammalian hematopoietic stem and progenitor cell function.

Directory of Open Access Journals (Sweden)

Weina Zhao

Full Text Available Significant insight into the signaling pathways leading to activation of the Rel transcription factor family, collectively termed NF-κB, has been gained. Less well understood is how subsets of NF-κB-dependent genes are regulated in a signal specific manner. The SIMPL protein (signaling molecule that interacts with mouse pelle-like kinase is required for full Tumor Necrosis Factor-α (TNFα induced NF-κB activity. We show that SIMPL is required for steady-state hematopoiesis and the expression of a subset of TNFα induced genes whose products regulate hematopoietic cell activity. To gain insight into the mechanism through which SIMPL modulates gene expression we focused on the Tnf gene, an immune response regulator required for steady-state hematopoiesis. In response to TNFα SIMPL localizes to the Tnf gene promoter where it modulates the initiation of Tnf gene transcription. SIMPL binding partners identified by mass spectrometry include proteins involved in transcription and the interaction between SIMPL and MED1 was characterized in more detail. In response to TNFα, SIMPL is found in p65-MED1 complexes where SIMPL enhances p65/MED1/SIMPL complex formation. Together our results indicate that SIMPL functions as a TNFα-dependent p65 co-activator by facilitating the recruitment of MED1 to p65 containing transcriptional complexes to control the expression of a subset of TNFα-induced genes.

A high-yield co-expression system for the purification of an intact drs2p-cdc50p lipid flippase complex, critically dependent on and stabilized by phosphatidylinositol-4-phosphate

DEFF Research Database (Denmark)

Azouaoui, Hassina; Montigny, Cédric; Ash, Miriam-Rose

2014-01-01

P-type ATPases from the P4 subfamily (P4-ATPases) are energy-dependent transporters, which are thought to establish lipid asymmetry in eukaryotic cell membranes. Together with their Cdc50 accessory subunits, P4-ATPases couple ATP hydrolysis to lipid transport from the exoplasmic to the cytoplasmic...... leaflet of plasma membranes, late Golgi membranes, and endosomes. To gain insights into the structure and function of these important membrane pumps, robust protocols for expression and purification are required. In this report, we present a procedure for high-yield co-expression of a yeast flippase......, the Drs2p-Cdc50p complex. After recovery of yeast membranes expressing both proteins, efficient purification was achieved in a single step by affinity chromatography on streptavidin beads, yielding ∼1-2 mg purified Drs2p-Cdc50p complex per liter of culture. Importantly, the procedure enabled us to recover...

NF-κB p50 subunit knockout impairs late LTP and alters long term memory in the mouse hippocampus

Directory of Open Access Journals (Sweden)

Oikawa Kensuke

2012-07-01

Full Text Available Abstract Background Nuclear factor kappa B (NF-κB is a transcription factor typically expressed with two specific subunits (p50, p65. Investigators have reported that NF-κB is activated during the induction of in vitro long term potentiation (LTP, a paradigm of synaptic plasticity and correlate of memory, suggesting that NF-κB may be necessary for some aspects of memory encoding. Furthermore, NF-κB has been implicated as a potential requirement in behavioral tests of memory. Unfortunately, very little work has been done to explore the effects of deleting specific NF-κB subunits on memory. Studies have shown that NF-κB p50 subunit deletion (p50−/− leads to memory deficits, however some recent studies suggest the contrary where p50−/− mice show enhanced memory in the Morris water maze (MWM. To more critically explore the role of the NF-κB p50 subunit in synaptic plasticity and memory, we assessed long term spatial memory in vivo using the MWM, and synaptic plasticity in vitro utilizing high frequency stimuli capable of eliciting LTP in slices from the hippocampus of NF-κB p50−/− versus their controls (p50+/+. Results We found that the lack of the NF-κB p50 subunit led to significant decreases in late LTP and in selective but significant alterations in MWM tests (i.e., some improvements during acquisition, but deficits during retention. Conclusions These results support the hypothesis that the NF-κ p50 subunit is required in long term spatial memory in the hippocampus.

Oral administration of Lactococcus lactis-expressing heat shock protein 65 and tandemly repeated IA2P2 prevents type 1 diabetes in NOD mice.

Science.gov (United States)

Liu, Kun-Feng; Liu, Xiao-Rui; Li, Guo-Liang; Lu, Shi-Ping; Jin, Liang; Wu, Jie

2016-06-01

Type 1 diabetes mellitus (T1DM) is a chronic autoimmune disease characterized by the destruction of insulin-secreting β cells upon autoreactive T cell attack. Oral administration of autoantigens is an attractive approach to treating T1DM, but an effective carrier should be used in order to protect antigens. Lactococcus lactis, a safe engineering strain, was used for this task in the present study. Two recombinant L. lactis expressing protein HSP65-6IA2P2 were used and be investigated the effects and mechanisms against T1DM in NOD mice. Our findings demonstrate that recombinant L. lactis strains can successfully both deliver antigens to intestinal mucosa and maintain the epitopes for a long time in NOD mice. Oral administration of recombinant L. lactis could prevent hyperglycemia, improve glucose tolerance, and reduce insulitis by inhibiting antigen-specific proliferation of T cells, augmenting regulatory immune reactions, and balancing ratios of Th17/Tregs and Th1/Th2. These results prove that orally administrated L. lactis expressing HSP65-6IA2P2 is an effective approach for the prevention of T1DM in NOD mice. Copyright © 2016 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.

High CpG island methylation of p16 gene and loss of p16 protein ...

Indian Academy of Sciences (India)

The study subjects consisted of 75 healthy ... that p16 protein expression was significantly lower in ToF group compared to ... in p16 promoters in ToF patients was negatively correlated with p16 protein ... studies, human foetal ventricular cardiomyocytes (HFCs) are ..... oral epithelial dysplasia: a prospective cohort study.

Protein expression of P13K and P53 in prediction of response to radiotherapy in cervical cancer

International Nuclear Information System (INIS)

Teja Kisnanto; Devita Tetriana; Iin Kurnia; Sudiono S; Mellova Amir; Budiningsih Siregar; Ramli; Andrijono; Setiawan Soetopo; Irwan; Tjahya Kurjana; Bethy S Hernowo; Maringan DL Tobing

2015-01-01

Cervical cancer is a malignant disease that is common in women and is the first order of malignant disease in Indonesia. Radiotherapy is the main treatment on cervical cancer, especially at an advanced stage (IIB-IIB). P13K and P-53 protein plays a role in the regulation of apoptosis (programmed cell death). The purpose of this study was to determine the protein expression of P13K and P-53 in the prediction of response to radiotherapy action in patients with cervical cancer. Microscopic preparations obtained from biopsy tissue cancer (IIB-IIIB) to 20 patients from RSCM and RSHS. The method used is the method of immunohistochemistry using P13K and P-53 protein biomarkers in cervical cancer tissue preparations. P13K protein expression value obtained by the method of immuno reactive Score (IRS). P13K protein positive expression marked in blue on the cell cytoplasm and P-53 protein is characterized by brown or dark colors contained in the cell nucleus. Results showed that IRS value by 10% a negative P13K, P13K IRS weaker by 70%, IRS P13K was at 15%, and the IRS P13K stronger by 5%. While the index positive P-53 was obtained by 75% and negative P-53 index by 5%. Radiotherapy response analysis showed that there were 75% good response and 25% a bad response. The conclusion from this study is the expression of the protein P13K and P-53 for response prediction of radiotherapy IRS P13K values obtained in response to both radiotherapy is higher compared with radiotherapy response is bad, and the P-53 protein is not found differences in response to radiotherapy between positive and negative expressions. (author)

Anti-mutated citrullinated vimentin (anti-MCV) and anti-65 kDa heat shock protein (anti-hsp65): new biomarkers in ankylosing spondylitis.

Science.gov (United States)

Bodnár, Nóra; Szekanecz, Zoltán; Prohászka, Zoltán; Kemény-Beke, Adám; Némethné-Gyurcsik, Zsuzsanna; Gulyás, Katalin; Lakos, Gabriella; Sipka, Sándor; Szántó, Sándor

2012-01-01

Citrullination as well as anti-citrullinated protein/peptide antibodies (ACPA) have been implicated in the pathogenesis of rheumatoid arthritis (RA). While ACPAs are specific and sensitive markers for RA, there have been hardly any reports regarding ACPAs in ankylosing spondylitis (AS). The possible role of antibodies to Mycobacterial 65 kDa heat shock protein (hsp65) has not been characterized in AS. As new laboratory biomarkers of AS are needed, we investigated the prevalence of anti-mutated citrullinated vimentin (MCV) and anti-hsp65 antibodies in AS. Altogether 43 AS and 44 healthy controls were included in the study. Anti-MCV and anti-hsp65 were determined in sera by commercial and in-house ELISA, respectively. Serum autoantibody levels were correlated with ESR, CRP, HLA-B27 status, smoking habits, pain intensity, BASDAI, BASFI and BASMI indices. Patients with AS had significantly higher serum anti-MCV levels (17.3 U/mL, range: 8.3-31.5 U/mL) in comparison to healthy subjects (8.9 U/mL, range: 5.4-13.3 U/mL) (p20 U/mL). The mean anti-hsp65 concentration in AS sera was 124.8 AU/mL (range: 27.2-1000 AU/mL), while controls exerted significantly lower anti-hsp65 levels (mean: 51.8 AU/mL; range: 22.5-88.5 AU/mL) (p<0.001). Correlation analysis revealed that both anti-MCV positivity (r=0.613; p=0.012) and absolute serum anti-MCV levels (r=0.553; p=0.021) correlated with anti-hsp65 levels. Anti-MCV positivity also correlated with ESR (r=0.437; p=0.03). Anti-MCV and anti-hsp65 may be novel biomarkers in AS. Copyright © 2011. Published by Elsevier SAS.

Two Crinivirus-specific proteins of Lettuce infectious yellows virus (LIYV), P26 and P9, are self-interacting.

Science.gov (United States)

Stewart, Lucy R; Hwang, Min Sook; Falk, Bryce W

2009-11-01

Interactions of Lettuce infectious yellows virus (LIYV)-encoded proteins were tested by yeast-two-hybrid (Y2H) assays. LIYV-encoded P34, Hsp70h, P59, CP, CPm, and P26 were tested in all possible pairwise combinations. Interaction was detected only for the P26-P26 combination. P26 self-interaction domains were mapped using a series of N- and C-terminal truncations. Orthologous P26 proteins from the criniviruses Beet pseudoyellows virus (BPYV), Cucurbit yellow stunting disorder virus (CYSDV), and Lettuce chlorosis virus (LCV) were also tested, and each exhibited strong self-interaction but no interaction with orthologous proteins. Two small putative proteins encoded by LIYV RNA2, P5 and P9, were also tested for interactions with the six aforementioned LIYV proteins and each other. No interactions were detected for P5, but P9-P9 self-interaction was detected. P26- and P9-encoding genes are present in all described members of the genus Crinivirus, but are not present in other members of the family Closteroviridae. LIYV P26 has previously been demonstrated to induce a unique LIYV cytopathology, plasmalemma deposits (PLDs), but no role is yet known for P9.

Functional characterization of a competitive peptide antagonist of p65 in human macrophage-like cells suggests therapeutic potential for chronic inflammation

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Srinivasan M

2014-12-01

Full Text Available Mythily Srinivasan,1 Corinne Blackburn,1 Debomoy K Lahiri2,3 1Department of Oral Pathology, Medicine and Radiology, Indiana University School of Dentistry, 2Institute of Psychiatry Research, Department of Psychiatry, 3Department of Medical and Molecular Genetics, School of Medicine, Indiana University-Purdue University, Indianapolis, IN, USA Abstract: Glucocorticoid-induced leucine zipper (GILZ is a glucocorticoid responsive protein that links the nuclear factor-kappa B (NFκB and the glucocorticoid signaling pathways. Functional and binding studies suggest that the proline-rich region at the carboxy terminus of GILZ binds the p65 subunit of NFκB and suppresses the immunoinflammatory response. A widely-used strategy in the discovery of peptide drugs involves exploitation of the complementary surfaces of naturally occurring binding partners. Previously, we observed that a synthetic peptide (GILZ-P derived from the proline-rich region of GILZ bound activated p65 and ameliorated experimental encephalomyelitis. Here we characterize the secondary structure of GILZ-P by circular dichroic analysis. GILZ-P adopts an extended polyproline type II helical conformation consistent with the structural conformation commonly observed in interfaces of transient intermolecular interactions. To determine the potential application of GILZ-P in humans, we evaluated the toxicity and efficacy of the peptide drug in mature human macrophage-like THP-1 cells. Treatment with GILZ-P at a wide range of concentrations commonly used for peptide drugs was nontoxic as determined by cell viability and apoptosis assays. Functionally, GILZ-P suppressed proliferation and glutamate secretion by activated macrophages by inhibiting nuclear translocation of p65. Collectively, our data suggest that the GILZ-P has therapeutic potential in chronic CNS diseases where persistent inflammation leads to neurodegeneration such as multiple sclerosis and Alzheimer’s disease. Keywords

Role of p73 Dinucleotide Polymorphism in Prostate Cancer and p73 Protein Isoform Balance

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L. Michael Carastro

2014-01-01

Full Text Available Background. Molecular markers for prostate cancer (PCa risks are currently lacking. Here we address the potential association of a dinucleotide polymorphism (DNP in exon 2 of the p73 gene with PCa risk/progression and discern any disruption of p73 protein isoforms levels in cells harboring a p73 DNP allele. Methods. We investigated the association between p73 DNP genotype and PCa risk/aggressiveness and survival by fitting logistic regression models in 1,292 incident cases and 682 controls. Results. Although we detected no association between p73 DNP and PCa risk, a significant inverse relationship between p73 DNP and PCa aggressiveness (AT/AT + GC/AT versus GC/GC, OR = 0.55, 95%Cl = 0.31–0.99 was detected. Also, p73 DNP is marginally associated with overall death (dominant model, HR = 0.76, 95%Cl = 0.57–1.00, P=0.053 as well as PCa specific death (HR = 0.69, 95%Cl = 0.45–1.06, P=0.09. Western blot analyses for p73 protein isoforms indicate that cells heterozygous for the p73 DNP have lower levels of ∆Np73 relative to TAp73 (P<0.001. Conclusions. Our findings are consistent with an association between p73 DNP and low risk for PCa aggressiveness by increasing the expressed TAp73/∆Np73 protein isoform ratio.

P50 suppression deficits and psychopathology in Han Chinese patients with schizophrenia.

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Zhu, Xiao Lin; Tan, Shu Ping; Wang, Zhi Ren; Zhang, Jin Guo; Li, Dong; Fan, Feng Mei; Zhao, Yan Li; Zou, Yi Zhuang; Tan, Yun Long; Yang, Fu De; Zhang, Xiang Yang

2017-07-13

Numerous studies have reported P50 gating deficits in schizophrenia, though with mixed results. Moreover, few studies have explored the association between P50 gating deficits and psychopathology in Chinese patients with schizophrenia. In the present study, we investigated the P50 auditory sensory gating patterns and their correlations with clinical symptoms in a large sample of Han Chinese patients with schizophrenia. We assessed P50 sensory gating with a 64-channel electroencephalography system in 133 patients with schizophrenia and 148 healthy controls. The schizophrenia symptomatology was assessed with the Positive and Negative Syndrome Scale (PANSS). Patients with schizophrenia had a significantly higher P50 gating ratio (pschizophrenia. These findings suggest that the P50 sensory gating deficits identified in Chinese patients with schizophrenia may not be involved in the psychopathology of the illness. Copyright © 2017 Elsevier B.V. All rights reserved.

Disparate effects of p24alpha and p24delta on secretory protein transport and processing.

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Jeroen R P M Strating

Full Text Available BACKGROUND: The p24 family is thought to be somehow involved in endoplasmic reticulum (ER-to-Golgi protein transport. A subset of the p24 proteins (p24alpha(3, -beta(1, -gamma(3 and -delta(2 is upregulated when Xenopus laevis intermediate pituitary melanotrope cells are physiologically activated to produce vast amounts of their major secretory cargo, the prohormone proopiomelanocortin (POMC. METHODOLOGY/PRINCIPAL FINDINGS: Here we find that transgene expression of p24alpha(3 or p24delta(2 specifically in the Xenopus melanotrope cells in both cases causes an effective displacement of the endogenous p24 proteins, resulting in severely distorted p24 systems and disparate melanotrope cell phenotypes. Transgene expression of p24alpha(3 greatly reduces POMC transport and leads to accumulation of the prohormone in large, ER-localized electron-dense structures, whereas p24delta(2-transgenesis does not influence the overall ultrastructure of the cells nor POMC transport and cleavage, but affects the Golgi-based processes of POMC glycomaturation and sulfation. CONCLUSIONS/SIGNIFICANCE: Transgenic expression of two distinct p24 family members has disparate effects on secretory pathway functioning, illustrating the specificity and non-redundancy of our transgenic approach. We conclude that members of the p24 family furnish subcompartments of the secretory pathway with specific sets of machinery cargo to provide the proper microenvironments for efficient and correct secretory protein transport and processing.

Punica granatum L. Fruit Aqueous Extract Suppresses Reactive Oxygen Species-Mediated p53/p65/miR-145 Expressions followed by Elevated Levels of irs-1 in Alloxan-Diabetic Rats.

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Gharib, Ehsan; Montasser Kouhsari, Shideh; Izad, Maryam

2018-01-01

Reactive oxygen species (ROS) is an apoptosis inducer in pancreatic β-cells that stimulates p53/p65 mediated microRNA (miR)-145 expression. Punica granatum L. (pomegranate) is an antioxidant fruit that attenuates ROS generation. This study examines the effects of pomegranate fruit aqueous extract (PGE) on the levels of ROS, p53, p65, miR-145, and its target insulin receptor substrate 1 (irs-1) mRNA in Alloxan-diabetic male Wistar rats. In this experimental study, diabetic rats received different doses of PGE. The effects of the PGE polyphenols were examined through a long-term PGE treatment period model, followed by an evaluation of the plasma and tissue contents of free fatty acids (FFAs), triglycerides (TG), and glycogen compared with diabetic controls (DC) and normal controls (NC). We used real-time polymerase chain reaction (PCR) to investigate the modulation of p53, p65, miR-145, and irs-1 expression levels. There was a noticeable reduction in fasting blood glucose (FBG) and ROS generation compared to DC. We observed marked decreases in p53, p65, miR-145 expression levels followed by an elevated level of irs-1, which contributed to improvement in insulin sensitivity. PGE administration downregulated miR-145 levels in Alloxan-diabetic Wistar rats by suppression of ROS-mediated p53 and p65 overexpression. Copyright© by Royan Institute. All rights reserved.

Calcium-dependent and -independent binding of the pentraxin serum amyloid P component to glycosaminoglycans and amyloid proteins

DEFF Research Database (Denmark)

Danielsen, B; Sørensen, I J; Nybo, Mads

1997-01-01

precursor protein beta2M was observed. This binding was also enhanced at slightly acid pH, most pronounced at pH 5.0. The results of this study indicate that SAP can exhibit both Ca2(+)-dependent and -independent binding to ligands involved in amyloid fibril formation and that the binding is enhanced under...... and beta2M) by ELISA. An increase in the dose-dependent binding of SAP to heparan sulfate, AA-protein and beta2M was observed as the pH decreased from 8.0 to 5.0. Furthermore, a lower, but significant Ca2(+)-independent binding of SAP to heparan sulfate, dermatan sulfate, AA protein and the amyloid...

Tuning the pH-shift protein-isolation method for maximum hemoglobin-removal from blood rich fish muscle.

Science.gov (United States)

Abdollahi, Mehdi; Marmon, Sofia; Chaijan, Manat; Undeland, Ingrid

2016-12-01

A main challenge preventing optimal use of protein isolated from unconventional raw materials (e.g., small pelagic fish and fish by-products) using the pH-shift method is the difficulty to remove enough heme-pigments. Here, the distribution of hemoglobin (Hb) in the different fractions formed during pH-shift processing was studied using Hb-fortified cod mince. Process modifications, additives and prewashing were then investigated to further facilitate Hb-removal. The alkaline pH-shift process version could remove considerably more Hb (77%) compared to the acidic version (37%) when proteins were precipitated at pH 5.5; most Hb was removed during dewatering. Protein precipitation at pH 6.5 improved total Hb removal up to 91% and 74% during alkaline and acid processing, respectively. Adding phytic acid to the first supernatant of the alkaline process version yielded 93% Hb removal. Combining one prewash with phytic acid at pH 5.5 followed by alkaline/acid pH-shift processing increased Hb removal up to 96/92%. Copyright © 2016 Elsevier Ltd. All rights reserved.

Crystallization and preliminary X-ray diffraction analysis of phospholipid-bound Sfh1p, a member of the Saccharomyces cerevisiae Sec14p-like phosphatidylinositol transfer protein family

International Nuclear Information System (INIS)

Schaaf, Gabriel; Betts, Laurie; Garrett, Teresa A.; Raetz, Christian R. H.; Bankaitis, Vytas A.

2006-01-01

Yeast Sfh1p, a close homolog of the Sec14p phosphatidylinositol transfer protein, was crystallized in the absence of detergent. X-ray data have been collected to 2.5 Å. Sec14p is the major phosphatidylinositol (PtdIns)/phosphatidylcholine (PtdCho) transfer protein in the budding yeast Saccharomyces cerevisiae and is the founding member of a large eukaryotic protein superfamily. This protein catalyzes the exchange of either PtdIns or PtdCho between membrane bilayers in vitro and this exchange reaction requires no external input of energy or of other protein cofactors. Despite the previous elucidation of the crystal structure of a detergent-bound form of Sec14p, the conformational changes that accompany the phospholipid-exchange reaction remain undefined. Moreover, a structural appreciation of how Sec14p or its homologs bind their various phospholipid substrates remains elusive. Here, the purification and crystallization of yeast Sfh1p, the protein most closely related to Sec14p, are reported. A combination of electrospray ionization mass-spectrometry and collision-induced decomposition mass-spectrometry methods indicate that recombinant Sfh1p loads predominantly with phosphatidylethanolamine. Unlike phospholipid-bound forms of Sec14p, this form of Sfh1p crystallizes readily in the absence of detergent. Sfh1p crystals diffract to 2.5 Å and belong to the orthorhombic primitive space group P2 1 2 1 2 1 , with unit-cell parameters a = 49.40, b = 71.55, c = 98.21 Å, α = β = γ = 90°. One Sfh1p molecule is present in the asymmetric unit (V M = 2.5 Å 3 Da −1 ; V s = 50%). Crystallization of a phospholipid-bound Sec14p-like protein is a critical first step in obtaining the first high-resolution picture of how proteins of the Sec14p superfamily bind their phospholipid ligands. This information will significantly extend our current understanding of how Sec14p-like proteins catalyze phospholipid exchange

Nuclear IL-33 is a transcriptional regulator of NF-{kappa}B p65 and induces endothelial cell activation

Energy Technology Data Exchange (ETDEWEB)

Choi, Yeon-Sook; Park, Jeong Ae; Kim, Jihye; Rho, Seung-Sik; Park, Hyojin [Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749 (Korea, Republic of); Kim, Young-Myeong [Department of Molecular and Cellular Biochemistry, School of Medicine, Kangwon National University, Chuncheon (Korea, Republic of); Kwon, Young-Guen, E-mail: ygkwon@yonsei.ac.kr [Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749 (Korea, Republic of)

2012-05-04

Highlights: Black-Right-Pointing-Pointer IL-33 as nuclear factor regulated expression of ICAM-1 and VCAM-1. Black-Right-Pointing-Pointer Nuclear IL-33 increased the transcription of NF-{kappa}B p65 by binding to the p65 promoter. Black-Right-Pointing-Pointer Nuclear IL-33 controls NF-{kappa}B-dependent inflammatory responses. -- Abstract: Interleukin (IL)-33, an IL-1 family member, acts as an extracellular cytokine by binding its cognate receptor, ST2. IL-33 is also a chromatin-binding transcriptional regulator highly expressed in the nuclei of endothelial cells. However, the function of IL-33 as a nuclear factor is poorly defined. Here, we show that IL-33 is a novel transcriptional regulator of the p65 subunit of the NF-{kappa}B complex and is involved in endothelial cell activation. Quantitative reverse transcriptase PCR and Western blot analyses indicated that IL-33 mediates the expression of intercellular adhesion molecule (ICAM)-1 and vascular cell adhesion molecule (VCAM)-1 in endothelial cells basally and in response to tumor necrosis factor-{alpha}-treatment. IL-33-induced ICAM-1/VCAM-1 expression was dependent on the regulatory effect of IL-33 on the nuclear factor (NF)-{kappa}B pathway; NF-{kappa}B p65 expression was enhanced by IL-33 overexpression and, conversely, reduced by IL-33 knockdown. Moreover, NF-{kappa}B p65 promoter activity and chromatin immunoprecipitation analysis revealed that IL-33 binds to the p65 promoter region in the nucleus. Our data provide the first evidence that IL-33 in the nucleus of endothelial cells participates in inflammatory reactions as a transcriptional regulator of NF-{kappa}B p65.

Prion protein (PrP) gene-knockout cell lines: insight into functions of the PrP

Science.gov (United States)

Sakudo, Akikazu; Onodera, Takashi

2015-01-01

Elucidation of prion protein (PrP) functions is crucial to fully understand prion diseases. A major approach to studying PrP functions is the use of PrP gene-knockout (Prnp−/−) mice. So far, six types of Prnp−/− mice have been generated, demonstrating the promiscuous functions of PrP. Recently, other PrP family members, such as Doppel and Shadoo, have been found. However, information obtained from comparative studies of structural and functional analyses of these PrP family proteins do not fully reveal PrP functions. Recently, varieties of Prnp−/− cell lines established from Prnp−/− mice have contributed to the analysis of PrP functions. In this mini-review, we focus on Prnp−/− cell lines and summarize currently available Prnp−/− cell lines and their characterizations. In addition, we introduce the recent advances in the methodology of cell line generation with knockout or knockdown of the PrP gene. We also discuss how these cell lines have provided valuable insights into PrP functions and show future perspectives. PMID:25642423

Immunohistochemical analysis of P53 protein in odontogenic cysts

Science.gov (United States)

Gaballah, Essam Taher M.A.; Tawfik, Mohamed A.

2010-01-01

The p53 is a well-known tumor suppressor gene, the mutations of which are closely related to the decreased differentiation of cells. Findings of studies on immunohistochemical P53 expression in odontogenic cysts are controversial. The present study was carried-out to investigate the immunohistochemical expression of P53 protein in odontogenic cysts. Thirty paraffin blocks of diagnosed odontogenic cysts were processed to determine the immunohistochemical expression of P53 protein. Nine of the 11 odontogenic keratocysts (81.8%) expressed P53, one of three dentigerous cyst cases expressed P53, while none of the 16 radicular cysts expressed P53 protein. The findings of the present work supported the reclassification of OKC as keratocystic odontogenic tumor. PMID:23960493

Plasmodium berghei NK65 in Combination with IFN-γ Induces Endothelial Glucocorticoid Resistance via Sustained Activation of p38 and JNK

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Zielińska, Karolina A.; de Cauwer, Lode; Knoops, Sofie; Van der Molen, Kristof; Sneyers, Alexander; Thommis, Jonathan; De Souza, J. Brian; Opdenakker, Ghislain; De Bosscher, Karolien; Van den Steen, Philippe E.

2017-01-01

Malaria-associated acute respiratory distress syndrome (MA-ARDS) is an often lethal complication of malaria. Currently, no adequate therapy for this syndrome exists. Although glucocorticoids (GCs) have been used to improve clinical outcome of ARDS, their therapeutic benefits remain unclear. We previously developed a mouse model of MA-ARDS, in which dexamethasone treatment revealed GC resistance. In the present study, we investigated GC sensitivity of mouse microvascular lung endothelial cells stimulated with interferon-γ (IFN-γ) and Plasmodium berghei NK65 (PbNK65). Upon challenge with IFN-γ alone, dexamethasone inhibited the expression of CCL5 (RANTES) by 90% and both CCL2 (MCP-1) and CXCL10 (IP-10) by 50%. Accordingly, whole transcriptome analysis revealed that dexamethasone differentially affected several gene clusters and in particular inhibited a large cluster of IFN-γ-induced genes, including chemokines. In contrast, combined stimulation with IFN-γ and PbNK65 extract impaired inhibitory actions of GCs on chemokine release, without affecting the capacity of the GC receptor to accumulate in the nucleus. Subsequently, we investigated the effects of GCs on two signaling pathways activated by IFN-γ. Dexamethasone left phosphorylation and protein levels of signal transducer and activator of transcription 1 (STAT1) unhampered. In contrast, dexamethasone inhibited the IFN-γ-induced activation of two mitogen-activated protein kinases (MAPK), JNK, and p38. However, PbNK65 extract abolished the inhibitory effects of GCs on MAPK signaling, inducing GC resistance. These data provide novel insights into the mechanisms of GC actions in endothelial cells and show how malaria may impair the beneficial effects of GCs. PMID:29033931

Plasmodium berghei NK65 in Combination with IFN-γ Induces Endothelial Glucocorticoid Resistance via Sustained Activation of p38 and JNK

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Karolina A. Zielińska

2017-09-01

Full Text Available Malaria-associated acute respiratory distress syndrome (MA-ARDS is an often lethal complication of malaria. Currently, no adequate therapy for this syndrome exists. Although glucocorticoids (GCs have been used to improve clinical outcome of ARDS, their therapeutic benefits remain unclear. We previously developed a mouse model of MA-ARDS, in which dexamethasone treatment revealed GC resistance. In the present study, we investigated GC sensitivity of mouse microvascular lung endothelial cells stimulated with interferon-γ (IFN-γ and Plasmodium berghei NK65 (PbNK65. Upon challenge with IFN-γ alone, dexamethasone inhibited the expression of CCL5 (RANTES by 90% and both CCL2 (MCP-1 and CXCL10 (IP-10 by 50%. Accordingly, whole transcriptome analysis revealed that dexamethasone differentially affected several gene clusters and in particular inhibited a large cluster of IFN-γ-induced genes, including chemokines. In contrast, combined stimulation with IFN-γ and PbNK65 extract impaired inhibitory actions of GCs on chemokine release, without affecting the capacity of the GC receptor to accumulate in the nucleus. Subsequently, we investigated the effects of GCs on two signaling pathways activated by IFN-γ. Dexamethasone left phosphorylation and protein levels of signal transducer and activator of transcription 1 (STAT1 unhampered. In contrast, dexamethasone inhibited the IFN-γ-induced activation of two mitogen-activated protein kinases (MAPK, JNK, and p38. However, PbNK65 extract abolished the inhibitory effects of GCs on MAPK signaling, inducing GC resistance. These data provide novel insights into the mechanisms of GC actions in endothelial cells and show how malaria may impair the beneficial effects of GCs.

Intracellular Distribution of Capsid-Associated pUL77 of Human Cytomegalovirus and Interactions with Packaging Proteins and pUL93.

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Köppen-Rung, Pánja; Dittmer, Alexandra; Bogner, Elke

2016-07-01

DNA packaging into procapsids is a common multistep process during viral maturation in herpesviruses. In human cytomegalovirus (HCMV), the proteins involved in this process are terminase subunits pUL56 and pUL89, which are responsible for site-specific cleavage and insertion of the DNA into the procapsid via portal protein pUL104. However, additional viral proteins are required for the DNA packaging process. We have shown previously that the plasmid that encodes capsid-associated pUL77 encodes another potential player during capsid maturation. Pulse-chase experiments revealed that pUL77 is stably expressed during HCMV infection. Time course analysis demonstrated that pUL77 is expressed in the early late part of the infectious cycle. The sequence of pUL77 was analyzed to find nuclear localization sequences (NLSs), revealing monopartite NLSm at the N terminus and bipartite NLSb in the middle of pUL77. The potential NLSs were inserted into plasmid pHM829, which encodes a chimeric protein with β-galactosidase and green fluorescent protein. In contrast to pUL56, neither NLSm nor NLSb was sufficient for nuclear import. Furthermore, we investigated by coimmunoprecipitation whether packaging proteins, as well as pUL93, the homologue protein of herpes simplex virus 1 pUL17, are interaction partners of pUL77. The interactions between pUL77 and packaging proteins, as well as pUL93, were verified. We showed that the capsid-associated pUL77 is another potential player during capsid maturation of HCMV. Protein UL77 (pUL77) is a conserved core protein of HCMV. This study demonstrates for the first time that pUL77 has early-late expression kinetics during the infectious cycle and an intrinsic potential for nuclear translocation. According to its proposed functions in stabilization of the capsid and anchoring of the encapsidated DNA during packaging, interaction with further DNA packaging proteins is required. We identified physical interactions with terminase subunits pUL56 and p

p,p'-Dichlorodiphenyltrichloroethane (p,p'-DDT) and p,p'-dichlorodiphenyldichloroethylene (p,p'-DDE) repress prostate specific antigen levels in human prostate cancer cell lines.

Science.gov (United States)

Wong, Lilian I L; Labrecque, Mark P; Ibuki, Naokazu; Cox, Michael E; Elliott, John E; Beischlag, Timothy V

2015-03-25

Despite stringent restrictions on their use by many countries since the 1970s, the endocrine disrupting chemicals, DDT and DDE are still ubiquitous in the environment. However, little attention has been directed to p,p'-DDT and the anti-androgen, p,p'-DDE on androgen receptor (AR) target gene transcription in human cells. Inhibitors of androgenic activity may have a deleterious clinical outcome in prostate cancer screens and progression, therefore we determined whether environmentally relevant concentrations of p,p'-DDT and p,p'-DDE negatively impact AR-regulated expression of prostate-specific antigen (PSA), and other AR target genes in human LNCaP and VCaP prostate cancer cells. Quantitative real-time PCR and immuno-blotting techniques were used to measure intracellular PSA, PSMA and AR mRNA and protein levels. We have shown for the first time that p,p'-DDT and p,p'-DDE repressed R1881-inducible PSA mRNA and protein levels in a dose-dependent manner. Additionally, we used the fully automated COBAS PSA detection system to determine that extracellular PSA levels were also significantly repressed. These chemicals achieve this by blocking the recruitment of AR to the PSA promoter region at 10 μM, as demonstrated by the chromatin immunoprecipitation (ChIP) in LNCaP cells. Both p,p'-DDT and p,p'-DDE repressed R1881-inducible AR protein accumulation at 10 μM. Thus, we conclude that men who have been exposed to either DDT or DDE may produce a false-negative PSA test when screening for prostate cancer, resulting in an inaccurate clinical diagnosis. More importantly, prolonged exposure to these anti-androgens may mimic androgen ablation therapy in individuals with prostate cancer, thus exacerbating the condition by inadvertently forcing adaptation to this stress early in the disease. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Significant difference in p53 and p21 protein immunoreactivity in HPV 16 positive and HPV negative breast carcinomas

International Nuclear Information System (INIS)

Hennig, E.M.; Norwegian Radium Hospital, Oslo; Kvinnsland, S.; Holm, R.; Nesland, J.M.

1999-01-01

Human papillomavirus (HPV) 16 has previously been found in 19/41 breast carcinomas (46%) in women with a history of HPV 16 positive CIN III lesions. There was no significant difference in distribution of histological subtypes, mean or median tumour diameter or number of regional lymph node metastases in the HPV positive and HPV negative breast carcinoma groups. P53, p21 and c-erbB-2 proteins were analyzed by immunohistochemistry in the HPV 16 positive and HPV negative breast carcinomas. There was a significant difference in p53 and p21 protein immunoreactivity between HPV 16 positive and HPV negative breast carcinomas (p=0.0091 and p=0.0040), with a significant less detectable p53 and p21 protein immunoreactivity in the HPV 16 positive cases. There was also a significant difference in the coexpression of p53/p21 between the HPV 16 positive and HPV 16 negative breast carcinomas (p=0.002). No significant difference in immunostaining for c-erbB-2 protein in the two groups was found (p=0.15), or for the coexpression of p53/c-erbB-2 (p=0.19). The significantly lower expression of p53 and p21 proteins in HPV 16 positive than in HPV 16 negative breast carcinomas supports the hypothesis of inactivation and degradation of wild-type p53 proteins by HPV 16 E6 and that p53 mutation is not necessary for transformation in the HPV 16 positive cases. (orig.)

Effects of ionizing radiation on expression of P21 protein in Jurkat cell line and p21 gene in thymocytes and splenocytes of mice

International Nuclear Information System (INIS)

Ni Guanying; Wu Ning; Guo Haizhuo; Jin Shunzi

2011-01-01

Objective: To investigate the effects of ionizing radiation on the expression of P21 protein in Jurkat cell line and p21 gene in thymocytes and splenocytes of mice. Methods: Flow cytometry (FCM) was used to analyze the expression of P21 protein in Jurkat cells at 12 and 24 h after irradiation to 0, 0.5, 1.0, 2.0, 4.0, and 6.0 Gy. Real-time PCR was used to detect the expression of p21 gene in thymocytes and splenocytes of mice at 4 and 24 h after irradiation to 0, 0.5, 1.0, 2.0, 4.0, and 6.0 Gy. Multi-staining was used to analyze the micronucleus rates of Rct in bone marrow. Results: The expressions of P21 protein were increased in a dose-dependent manner during 0.5-4.0 Gy (t=-24.23 - -3.96, P<0.05), but decreased at 6.0 Gy at 12 and 24 h post-irradiation (t=-11.19, -14.50, P<0.05). The expressions of p21 gene in both thymocytes and splenocytes of mice were increased in dose-dependent manner in the range of 0-6.0 Gy (including 6.0 Gy) (t=-29.96-8.80, P<0.05), and reached to the peak at 6.0 Gy at 4 and 24 h post-irradiation (t=-11.84 - -3.42, P<0.05), except thymocytes at 4 h and 1.0 Gy post-irradiation (t=-3.42, P>0.05). Conclusions: The expressions of P21 protein and p21 gene could be increased by X-ray irradiation, which shows good dose-dependent manners in certain range of dose. (authors)

Predicting pKa for proteins using COSMO-RS

DEFF Research Database (Denmark)

Andersson, Martin Peter; Jensen, Jan Halborg; Stipp, Susan Louise Svane

2013-01-01

We have used the COSMO-RS implicit solvation method to calculate the equilibrium constants, pKa, for deprotonation of the acidic residues of the ovomucoid inhibitor protein, OMTKY3. The root mean square error for comparison with experimental data is only 0.5 pH units and the maximum error 0.8 p......H units. The results show that the accuracy of pKa prediction using COSMO-RS is as good for large biomolecules as it is for smaller inorganic and organic acids and that the method compares very well to previous pKa predictions of the OMTKY3 protein using Quantum Mechanics/Molecular Mechanics. Our approach...

A proteomic screen reveals the mitochondrial outer membrane protein Mdm34p as an essential target of the F-box protein Mdm30p.

Science.gov (United States)

Ota, Kazuhisa; Kito, Keiji; Okada, Satoshi; Ito, Takashi

2008-10-01

Ubiquitination plays various critical roles in eukaryotic cellular regulation and is mediated by a cascade of enzymes including ubiquitin protein ligase (E3). The Skp1-Cullin-F-box protein complex comprises the largest E3 family, in each member of which a unique F-box protein binds its targets to define substrate specificity. Although genome sequencing uncovers a growing number of F-box proteins, most of them have remained as "orphans" because of the difficulties in identification of their substrates. To address this issue, we tested a quantitative proteomic approach by combining the stable isotope labeling by amino acids in cell culture (SILAC), parallel affinity purification (PAP) that we had developed for efficient enrichment of ubiquitinated proteins, and mass spectrometry (MS). We applied this SILAC-PAP-MS approach to compare ubiquitinated proteins between yeast cells with and without over-expressed Mdm30p, an F-box protein implicated in mitochondrial morphology. Consequently, we identified the mitochondrial outer membrane protein Mdm34p as a target of Mdm30p. Furthermore, we found that mitochondrial defects induced by deletion of MDM30 are not only recapitulated by a mutant Mdm34p defective in interaction with Mdm30p but alleviated by ubiquitination-mimicking forms of Mdm34p. These results indicate that Mdm34p is a physiologically important target of Mdm30p.

Correlation of random urine protein creatinine (P-C) ratio with 24-hour urine protein and P-C ratio, based on physical activity: a pilot study.

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Sadjadi, Seyed-Ali; Jaipaul, Navin

2010-09-07

Quantification of proteinuria is usually predicated upon 24-hour urine collection. Multiple factors influence urine collection and the rate of protein and creatinine excretion. Urine collection is often incomplete, and therefore creatinine and protein excretion rates are underestimated. A random urine protein-creatinine (P-C) ratio has been shown over the years to be a reliable alternative to the 24-hour collection for detection and follow up of proteinuria. However, urine protein excretion may be influenced by physical activity. We studied 48 patients with proteinuria and varying levels of physical activity to determine the correlation between the measures of urine protein excretion. The correlation coefficient (r) between 24-hour urine total protein and random urine P-C ratio was 0.75 (P r = 0.99 (P r = 0.95 (P bedridden patients; r = 0.44 (P = not significant [NS]) and r = 0.54 (P = NS) in semiactive patients; and r = 0.44 (P = NS) and r = 0.58 (P 3500 mg/day) and non-nephrotic (r = 0.84; P r = 0.99 (P r = 0.92 (P bedridden patients; r = 0.61 (P = NS) and r = 0.54 (P = NS) in semiactive patients; and r = 0.64 (P r = 0.52 (P < 0.05) in active patients with nephrotic and non-nephrotic range proteinuria, respectively. We conclude that the random urine P-C ratio is a reliable and practical way of estimating and following proteinuria, but its precision and accuracy may be affected by the level of patient physical activity.

Biochemical and Genetic Evidence that Enterococcus faecium L50 Produces Enterocins L50A and L50B, the sec-Dependent Enterocin P, and a Novel Bacteriocin Secreted without an N-Terminal Extension Termed Enterocin Q

Science.gov (United States)

Cintas, Luis M.; Casaus, Pilar; Herranz, Carmen; Håvarstein, Leiv Sigve; Holo, Helge; Hernández, Pablo E.; Nes, Ingolf F.

2000-01-01

Enterococcus faecium L50 grown at 16 to 32°C produces enterocin L50 (EntL50), consisting of EntL50A and EntL50B, two unmodified non-pediocin-like peptides synthesized without an N-terminal leader sequence or signal peptide. However, the bacteriocin activity found in the cell-free culture supernatants following growth at higher temperatures (37 to 47°C) is not due to EntL50. A purification procedure including cation-exchange, hydrophobic interaction, and reverse-phase liquid chromatography has shown that the antimicrobial activity is due to two different bacteriocins. Amino acid sequences obtained by Edman degradation and DNA sequencing analyses revealed that one is identical to the sec-dependent pediocin-like enterocin P produced by E. faecium P13 (L. M. Cintas, P. Casaus, L. S. Håvarstein, P. E. Hernández, and I. F. Nes, Appl. Environ. Microbiol. 63:4321–4330, 1997) and the other is a novel unmodified non-pediocin-like bacteriocin termed enterocin Q (EntQ), with a molecular mass of 3,980. DNA sequencing analysis of a 963-bp region of E. faecium L50 containing the enterocin P structural gene (entP) and the putative immunity protein gene (entiP) reveals a genetic organization identical to that previously found in E. faecium P13. DNA sequencing analysis of a 1,448-bp region identified two consecutive but diverging open reading frames (ORFs) of which one, termed entQ, encodes a 34-amino-acid protein whose deduced amino acid sequence was identical to that obtained for EntQ by amino acid sequencing, showing that EntQ, similarly to EntL50A and EntL50B, is synthesized without an N-terminal leader sequence or signal peptide. The second ORF, termed orf2, was located immediately upstream of and in opposite orientation to entQ and encodes a putative immunity protein composed of 221 amino acids. Bacteriocin production by E. faecium L50 showed that EntP and EntQ are produced in the temperature range from 16 to 47°C and maximally detected at 47 and 37 to 47�

SUN family proteins Sun4p, Uth1p and Sim1p are secreted from Saccharomyces cerevisiae and produced dependently on oxygen level.

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Evgeny Kuznetsov

Full Text Available The SUN family is comprised of proteins that are conserved among various yeasts and fungi, but that are absent in mammals and plants. Although the function(s of these proteins are mostly unknown, they have been linked to various, often unrelated cellular processes such as those connected to mitochondrial and cell wall functions. Here we show that three of the four Saccharomyces cerevisiae SUN family proteins, Uth1p, Sim1p and Sun4p, are efficiently secreted out of the cells in different growth phases and their production is affected by the level of oxygen. The Uth1p, Sim1p, Sun4p and Nca3p are mostly synthesized during the growth phase of both yeast liquid cultures and colonies. Culture transition to slow-growing or stationary phases is linked with a decreased cellular concentration of Sim1p and Sun4p and with their efficient release from the cells. In contrast, Uth1p is released mainly from growing cells. The synthesis of Uth1p and Sim1p, but not of Sun4p, is repressed by anoxia. All four proteins confer cell sensitivity to zymolyase. In addition, Uth1p affects cell sensitivity to compounds influencing cell wall composition and integrity (such as Calcofluor white and Congo red differently when growing on fermentative versus respiratory carbon sources. In contrast, Uth1p is essential for cell resistance to boric acids irrespective of carbon source. In summary, our novel findings support the hypothesis that SUN family proteins are involved in the remodeling of the yeast cell wall during the various phases of yeast culture development and under various environmental conditions. The finding that Uth1p is involved in cell sensitivity to boric acid, i.e. to a compound that is commonly used as an important antifungal in mycoses, opens up new possibilities of investigating the mechanisms of boric acid's action.

File list: Unc.PSC.50.AllAg.P19 [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available Unc.PSC.50.AllAg.P19 mm9 Unclassified Pluripotent stem cell P19 SRX1098121,SRX10981...20,SRX1098122,SRX1098124,SRX1098123,SRX1098125 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Unc.PSC.50.AllAg.P19.bed ...

Alfalfa mosaic virus replicase proteins P1 and P2 interact and colocalize at the vacuolar membrane

NARCIS (Netherlands)

Heijden, van der M.W.; Carette, J.E.; Reinhoud, P.J.; Haegi, A.; Bol, J.F.

2001-01-01

Replication of Alfalfa mosaic virus (AMV) RNAs depends on the virus-encoded proteins P1 and P2. P1 contains methyltransferase- and helicase-like domains, and P2 contains a polymerase-like domain. Coimmunoprecipitation experiments revealed an interaction between in vitro translated-P1 and P2 and

Identification of an intracellular protein that specifically interacts with photoaffinity-labeled oncogenic p21 protein

International Nuclear Information System (INIS)

Lee, G.; Ronai, Z.A.; Pincus, M.R.; Brandt-Rauf, P.W.; Weinstein, I.B.; Murphy, R.B.; Delohery, T.M.; Nishimura, S.; Yamaizumi, Z.

1989-01-01

An oncogenic 21-kDa (p21) protein (Harvey RAS protein with Val-12) has been covalently modified with a functional reagent that contains a photoactivatable aromatic azide group. This modified p21 protein has been introduced quantitatively into NIH 3T3 cells using an erythrocyte-mediated fusion technique. The introduced p21 protein was capable of inducing enhanced pinocytosis and DNA synthesis in the recipient cells. To identify the putative intracellular protein(s) that specifically interact with modified p21 protein, the cells were pulsed with [ 35 S]methionine at selected times after fusion and then UV-irradiated to activate the azide group. The resulting nitrene covalently binds to amino acid residues in adjacent proteins, thus linking the p21 protein to these proteins. The cells were then lysed, and the lysate was immunoprecipitated with the anti-p21 monoclonal antibody Y13-259. The immunoprecipitate was analyzed by SDS/PAGE to identify p21 - protein complexes. By using this technique, the authors found that three protein complexes of 51, 64, and 82 kDa were labeled specifically and reproducibly. The most prominent band is the 64-kDa protein complex that shows a time-dependent rise and fall, peaking within a 5-hr period after introduction of the p21 protein the cells. These studies provide evidence that in vitro the p21 protein becomes associated with a protein whose mass is about 43 kDa. They suggest that the formation of this complex may play a role in mediating early events involved with cell transformation induced by RAS oncogenes

Analysis of the protein-protein interactions between the human acidic ribosomal P-proteins: evaluation by the two hybrid system

DEFF Research Database (Denmark)

Tchórzewski, M; Boldyreff, B; Issinger, O

2000-01-01

The surface acidic ribosomal proteins (P-proteins), together with ribosomal core protein P0 form a multimeric lateral protuberance on the 60 S ribosomal subunit. This structure, also called stalk, is important for efficient translational activity of the ribosome. In order to shed more light...... forms the 60 S ribosomal stalk: P0-(P1/P2)(2). Additionally, mutual interactions among human and yeast P-proteins were analyzed. Heterodimer formation could be observed between human P2 and yeast P1 proteins....

Biochemical and functional analysis of two Plasmodium falciparum blood-stage 6-cys proteins: P12 and P41.

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Tana Taechalertpaisarn

Full Text Available The genomes of Plasmodium parasites that cause malaria in humans, other primates, birds, and rodents all encode multiple 6-cys proteins. Distinct 6-cys protein family members reside on the surface at each extracellular life cycle stage and those on the surface of liver infective and sexual stages have been shown to play important roles in hepatocyte growth and fertilization respectively. However, 6-cys proteins associated with the blood-stage forms of the parasite have no known function. Here we investigate the biochemical nature and function of two blood-stage 6-cys proteins in Plasmodium falciparum, the most pathogenic species to afflict humans. We show that native P12 and P41 form a stable heterodimer on the infective merozoite surface and are secreted following invasion, but could find no evidence that this complex mediates erythrocyte-receptor binding. That P12 and P41 do not appear to have a major role as adhesins to erythrocyte receptors was supported by the observation that antisera to these proteins did not substantially inhibit erythrocyte invasion. To investigate other functional roles for these proteins their genes were successfully disrupted in P. falciparum, however P12 and P41 knockout parasites grew at normal rates in vitro and displayed no other obvious phenotypic changes. It now appears likely that these blood-stage 6-cys proteins operate as a pair and play redundant roles either in erythrocyte invasion or in host-immune interactions.

CHCHD10 mutations p.R15L and p.G66V cause motoneuron disease by haploinsufficiency.

Science.gov (United States)

Brockmann, Sarah J; Freischmidt, Axel; Oeckl, Patrick; Müller, Kathrin; Ponna, Srinivas K; Helferich, Anika M; Paone, Christoph; Reinders, Jörg; Kojer, Kerstin; Orth, Michael; Jokela, Manu; Auranen, Mari; Udd, Bjarne; Hermann, Andreas; Danzer, Karin M; Lichtner, Peter; Walther, Paul; Ludolph, Albert C; Andersen, Peter M; Otto, Markus; Kursula, Petri; Just, Steffen; Weishaupt, Jochen H

2018-02-15

Mutations in the mitochondrially located protein CHCHD10 cause motoneuron disease by an unknown mechanism. In this study, we investigate the mutations p.R15L and p.G66V in comparison to wild-type CHCHD10 and the non-pathogenic variant p.P34S in vitro, in patient cells as well as in the vertebrate in vivo model zebrafish. We demonstrate a reduction of CHCHD10 protein levels in p.R15L and p.G66V mutant patient cells to approximately 50%. Quantitative real-time PCR revealed that expression of CHCHD10 p.R15L, but not of CHCHD10 p.G66V, is already abrogated at the mRNA level. Altered secondary structure and rapid protein degradation are observed with regard to the CHCHD10 p.G66V mutant. In contrast, no significant differences in expression, degradation rate or secondary structure of non-pathogenic CHCHD10 p.P34S are detected when compared with wild-type protein. Knockdown of CHCHD10 expression in zebrafish to about 50% causes motoneuron pathology, abnormal myofibrillar structure and motility deficits in vivo. Thus, our data show that the CHCHD10 mutations p.R15L and p.G66V cause motoneuron disease primarily based on haploinsufficiency of CHCHD10. © The Author(s) 2018. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Protein mobilities and P-selectin storage in Weibel-Palade bodies.

Science.gov (United States)

Kiskin, Nikolai I; Hellen, Nicola; Babich, Victor; Hewlett, Lindsay; Knipe, Laura; Hannah, Matthew J; Carter, Tom

2010-09-01

Using fluorescence recovery after photobleaching (FRAP) we measured the mobilities of EGFP-tagged soluble secretory proteins in the endoplasmic reticulum (ER) and in individual Weibel-Palade bodies (WPBs) at early (immature) and late (mature) stages in their biogenesis. Membrane proteins (P-selectin, CD63, Rab27a) were also studied in individual WPBs. In the ER, soluble secretory proteins were mobile; however, following insertion into immature WPBs larger molecules (VWF, Proregion, tPA) and P-selectin became immobilised, whereas small proteins (ssEGFP, eotaxin-3) became less mobile. WPB maturation led to further decreases in mobility of small proteins and CD63. Acute alkalinisation of mature WPBs selectively increased the mobilities of small soluble proteins without affecting larger molecules and the membrane proteins. Disruption of the Proregion-VWF paracrystalline core by prolonged incubation with NH(4)Cl rendered P-selectin mobile while VWF remained immobile. FRAP of P-selectin mutants revealed that immobilisation most probably involves steric entrapment of the P-selectin extracellular domain by the Proregion-VWF paracrystal. Significantly, immobilisation contributed to the enrichment of P-selectin in WPBs; a mutation of P-selectin preventing immobilisation led to a failure of enrichment. Together these data shed new light on the transitions that occur for soluble and membrane proteins following their entry and storage into post-Golgi-regulated secretory organelles.

Angiotensin II stimulates basolateral 50-pS K channels in the thick ascending limb.

Science.gov (United States)

Wang, Mingxiao; Luan, Haiyan; Wu, Peng; Fan, Lili; Wang, Lijun; Duan, Xinpeng; Zhang, Dandan; Wang, Wen-Hui; Gu, Ruimin

2014-03-01

We used the patch-clamp technique to examine the effect of angiotensin II (ANG II) on the basolateral K channels in the thick ascending limb (TAL) of the rat kidney. Application of ANG II increased the channel activity and the current amplitude of the basolateral 50-pS K channel. The stimulatory effect of ANG II on the K channels was completely abolished by losartan, an inhibitor of type 1 angiotensin receptor (AT1R), but not by PD123319, an AT2R antagonist. Moreover, inhibition of phospholipase C (PLC) and protein kinase C (PKC) also abrogated the stimulatory effect of ANG II on the basolateral K channels in the TAL. This suggests that the stimulatory effect of ANG II on the K channels was induced by activating PLC and PKC pathways. Western blotting demonstrated that ANG II increased the phosphorylation of c-Src at tyrosine residue 416, an indication of c-Src activation. This effect was mimicked by PKC stimulator but abolished by calphostin C. Moreover, inhibition of NADPH oxidase (NOX) also blocked the effect of ANG II on c-Src tyrosine phosphorylation. The role of Src-family protein tyrosine kinase (SFK) in mediating the effect of ANG II on the basolateral K channel was further suggested by the experiments in which inhibition of SFK abrogated the stimulatory effect of ANG II on the basolateral 50-pS K channel. We conclude that ANG II increases basolateral 50-pS K channel activity via AT1R and that activation of AT1R stimulates SFK by a PLC-PKC-NOX-dependent mechanism.

Construction of a time-of-flight neutron spectrometer for reaction angles 000 and study of the reaction 65Cu(p,xn) 65Zn for Esub(p)=26.7 MeV

International Nuclear Information System (INIS)

Holler, Y.

1984-01-01

At the Hamburg Isochronous Cyclotron a novel time-of-flight neutron spectrometer was designed, constructed, and tested by means of a for the planned application typical nuclear reaction. The apparature was optimized for the measurement of continuous, structure-deficient neutron spectra in a wide angular range at a reproducible, as low as possible scattering neutron background. Such a facility is fitted to the strengths of the Hamburg cyclotron and allows to study questions on the precompound emission and on the inelastic projectile ( 3 He) breakup. The final test was performed with the reaction 65 Cu(p,xn) 65 Zn at Esub(p)=26.7 MeV for which already comparable data over a smaller angular range were present. In the analysis of the measurement results performed regarding the precompound effects the hybrid-exciton model calculations let recognize essential deviations at high neutron energies in the range of the extreme reaction angles. (orig./HSI) [de

Identification of an intracellular protein that specifically interacts with photoaffinity-labeled oncogenic p21 protein.

Science.gov (United States)

Lee, G; Ronai, Z A; Pincus, M R; Brandt-Rauf, P W; Murphy, R B; Delohery, T M; Nishimura, S; Yamaizumi, Z; Weinstein, I B

1989-11-01

An oncogenic 21-kDa (p21) protein (Harvey RAS protein with Val-12) has been covalently modified with a functional reagent that contains a photoactivatable aromatic azide group. This modified p21 protein has been introduced quantitatively into NIH 3T3 cells using an erythrocyte-mediated fusion technique. The introduced p21 protein was capable of inducing enhanced pinocytosis and DNA synthesis in the recipient cells. To identify the putative intracellular protein(s) that specifically interact with the modified p21 protein, the cells were pulsed with [35S]methionine at selected times after fusion and then UV-irradiated to activate the azide group. The resulting nitrene covalently binds to amino acid residues in adjacent proteins, thus linking the p21 protein to these proteins. The cells were then lysed, and the lysate was immunoprecipitated with the anti-p21 monoclonal antibody Y13-259. The immunoprecipitate was analyzed by SDS/PAGE to identify p21-protein complexes. By using this technique, we found that three protein complexes of 51, 64, and 82 kDa were labeled specifically and reproducibly. The most prominent band is the 64-kDa protein complex that shows a time-dependent rise and fall, peaking within a 5-hr period after introduction of the p21 protein into the cells. These studies provide evidence that in vitro the p21 protein becomes associated with a protein whose mass is about 43 kDa. We suggest that the formation of this complex may play a role in mediating early events involved with cell transformation induced by RAS oncogenes.

The Prognostic Role of NEDD9 and P38 Protein Expression Levels in Urinary Bladder Transitional Cell Carcinoma

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Ola A. Harb

2017-01-01

Full Text Available Background. The most common malignant tumor of the urinary bladder is transitional cell carcinoma (TCC. Neural precursor cell-expressed developmentally downregulated protein 9 (NEDD9 is found to be a cell adhesion mediator. P38 Mitogen-Activated Protein Kinase is a serine/threonine kinases member which can mediate carcinogenesis through intracellular signaling. Methods. To assess their prognostic role; NEDD9 and p38 protein were evaluated in sections from 50 paraffin blocks of TCC. Results. The high expressions of NEDD9 and p38 protein were significantly associated with grade, stage, distant metastasis (p<0.001, number of tumors, lymph node metastasis, and tumor size (p<0.001, 0.002; 0.018, <0.001; and 0.004, 0.007, respectively. High NEDD9 and p38 detection had a worse 3-year OS (p=0.041 and <0.001, respectively. By multivariate analysis the NEDD9 and p38 protein expression levels and various clinicopathological criteria including gender, grade, stage of the tumor, and regional lymph node involvement were independent prognostic parameters of TCC of the urinary bladder patients’ outcome. Conclusion. NEDD9 and p38 protein expressions were poor prognostic markers of TCC.

Mismatch Negativity and P50 Sensory Gating in Abstinent Former Cannabis Users

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Samantha J. Broyd

2016-01-01

Full Text Available Prolonged heavy exposure to cannabis is associated with impaired cognition and brain functional and structural alterations. We recently reported attenuated mismatch negativity (MMN and altered P50 sensory gating in chronic cannabis users. This study investigated the extent of brain functional recovery (indexed by MMN and P50 in chronic users after cessation of use. Eighteen ex-users (median 13.5 years prior regular use; median 3.5 years abstinence and 18 nonusers completed (1 a multifeature oddball task with duration, frequency, and intensity deviants and (2 a P50 paired-click paradigm. Trend level smaller duration MMN amplitude and larger P50 ratios (indicative of poorer sensory gating were observed in ex-users compared to controls. Poorer P50 gating correlated with prior duration of cannabis use. Duration of abstinence was positively correlated with duration MMN amplitude, even after controlling for age and duration of cannabis use. Impaired sensory gating and attenuated MMN amplitude tended to persist in ex-users after prolonged cessation of use, suggesting a lack of full recovery. An association with prolonged duration of prior cannabis use may indicate persistent cannabis-related alterations to P50 sensory gating. Greater reductions in MMN amplitude with increasing abstinence (positive correlation may be related to either self-medication or an accelerated aging process.

Synthesis of P-protein in mature phloem of Cucurbita maxima.

Science.gov (United States)

Nuske, J; Eschrich, W

1976-01-01

Cotyledons of Cucurbita maxima Duch. seedlings were provided with (14)C-labeled amino acids for 12 h. Besides the bulk of labeled amino acids the sieve-tube exudate also carried labeled proteins. 80% of the incorporated radioactivity was found in the P-protein, 20% in a neutral protein, and traces were found in acidic proteins after fractionation on diethyl-aminoethyl cellulose columns. The radioactive elutes were characterized by autoradiographs of both disc- and sodium dodecyl sulfate-gelelectropherograms, and by isoelectric focusing. The P-protein fraction appeared with the void volume from the diethylaminoethyl-cellulose column. Obviously, this is the protein that gels when oxidized and that is reversibly precipitable giving rise to filaments when processed for electron microscopy. Its main component has a molecular weight of 115,000 Dalton. By isoelectric focusing this fraction separated into 3 proteins with isoelectric points of 9.8, 9.4, and 9.2. The isoelectric point 9.2-protein probably is identical with an oligomer of a 30,000 Dalton protein with neutral isoelectric point, which keeps 20% of the incorporated label. Microautoradiographs suggest that the labeled proteins were synthesized in companion cells. The results indicate that P-protein of Cucurbita maxima is synthesized continuously in mature phloem. It can be assumed that P-protein has a relatively high turn-over rate. Therefore it seems unlikely that P-protein is a "structural" protein.

Correlation of random urine protein creatinine (P-C ratio with 24-hour urine protein and P-C ratio, based on physical activity: a pilot study

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Seyed-Ali Sadjadi

2010-07-01

Full Text Available Seyed-Ali Sadjadi1,2, Navin Jaipaul1,21Jerry L Pettis Memorial VA Medical Center, 2Loma Linda University School of Medicine, Loma Linda, CA, USAAbstract: Quantification of proteinuria is usually predicated upon 24-hour urine collection. Multiple factors influence urine collection and the rate of protein and creatinine excretion. Urine collection is often incomplete, and therefore creatinine and protein excretion rates are underestimated. A random urine protein-creatinine (P-C ratio has been shown over the years to be a reliable alternative to the 24-hour collection for detection and follow up of proteinuria. However, urine protein excretion may be influenced by physical activity. We studied 48 patients with proteinuria and varying levels of physical activity to determine the correlation between the measures of urine protein excretion. The correlation coefficient (r between 24-hour urine total protein and random urine P-C ratio was 0.75 (P < 0.01 in the overall study population, but varied according to the level of proteinuria and physical activity in a stratified analysis: r = 0.99 (P < 0.001 and r = 0.95 (P < 0.01 in bedridden patients; r = 0.44 (P = not significant [NS] and r = 0.54 (P = NS in semiactive patients; and r = 0.44 (P = NS and r = 0.58 (P < 0.05 in active patients with nephrotic- (>3500 mg/day and non-nephrotic (<3500 mg/day range proteinuria, respectively. The correlation appeared to be stronger between random urine and 24-hour urine P-C ratio for the overall study population (r = 0.84; P < 0.001, and when stratified according to the level of proteinuria and physical activity: r = 0.99 (P < 0.001 and r = 0.92 (P < 0.01 in bedridden patients; r = 0.61 (P = NS and r = 0.54 (P = NS in semiactive patients; and r = 0.64 (P < 0.02 and r = 0.52 (P < 0.05 in active patients with nephrotic and non-nephrotic range proteinuria, respectively. We conclude that the random urine P-C ratio is a reliable and practical way of estimating and

Phosphorylation of the human respiratory syncytial virus P protein mediates M2-2 regulation of viral RNA synthesis, a process that involves two P proteins.

Science.gov (United States)

Asenjo, Ana; Villanueva, Nieves

2016-01-04

The M2-2 protein regulates the balance between human respiratory syncytial virus (HRSV) transcription and replication. Here it is shown that M2-2 mediated transcriptional inhibition is managed through P protein phosphorylation. Transcription inhibition by M2-2 of the HRSV based minigenome pRSVluc, required P protein phosphorylation at serines (S) in positions 116, 117, 119 and increased inhibition is observed if S232 or S237 is also phosphorylated. Phosphorylation of these residues is required for viral particle egression from infected cells. Viral RNA synthesis complementation assays between P protein variants, suggest that two types of P proteins participate in the process as components of RNA dependent RNA polymerase (RdRp). Type I is only functional when, as a homotetramer, it is bound to N and L proteins through residues 203-241. Type II is functionally independent of these interactions and binds to N protein at a region outside residues 232-241. P protein type I phosphorylation at S116, S117 and S119, did not affect the activity of RdRp but this phosphorylation in type II avoids its interaction with N protein and impairs RdRp functionality for transcription and replication. Structural changes in the RdRp, mediated by phosphorylation turnover at the indicated residues, in the two types of P proteins, may result in a fine adjustment, late in the infectious cycle, of transcription, replication and progression in the morphogenetic process that ends in egression of the viral particles from infected cells. Copyright © 2015 Elsevier B.V. All rights reserved.

File list: InP.CDV.50.AllAg.Cardiomyocytes [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.CDV.50.AllAg.Cardiomyocytes mm9 Input control Cardiovascular Cardiomyocytes SRX...1121694 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.CDV.50.AllAg.Cardiomyocytes.bed ...

The amino terminal end determines the stability and assembling capacity of eukaryotic ribosomal stalk proteins P1 and P2.

Science.gov (United States)

Camargo, Hendricka; Nusspaumer, Gretel; Abia, David; Briceño, Verónica; Remacha, Miguel; Ballesta, Juan P G

2011-05-01

The eukaryotic ribosomal proteins P1 and P2 bind to protein P0 through their N-terminal domain to form the essential ribosomal stalk. A mutational analysis points to amino acids at positions 2 and 3 as determinants for the drastic difference of Saccharomyces cerevisiae P1 and P2 half-life, and suggest different degradation mechanisms for each protein type. Moreover, the capacity to form P1/P2 heterodimers is drastically affected by mutations in the P2β four initial amino acids, while these mutations have no effect on P1β. Binding of P2β and, to a lesser extent, P1β to the ribosome is also seriously affected showing the high relevance of the amino acids in the first turn of the NTD α-helix 1 for the stalk assembly. The negative effect of some mutations on ribosome binding can be reversed by the presence of the second P1/P2 couple in the ribosome, indicating a stabilizing structural influence between the two heterodimers. Unexpectedly, some mutations totally abolish heterodimer formation but allow significant ribosome binding and, therefore, a previous P1 and P2 association seems not to be an absolute requirement for stalk assembly. Homology modeling of the protein complexes suggests that the mutated residues can affect the overall protein conformation. © The Author(s) 2011. Published by Oxford University Press.

Genistein inhibits phorbol ester-induced NF-κB transcriptional activity and COX-2 expression by blocking the phosphorylation of p65/RelA in human mammary epithelial cells

Energy Technology Data Exchange (ETDEWEB)

Chung, Myung-Hoon; Kim, Do-Hee [Research Institute for Pharmaceutical Sciences, College of Pharmacy, Seoul National University, Seoul (Korea, Republic of); Na, Hye-Kyung [Department of Food and Nutrition, Sungshin Women' s University, Seoul (Korea, Republic of); Kim, Jung-Hwan; Kim, Ha-Na [Research Institute for Pharmaceutical Sciences, College of Pharmacy, Seoul National University, Seoul (Korea, Republic of); Haegeman, Guy [LEGEST, University of Gent (Belgium); Surh, Young-Joon, E-mail: surh@snu.ac.kr [Research Institute for Pharmaceutical Sciences, College of Pharmacy, Seoul National University, Seoul (Korea, Republic of); Department of Molecular Medicine and Biopharmaceutical Sciences, Graduate School of Convergence Science and Technology, Seoul (Korea, Republic of); Cancer Research Institute, Seoul National University, Seoul (Korea, Republic of)

2014-10-15

Genistein, an isoflavone present in soy products, has chemopreventive effects on mammary carcinogenesis. In the present study, we have investigated the effects of genistein on phorbol ester-induced expression of cyclooxygenase-2 (COX-2) that plays an important role in the pathophysiology of inflammation-associated carcinogenesis. Pretreatment of cultured human breast epithelial (MCF10A) cells with genistein reduced COX-2 expression induced by 12-O-tetradecanoylphorbol-13-acetate (TPA). There are multiple lines of evidence supporting that the induction of COX-2 is regulated by the eukaryotic transcription factor NF-κB. Genistein failed to inhibit TPA-induced nuclear translocation and DNA binding of NF-κB as well as degradation of IκB. However, genistein abrogated the TPA-induced transcriptional activity of NF-κB as determined by the luciferase reporter gene assay. Genistein inhibited phosphorylation of the p65 subunit of NF-κB and its interaction with cAMP regulatory element-binding protein-binding protein (CBP)/p300 and TATA-binding protein (TBP). TPA-induced NF-κB phosphorylation was abolished by pharmacological inhibition of extracellular signal-regulated kinase (ERK). Likewise, pharmacologic inhibition or dominant negative mutation of ERK suppressed phosphorylation of p65. The above findings, taken together, suggest that genistein inhibits TPA-induced COX-2 expression in MCF10A cells by blocking ERK-mediated phosphorylation of p65 and its subsequent interaction with CBP and TBP.

Caspase-3-mediated cleavage of p65/RelA results in a carboxy-terminal fragment that inhibits IκBα and enhances HIV-1 replication in human T lymphocytes

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Alcamí José

2008-12-01

Full Text Available Abstract Background Degradation of p65/RelA has been involved in both the inhibition of NF-κB-dependent activity and the onset of apoptosis. However, the mechanisms of NF-κB degradation are unclear and can vary depending on the cell type. Cleavage of p65/RelA can produce an amino-terminal fragment that was shown to act as a dominant-negative inhibitor of NF-κB, thereby promoting apoptosis. However, the opposite situation has also been described and the production of a carboxy-terminal fragment that contains two potent transactivation domains has also been related to the onset of apoptosis. In this context, a carboxy-terminal fragment of p65/RelA (ΔNH2p65, detected in non-apoptotic human T lymphocytes upon activation, has been studied. T cells constitute one of the long-lived cellular reservoirs of the human immunodeficiency virus type 1 (HIV-1. Because NF-κB is the most important inducible element involved in initiation of HIV-1 transcription, an adequate control of NF-κB response is of paramount importance for both T cell survival and viral spread. Its major inhibitor IκBα constitutes a master terminator of NF-κB response that is complemented by degradation of p65/RelA. Results and conclusions In this study, the function of a caspase-3-mediated carboxy-terminal fragment of p65/RelA, which was detected in activated human peripheral blood lymphocytes (PBLs, was analyzed. Cells producing this truncated p65/RelA did not undergo apoptosis but showed a high viability, in spite of caspase-3 activation. ΔNH2p65 lacked most of DNA-binding domain but retained the dimerization domain, NLS and transactivation domains. Consequently, it could translocate to the nucleus, associate with NF-κB1/p50 and IκBα, but could not bind -κB consensus sites. However, although ΔNH2p65 lacked transcriptional activity by itself, it could increase NF-κB activity in a dose-dependent manner by hijacking IκBα. Thus, its expression resulted in a persistent

Functional analyses of GB virus B p13 protein: development of a recombinant GB virus B hepatitis virus with a p7 protein

DEFF Research Database (Denmark)

Takikawa, Shingo; Engle, Ronald E; Emerson, Suzanne U

2006-01-01

GB virus B (GBV-B), which infects tamarins, is the virus most closely related to hepatitis C virus (HCV). HCV has a protein (p7) that is believed to form an ion channel. It is critical for viability. In vitro studies suggest that GBV-B has an analogous but larger protein (p13). We found...... plus part of p7) was nonviable. However, a mutant lacking amino acid 614-669 (p6) produced high titer viremia and acute resolving hepatitis; viruses recovered from both animals lacked the deleted sequence and had no other mutations. Thus, p6 was dispensable but p7 was essential for infectivity...... processing at both sites, suggesting that p13 is processed into two components (p6 and p7). Mutants with substitution at amino acid 669 or 681 were viable in vivo, but the recovered viruses had changes at amino acid 669 and 681, respectively, which restored cleavage. A mutant lacking amino acid 614-681 (p6...

Functional analyses of GB virus B p13 protein: Development of a recombinant GB virus B hepatitis virus with a p7 protein

DEFF Research Database (Denmark)

Takikawa, Shingo; Engle, Ronald E; Emerson, Suzanne U

2006-01-01

GB virus B (GBV-B), which infects tamarins, is the virus most closely related to hepatitis C virus (HCV). HCV has a protein (p7) that is believed to form an ion channel. It is critical for viability. In vitro studies suggest that GBV-B has an analogous but larger protein (p13). We found...... plus part of p7) was nonviable. However, a mutant lacking amino acid 614-669 (p6) produced high titer viremia and acute resolving hepatitis; viruses recovered from both animals lacked the deleted sequence and had no other mutations. Thus, p6 was dispensable but p7 was essential for infectivity...... processing at both sites, suggesting that p13 is processed into two components (p6 and p7). Mutants with substitution at amino acid 669 or 681 were viable in vivo, but the recovered viruses had changes at amino acid 669 and 681, respectively, which restored cleavage. A mutant lacking amino acid 614-681 (p6...

Chronology of p53 protein accumulation in gastric carcinogenesis

NARCIS (Netherlands)

Craanen, M. E.; Blok, P.; Dekker, W.; Offerhaus, G. J.; Tytgat, G. N.

1995-01-01

p53 Protein accumulation in early gastric carcinoma was studied in relation to the histological type (Lauren classification) and the type of growth pattern, including the chronology of p53 protein accumulation during carcinogenesis. Forty five, paraffin embedded gastrectomy specimens from early

File list: InP.Liv.50.AllAg.Hepatocytes [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Liv.50.AllAg.Hepatocytes hg19 Input control Liver Hepatocytes SRX530185,SRX5301...83 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Liv.50.AllAg.Hepatocytes.bed ...

HSF1 and NF-κB p65 participate in the process of exercise preconditioning attenuating pressure overload-induced pathological cardiac hypertrophy

Energy Technology Data Exchange (ETDEWEB)

Xu, Tongyi [Department of Cardiothoracic Surgery, No. 401 Hospital of PLA, Qingdao (China); Department of Cardiothoracic Surgery, Changhai Hospital, Second Military Medical University, Shanghai (China); Zhang, Ben [Centre of Cardiovascular Surgery, Guangzhou General Hospital of Guangzhou Military Region, Guangzhou (China); Department of Cardiothoracic Surgery, Changhai Hospital, Second Military Medical University, Shanghai (China); Yang, Fan; Cai, Chengliang; Wang, Guokun [Department of Cardiothoracic Surgery, Changhai Hospital, Second Military Medical University, Shanghai (China); Han, Qingqi, E-mail: handoctor@gmail.com [Department of Cardiothoracic Surgery, Changhai Hospital, Second Military Medical University, Shanghai (China); Zou, Liangjian, E-mail: zouliangjiansh@gmail.com [Department of Cardiothoracic Surgery, Changhai Hospital, Second Military Medical University, Shanghai (China)

2015-05-08

Pathological cardiac hypertrophy, often accompanied by hypertension, aortic stenosis and valvular defects, is typically associated with myocyte remodeling and cardiac dysfunction. Exercise preconditioning (EP) has been proven to enhance the tolerance of the myocardium to cardiac ischemia-reperfusion injury. However, the effects of EP in pathological cardiac hypertrophy are rarely reported. 10-wk-old male Sprague–Dawley rats (n = 80) were randomly divided into four groups: sham, TAC, EP + sham and EP + TAC. Two EP groups were subjected to 4 weeks of treadmill training, and the EP + TAC and TAC groups were followed by TAC operations. The sham and EP + sham groups underwent the same operation without aortic constriction. Eight weeks after the surgery, we evaluated the effects of EP by echocardiography, morphology, and histology and observed the expressions of the associated proteins. Compared with the respective control groups, hypertrophy-related indicators were significantly increased in the TAC and EP + TAC groups (p < 0.05). However, between the TAC and EP + TAC groups, all of these changes were effectively inhibited by EP treatment (p < 0.05). Furthermore, EP treatment upregulated the expression of HSF1 and HSP70, increased the HSF1 levels in the nuclear fraction, inhibited the expression of the NF-κB p65 subunit, decreased the NF-κB p65 subunit levels in the nuclear fraction, and reduced the IL2 levels in the myocardia of rats. EP could effectively reduce the cardiac hypertrophic responses induced by TAC and may play a protective role by upregulating the expressions of HSF1 and HSP70, activating HSF1 and then inhibiting the expression of NF-κB p65 and nuclear translocation. - Highlights: • EP could effectively reduce the cardiac hypertrophic responses induced by TAC. • EP may play a protective role by upregulating the expressions of HSF1 and HSP70 and then activating HSF1. • EP may play a protective role by inhibiting the expression

Improving the expression of recombinant proteins in E. coli BL21 (DE3) under acetate stress: an alkaline pH shift approach.

Science.gov (United States)

Wang, Hengwei; Wang, Fengqing; Wang, Wei; Yao, Xueling; Wei, Dongzhi; Cheng, Hairong; Deng, Zixin

2014-01-01

Excess acetate has long been an issue for the production of recombinant proteins in E. coli cells. Recently, improvements in acetate tolerance have been achieved through the use of genetic strategies and medium supplementation with certain amino acids and pyrimidines. The aim of our study was to evaluate an alternative to improve the acetate tolerance of E. coli BL21 (DE3), a popular strain used to express recombinant proteins. In this work we reported the cultivation of BL21 (DE3) in complex media containing acetate at high concentrations. In the presence of 300 mM acetate, compared with pH 6.5, pH 7.5 improved cell growth by approximately 71%, reduced intracellular acetate by approximately 50%, and restored the expression of glutathione S-transferase (GST), green fluorescent protein (GFP) and cytochrome P450 monooxygenase (CYP). Further experiments showed that alkaline pHs up to 8.5 had little inhibition in the expression of GST, GFP and CYP. In addition, the detrimental effect of acetate on the reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) by the cell membrane, an index of cellular metabolic capacity, was substantially alleviated by a shift to alkaline pH values of 7.5-8.0. Thus, we suggest an approach of cultivating E. coli BL21 (DE3) at pH 8.0 ± 0.5 to minimize the effects caused by acetate stress. The proposed strategy of an alkaline pH shift is a simple approach to solving similar bioprocessing problems in the production of biofuels and biochemicals from sugars.

Binding of the cyclic AMP receptor protein of Escherichia coli and DNA bending at the P4 promoter of pBR322.

Science.gov (United States)

Brierley, I; Hoggett, J G

1992-07-01

The binding of the Escherichia coli cyclic AMP receptor protein (CRP) to its specific site on the P4 promoter of pBR322 has been studied by gel electrophoresis. Binding to the P4 site was about 40-50-fold weaker than to the principal CRP site on the lactose promoter at both low (0.01 M) and high (0.1 M) ionic strengths. CRP-induced bending at the P4 site was investigated from the mobilities of CRP bound to circularly permuted P4 fragments. The estimated bending angle, based on comparison with Zinkel & Crothers [(1990) Biopolymers 29, 29-38] A-tract bending standards, was found to be approximately 96 degrees, similar to that found for binding to the lac site. These observations suggest that there is not a simple relationship between strength of CRP binding and the extent of induced bending for different CRP sites. The apparent centre of bending in P4 is displaced about 6-8 bp away from the conserved TGTGA sequence and the P4 transcription start site.

File list: InP.Myo.50.AllAg.HSMM [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Myo.50.AllAg.HSMM hg19 Input control Muscle HSMM SRX067399,SRX067414,SRX038600,...SRX038601 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Myo.50.AllAg.HSMM.bed ...

File list: InP.Liv.50.AllAg.Hepatocytes [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Liv.50.AllAg.Hepatocytes mm9 Input control Liver Hepatocytes SRX019015,SRX55553...3,SRX1334843 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Liv.50.AllAg.Hepatocytes.bed ...

BiP clustering facilitates protein folding in the endoplasmic reticulum.

Directory of Open Access Journals (Sweden)

Marc Griesemer

2014-07-01

Full Text Available The chaperone BiP participates in several regulatory processes within the endoplasmic reticulum (ER: translocation, protein folding, and ER-associated degradation. To facilitate protein folding, a cooperative mechanism known as entropic pulling has been proposed to demonstrate the molecular-level understanding of how multiple BiP molecules bind to nascent and unfolded proteins. Recently, experimental evidence revealed the spatial heterogeneity of BiP within the nuclear and peripheral ER of S. cerevisiae (commonly referred to as 'clusters'. Here, we developed a model to evaluate the potential advantages of accounting for multiple BiP molecules binding to peptides, while proposing that BiP's spatial heterogeneity may enhance protein folding and maturation. Scenarios were simulated to gauge the effectiveness of binding multiple chaperone molecules to peptides. Using two metrics: folding efficiency and chaperone cost, we determined that the single binding site model achieves a higher efficiency than models characterized by multiple binding sites, in the absence of cooperativity. Due to entropic pulling, however, multiple chaperones perform in concert to facilitate the resolubilization and ultimate yield of folded proteins. As a result of cooperativity, multiple binding site models used fewer BiP molecules and maintained a higher folding efficiency than the single binding site model. These insilico investigations reveal that clusters of BiP molecules bound to unfolded proteins may enhance folding efficiency through cooperative action via entropic pulling.

File list: InP.Gon.50.AllAg.Testis [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Gon.50.AllAg.Testis hg19 Input control Gonad Testis SRX663452,SRX663439,SRX6634...45,SRX663442 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Gon.50.AllAg.Testis.bed ...

File list: InP.Gon.50.AllAg.Testis [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Gon.50.AllAg.Testis mm9 Input control Gonad Testis SRX112534,SRX349389,SRX02744...X112541,SRX118389 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Gon.50.AllAg.Testis.bed ...

File list: InP.Epd.50.AllAg.BJ [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Epd.50.AllAg.BJ hg19 Input control Epidermis BJ SRX490265,SRX130064,SRX080433,S...RX190018 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Epd.50.AllAg.BJ.bed ...

In vitro studies of immunoglobulin heavy-chain binding protein (BiP, GRP78). Interactions of BiP with newly synthesized proteins and adenine nucleotides

International Nuclear Information System (INIS)

Kassenbrock, C.K.

1988-01-01

Here we examine the interaction of BiP with newly synthesized polypeptides in an in vitro protein translations-translocation system. We find that BiP forms tight complexes with nonglycosylated yeast invertase and incorrectly disulfide-bonded prolactin but not with glycosylated invertase or correctly disulfide-bonded prolactin. Moreover, BiP associates detectably only with completed chains of prolactin, not with chains undergoing synthesis. We conclude that BiP recognizes and binds with high affinity to aberrantly folded or aberrantly glycosylated polypeptides in vitro, but not to all nascent chains as they are folding. BiP also binds APT and can be purified by APT affinity chromatography. We show that submicromolar levels of ATP or ADP decrease the rate of absorption of 125 I-BiP to nitrocellulose filters coated with protein or nonionic detergents. ATP and ADP also protect portions of BiP from proteolytic degradation. In contrast, micromolar levels of AMP increase the rate of adsorption and the rate of proteolytic degradation of BiP. We also show that an ATPase activity co-purifies with BiP, but its slow turnover number suggests a regulatory, rather than a functional role. The BiP-associated ATPase shares several properties with the related cytoplasmic protein, HSC70/clathrin uncoating ATPase

File list: InP.Bld.50.AllAg.PBMC [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Bld.50.AllAg.PBMC hg19 Input control Blood PBMC SRX151954,SRX099233,SRX099232,S...RX095365,SRX037434 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Bld.50.AllAg.PBMC.bed ...

Melatonin suppresses thyroid cancer growth and overcomes radioresistance via inhibition of p65 phosphorylation and induction of ROS

Directory of Open Access Journals (Sweden)

Zhen-Wei Zou

2018-06-01

Full Text Available Thyroid cancer is the most common endocrine carcinoma with increasing incidence worldwide and anaplastic subtypes are frequently associated with cancer related death. Radioresistance of thyroid cancer often leads to therapy failure and cancer-related death. In this study, we found that melatonin showed potent suppressive roles on NF-κB signaling via inhibition of p65 phosphorylation and generated redox stress in thyroid cancer including the anaplastic subtypes. Our data showed that melatonin significantly decreased cell viability, suppressed cell migration and induced apoptosis in thyroid cancer cell lines in vitro and impaired tumor growth in the subcutaneous mouse model in vivo. By contrast, irradiation of thyroid cancer cells resulted in elevated level of phosphorylated p65, which could be reversed by cotreatment with melatonin. Consequently, melatonin synergized with irradiation to induce cytotoxicity to thyroid cancer, especially in the undifferentiated subgroups. Taken together, our results suggest that melatonin may exert anti-tumor activities against thyroid carcinoma by inhibition of p65 phosphorylation and induction of reactive oxygen species. Radio-sensitization by melatonin may have clinical benefits in thyroid cancer. Keywords: Melatonin, Thyroid cancer, Radioresistance, p65, Reactive oxygen species

p38gamma and p38delta mitogen activated protein kinases (MAPKs, new stars in the MAPK galaxy

Directory of Open Access Journals (Sweden)

Alejandra eEscós

2016-04-01

Full Text Available The protein kinases p38γ and p38δ belong to the p38 mitogen-activated protein kinase (MAPK family. p38MAPK signalling controls many cellular processes and is one of the most conserved mechanisms in eukaryotes for the cellular response to environmental stress and inflammation. Although p38γ and p38δ are widely expressed, it is likely that they perform specific functions in different tissues. Their involvement in human pathologies such as inflammation-related diseases or cancer is starting to be uncovered. In this article we give a general overview and highlight recent advances made in defining the functions of p38γ and p38δ, focusing in innate immunity and inflammation. We consider the potential of the pharmacological targeting of MAPK pathways to treat autoimmune and inflammatory diseases and cancer

The prognostic significance of accumulation of p53 protein in stage III non-small cell lung cancer treated by radiotherapy

International Nuclear Information System (INIS)

Langendijk, J.A.; Thunnissen, F.B.J.M.; Lamers, R.J.S.; Jong, J.M.A. de; Velde, G.P.M. ten; Wouters, E.F.M.

1995-01-01

In the present study the prognostic significance of accumulation of nuclear p53 protein on survival and freedom from local progression was investigated. Formalin-fixed, paraffin-embedded sections obtained by bronchoscopy or mediastinoscopy were used to examine the expression of nuclear p53 protein using immunohistochemistry. In 37 cases (57%), overexpression of the p53 protein was detected. No relation was found between p53 expression and other pretreatment variables. Response to radiotherapy was found in 11 p53-negative cases (65%) versus 10 p53-positive cases (42%). Freedom from local progression was significantly better in the p53-negative cases as compared with the p53-positive cases. The p53-negative cases who responded to radiotherapy showed an excellent freedom from local progression rate after 2 years of 100%, whereas all p53-positive cases without response to radiotherapy showed local progression within 24 months. Overall survival between p53-negative and -positive cases did not differ, however the disease-specific survival was found to be worse in the p53-positive cases as compared to the negative cases (median survival 8.4 vs. 14.4 months (P < 0.05)). No correlation was found between p53 expression and the frequency of distant metastases. In conclusion, the results of this study suggest that p53 protein expression may be of prognostic value on freedom from local progression in non-small cell lung carcinoma

File list: InP.Neu.50.AllAg.Neurons [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Neu.50.AllAg.Neurons mm9 Input control Neural Neurons SRX799007,SRX798994,SRX79...9006,SRX798993,SRX798995,SRX799005 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Neu.50.AllAg.Neurons.bed ...

File list: InP.Oth.50.AllAg.Fibrosarcoma [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Oth.50.AllAg.Fibrosarcoma hg19 Input control Others Fibrosarcoma SRX253236,SRX2...53238,SRX253237,SRX253233,SRX253235,SRX253234 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Oth.50.AllAg.Fibrosarcoma.bed ...

File list: InP.Neu.50.AllAg.Forebrain [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Neu.50.AllAg.Forebrain mm9 Input control Neural Forebrain SRX377679,SRX377675,S...RX377677,SRX377673,SRX669236,SRX377671 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Neu.50.AllAg.Forebrain.bed ...

File list: InP.Gon.50.AllAg.Spermatocytes [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Gon.50.AllAg.Spermatocytes mm9 Input control Gonad Spermatocytes SRX332362,SRX9...X381472,SRX381467,SRX709671,SRX997036,SRX099892 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Gon.50.AllAg.Spermatocytes.bed ...

File list: InP.Neu.50.AllAg.Hippocampus [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Neu.50.AllAg.Hippocampus mm9 Input control Neural Hippocampus SRX248470,SRX7693...89,SRX517454,SRX517457,SRX216313,SRX248471 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Neu.50.AllAg.Hippocampus.bed ...

1,5-Anhydro-D-fructose attenuates lipopolysaccharide-induced cytokine release via suppression of NF-κB p65 phosphorylation

International Nuclear Information System (INIS)

Meng Xiaojie; Kawahara, Ko-ichi; Nawa, Yuko; Miura, Naoki; Shrestha, Binita; Tancharoen, Salunya; Sameshima, Hisayo; Hashiguchi, Teruto; Maruyama, Ikuro

2009-01-01

Lipopolysaccharide (LPS) stimulates macrophages by activating NF-κB, which contributes to the release of tumor necrosis factor (TNF)-α and interleukin (IL)-6. 1,5-anhydro-D-fructose (1,5-AF), a monosaccharide formed from starch and glycogen, exhibits anti-oxidant activity and enhances insulin secretion. This study examined the effects of 1,5-AF on LPS-induced inflammatory reactions and elucidated its molecular mechanisms. Before LPS challenge, mice were pretreated with 1,5-AF (38.5 mg/kg). We found that 1,5-AF pretreatment attenuated cytokine release into the serum, including TNF-α, IL-6 and macrophage chemoattractant protein (MCP)-1. Furthermore, pretreatment with 1,5-AF (500 μg/ml) attenuated cytokine release, and 1,5-AF directly inhibited the nuclear translocalization of the NF-κB p65 subunit in LPS-stimulated murine macrophage-like RAW264.7 cells. This inhibition was responsible for decreased LPS-induced phosphorylation on Ser536 of the NF-κB p65 subunit, which is a posttranslational modification involved in the non-canonical pathway. Collectively, these findings indicate that the anti-inflammatory activity of 1,5-AF occurs via inactivation of NF-κB.

Evaluation of the pH- and Thermal Stability of the Recombinant Green Fluorescent Protein (GFP) in the Presence of Sodium Chloride

Science.gov (United States)

Ishii, Marina; Kunimura, Juliana Sayuri; Jeng, Hélio Tallon; Vessoni Penna, Thereza Christina; Cholewa, Olivia

The thermal stability of recombinant green fluorescent protein (GFP) in sodium chloride (NaCl) solutions at different concentrations, pH, and temperatures was evaluated by assaying the loss of fluorescence intensity as a measure of denaturation. GFP, extracted from Escherichia coli cells by the three-phase partitioning method and purified through a butyl hydrophobic interaction chromatography (HIC) column, was diluted in water for injection (WFI) (pH 6.0-7.0) and in 10 mM buffer solutions (acetate, pH 5.0; phosphate, pH 7.0; and Tris-EDTA, pH 8.0) with 0.9-30% NaCl or without and incubated at 80-95°C. The extent of protein denaturation was expressed as a percentage of the calculated decimal reduction time (D-value). In acetate buffer (pH 4.84 ±0.12), the mean D-values for 90% reduction in GFP fluorescence ranged from 2.3 to 3.6 min, independent of NaCl concentration and temperature. GFP thermal stability diluted in WFI (pH 5.94±0.60) was half that observed in phosphate buffer (pH 6.08±0.60); but in both systems, D-values decreased linearly with increasing NaCl concentration, with D-values (at 80°C) ranging from 3.44, min (WFI) to 6.1 min (phosphate buffer), both with 30% NaCl. However, D-values in Tris-EDTA (pH 7.65±0.17) were directly dependent on the NaCl concentration and 5-10 times higher than D-values for GFP in WFI at 80°C. GFP pH-and thermal stability can be easily monitored by the convenient measure of fluorescence intensity and potentially be used as an indicator to monitor that processing times and temperatures were attained.

Atomic structure of the sweet-tasting protein thaumatin I at pH 8.0 reveals the large disulfide-rich region in domain II to be sensitive to a pH change

Energy Technology Data Exchange (ETDEWEB)

Masuda, Tetsuya, E-mail: t2masuda@kais.kyoto-u.ac.jp [Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Gokasho, Uji, Kyoto 611-0011 (Japan); Department of Natural Resources, Graduate School of Global Environmental Studies, Kyoto University, Gokasho, Uji, Kyoto 611-0011 (Japan); Ohta, Keisuke [Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Gokasho, Uji, Kyoto 611-0011 (Japan); Department of Natural Resources, Graduate School of Global Environmental Studies, Kyoto University, Gokasho, Uji, Kyoto 611-0011 (Japan); Mikami, Bunzo [Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Gokasho, Uji, Kyoto 611-0011 (Japan); Kitabatake, Naofumi [Department of Foods and Human Nutrition, Notre Dame Seishin University, Okayama 700-8516 (Japan); Tani, Fumito [Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Gokasho, Uji, Kyoto 611-0011 (Japan); Department of Natural Resources, Graduate School of Global Environmental Studies, Kyoto University, Gokasho, Uji, Kyoto 611-0011 (Japan)

2012-03-02

Highlights: Black-Right-Pointing-Pointer Structure of a recombinant thaumatin at pH 8.0 determined at a resolution of 1.0 A. Black-Right-Pointing-Pointer Substantial fluctuations of a loop in domain II was found in the structure at pH 8.0. Black-Right-Pointing-Pointer B-factors for Lys137, Lys163, and Lys187 were significantly affected by pH change. Black-Right-Pointing-Pointer An increase in mobility might play an important role in the heat-induced aggregation. -- Abstract: Thaumatin, an intensely sweet-tasting plant protein, elicits a sweet taste at 50 nM. Although the sweetness remains when thaumatin is heated at 80 Degree-Sign C for 4 h under acid conditions, it rapidly declines when heating at a pH above 6.5. To clarify the structural difference at high pH, the atomic structure of a recombinant thaumatin I at pH 8.0 was determined at a resolution of 1.0 A. Comparison to the crystal structure of thaumatin at pH 7.3 and 7.0 revealed the root-mean square deviation value of a C{alpha} atom to be substantially greater in the large disulfide-rich region of domain II, especially residues 154-164, suggesting that a loop region in domain II to be affected by solvent conditions. Furthermore, B-factors of Lys137, Lys163, and Lys187 were significantly affected by pH change, suggesting that a striking increase in the mobility of these lysine residues, which could facilitate a reaction with a free sulfhydryl residue produced via the {beta}-elimination of disulfide bonds by heating at a pH above 7.0. The increase in mobility of lysine residues as well as a loop region in domain II might play an important role in the heat-induced aggregation of thaumatin above pH 7.0.

Gravity-driven pH adjustment for site-specific protein pKa measurement by solution-state NMR

Science.gov (United States)

Li, Wei

2017-12-01

To automate pH adjustment in site-specific protein pKa measurement by solution-state NMR, I present a funnel with two caps for the standard 5 mm NMR tube. The novelty of this simple-to-build and inexpensive apparatus is that it allows automatic gravity-driven pH adjustment within the magnet, and consequently results in a fully automated NMR-monitored pH titration without any hardware modification on the NMR spectrometer.

LocateP: Genome-scale subcellular-location predictor for bacterial proteins

Directory of Open Access Journals (Sweden)

Zhou Miaomiao

2008-03-01

Full Text Available Abstract Background In the past decades, various protein subcellular-location (SCL predictors have been developed. Most of these predictors, like TMHMM 2.0, SignalP 3.0, PrediSi and Phobius, aim at the identification of one or a few SCLs, whereas others such as CELLO and Psortb.v.2.0 aim at a broader classification. Although these tools and pipelines can achieve a high precision in the accurate prediction of signal peptides and transmembrane helices, they have a much lower accuracy when other sequence characteristics are concerned. For instance, it proved notoriously difficult to identify the fate of proteins carrying a putative type I signal peptidase (SPIase cleavage site, as many of those proteins are retained in the cell membrane as N-terminally anchored membrane proteins. Moreover, most of the SCL classifiers are based on the classification of the Swiss-Prot database and consequently inherited the inconsistency of that SCL classification. As accurate and detailed SCL prediction on a genome scale is highly desired by experimental researchers, we decided to construct a new SCL prediction pipeline: LocateP. Results LocateP combines many of the existing high-precision SCL identifiers with our own newly developed identifiers for specific SCLs. The LocateP pipeline was designed such that it mimics protein targeting and secretion processes. It distinguishes 7 different SCLs within Gram-positive bacteria: intracellular, multi-transmembrane, N-terminally membrane anchored, C-terminally membrane anchored, lipid-anchored, LPxTG-type cell-wall anchored, and secreted/released proteins. Moreover, it distinguishes pathways for Sec- or Tat-dependent secretion and alternative secretion of bacteriocin-like proteins. The pipeline was tested on data sets extracted from literature, including experimental proteomics studies. The tests showed that LocateP performs as well as, or even slightly better than other SCL predictors for some locations and outperforms

File list: InP.Neu.50.AllAg.Brain [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Neu.50.AllAg.Brain mm9 Input control Neural Brain SRX236086,ERX132888,ERX132890...ERX646161,ERX513118,ERX646137,ERX646145,ERX513117,SRX150261 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Neu.50.AllAg.Brain.bed ...

File list: InP.Adl.50.AllAg.Ovary [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Adl.50.AllAg.Ovary dm3 Input control Adult Ovary SRX264598,SRX1120694,SRX507393...215629,SRX507394,SRX215627,SRX215623,SRX215625,SRX507386 http://dbarchive.biosciencedbc.jp/kyushu-u/dm3/assembled/InP.Adl.50.AllAg.Ovary.bed ...

A Cross-Species Study of PI3K Protein-Protein Interactions Reveals the Direct Interaction of P85 and SHP2

Science.gov (United States)

Breitkopf, Susanne B.; Yang, Xuemei; Begley, Michael J.; Kulkarni, Meghana; Chiu, Yu-Hsin; Turke, Alexa B.; Lauriol, Jessica; Yuan, Min; Qi, Jie; Engelman, Jeffrey A.; Hong, Pengyu; Kontaridis, Maria I.; Cantley, Lewis C.; Perrimon, Norbert; Asara, John M.

2016-02-01

Using a series of immunoprecipitation (IP) - tandem mass spectrometry (LC-MS/MS) experiments and reciprocal BLAST, we conducted a fly-human cross-species comparison of the phosphoinositide-3-kinase (PI3K) interactome in a drosophila S2R+ cell line and several NSCLC and human multiple myeloma cell lines to identify conserved interacting proteins to PI3K, a critical signaling regulator of the AKT pathway. Using H929 human cancer cells and drosophila S2R+ cells, our data revealed an unexpected direct binding of Corkscrew, the drosophila ortholog of the non-receptor protein tyrosine phosphatase type II (SHP2) to the Pi3k21B (p60) regulatory subunit of PI3K (p50/p85 human ortholog) but no association with Pi3k92e, the human ortholog of the p110 catalytic subunit. The p85-SHP2 association was validated in human cell lines, and formed a ternary regulatory complex with GRB2-associated-binding protein 2 (GAB2). Validation experiments with knockdown of GAB2 and Far-Western blots proved the direct interaction of SHP2 with p85, independent of adaptor proteins and transfected FLAG-p85 provided evidence that SHP2 binding on p85 occurred on the SH2 domains. A disruption of the SHP2-p85 complex took place after insulin/IGF1 stimulation or imatinib treatment, suggesting that the direct SHP2-p85 interaction was both independent of AKT activation and positively regulates the ERK signaling pathway.

p38 mitogen-activated protein kinase (p38MAPK) upregulates catalase levels in response to low dose H2O2 treatment through enhancement of mRNA stability.

Science.gov (United States)

Sen, Prosenjit; Chakraborty, Prabir Kumar; Raha, Sanghamitra

2005-08-15

V79 fibroblasts were repetitively stressed through multiple exposures to a low dose (30 microM) H2O2 in culture for 4 weeks. Catalase activity, protein levels and mRNA levels increased markedly (5-6-fold) during this time and these augmentations were inhibited by the simultaneous presence of SB203580, an inhibitor of p38 mitogen-activated protein kinase (p38MAPK). p38MAPK became dually phosphorylated and ATF-2, a p38MAPK substrate also became increasingly phosphorylated over the repetitive stress period. Short interfering RNA that induced effective silencing of p38MAPK, was used to silence p38MAPK in V79 fibroblasts. Silencing of p38MAPK drastically hindered the elevation in catalase (protein and mRNA) levels observed after a single low dose (50 microM) of H2O. The rise in catalase mRNA levels induced by low concentration (single and multiple dose) H2O2 treatment was established to be unconnected with transcriptional upregulation but was brought forth primarily by an enhancement in catalase mRNA stability through the action of p38MAPK. Therefore, our data strongly indicate that activation of p38MAPK is a key controlling step in the upregulation of catalase levels by low dose H2O2 treatment.

Arrestin-related proteins mediate pH signaling in fungi

OpenAIRE

Herranz, Silvia; Rodríguez, José M.; Bussink, Henk-Jan; Sánchez-Ferrero, Juan C.; Arst, Herbert N.; Peñalva, Miguel A.; Vincent, Olivier

2005-01-01

Metazoan arrestins bind to seven-transmembrane (7TM) receptors to regulate function. Aspergillus nidulans PalF, a protein involved in the fungal ambient pH signaling pathway, contains arrestin N-terminal and C-terminal domains and binds strongly to two different regions within the C-terminal cytoplasmic tail of the 7TM, putative pH sensor PalH. Upon exposure to alkaline ambient pH, PalF is phosphorylated and, like mammalian β-arrestins, ubiquitinated in a signal-dependent and 7TM protein-depe...

Changes in protein expression in p53 deleted spontaneous thymic lymphomas

DEFF Research Database (Denmark)

Honoré, Bent; Vorum, Henrik; Pedersen, Anders Elm

2004-01-01

with the protein expression in p53+/+ and p53-/- thymocytes. Only a minority (13 proteins) of the quantitatively changed proteins were common for the two thymic lymphoma cell lines, suggesting that the p53 deficiency mainly results in genetic dysfunctions which are individual for a given tumor. Two of the detected...... structure containing motifs of the glyoxalase-bleomycin resistance protein family (MDR) as deduced from the cDNA....

Saccharomyces cerevisiae GTPase complex: Gtr1p-Gtr2p regulates cell-proliferation through Saccharomyces cerevisiae Ran-binding protein, Yrb2p

International Nuclear Information System (INIS)

Wang Yonggang; Nakashima, Nobutaka; Sekiguchi, Takeshi; Nishimoto, Takeharu

2005-01-01

A Gtr1p GTPase, the GDP mutant of which suppresses both temperature-sensitive mutants of Saccharomyces cerevisiae RanGEF/Prp20p and RanGAP/Rna1p, was presently found to interact with Yrb2p, the S. cerevisiae homologue of mammalian Ran-binding protein 3. Gtr1p bound the Ran-binding domain of Yrb2p. In contrast, Gtr2p, a partner of Gtr1p, did not bind Yrb2p, although it bound Gtr1p. A triple mutant: yrb2Δ gtr1Δ gtr2Δ was lethal, while a double mutant: gtr1Δ gtr2Δ survived well, indicating that Yrb2p protected cells from the killing effect of gtr1Δ gtr2Δ. Recombinant Gtr1p and Gtr2p were purified as a complex from Escherichia coli. The resulting Gtr1p-Gtr2p complex was comprised of an equal amount of Gtr1p and Gtr2p, which inhibited the Rna1p/Yrb2 dependent RanGAP activity. Thus, the Gtr1p-Gtr2p cycle was suggested to regulate the Ran cycle through Yrb2p

Emulsion properties of pork myofibrillar protein in combination with microbial transglutaminase and calcium alginate under various pH conditions.

Science.gov (United States)

Hong, Geun Pyo; Min, Sang-Gi; Chin, Koo Bok

2012-01-01

In this study, the effects of microbial transglutaminase (MTG) and calcium alginate (CA) systems in combination with soybean oil on the emulsion properties of porcine myofibrillar protein (MP) were evaluated under various pH conditions. MTG was shown to improve emulsifying capacity and creaming stability, which increased with increasing pH values up to 6.5. The CA did not influence emulsifying capacity, but it improved the creaming stability of the MP-stabilized emulsions. Both MTG and CA enhanced the rheological properties, but their effects on the physical characteristics of the protein evidenced an opposite trend in relation to pH, i.e., the MTG system improved both the emulsion and gelling properties with increasing pH, whereas the CA system was effective when the pH was lowered. By combining the two MP gelling systems, a stable and pH-insensible emulsion could be produced. Copyright © 2011 Elsevier Ltd. All rights reserved.

Effects of Extremely Low Frequency Electromagnetic Fields on Melanogenesis through p-ERK and p-SAPK/JNK Pathways in Human Melanocytes

Directory of Open Access Journals (Sweden)

Yu-Mi Kim

2017-10-01

Full Text Available This study evaluated frequency-dependent effects of extremely low frequency electromagnetic fields (ELF-EMFs on melanogenesis by melanocytes in vitro. Melanocytes were exposed to 2 mT EMFs at 30–75 Hz for 3 days before melanogenesis was examined. Exposure to ELF-EMFs at 50 and 60 Hz induced melanogenic maturation without cell damage, without changing cell proliferation and mitochondrial activity. Melanin content and tyrosinase activity of cells exposed to 50 Hz were higher than in controls, and mRNA expression of tyrosinase-related protein-2 was elevated relative to controls at 50 Hz. Phosphorylated cyclic adenosine monophosphate response element-binding protein (p-CREB levels were higher than controls in cells exposed to ELF-EMFs at 50–75 Hz. Immunohistochemical staining showed that melanocyte-specific markers (HMB45, Melan-A were strongly expressed in cells exposed to EMFs at 50 and 60 Hz compared to controls. Thus, exposure to ELF-EMFs at 50 Hz could stimulate melanogenesis in melanocytes, through activation of p-CREB and p-p38 and inhibition of phosphorylated extracellular signal-regulated protein kinase and phosphorylated stress-activated protein kinase/c-Jun N-terminal kinase. The results may form the basis of an appropriate anti-gray hair treatment or be applied in a therapeutic device for inducing repigmentation in the skin of vitiligo patients.

Transient degradation of NF-κB proteins in macrophages after interaction with mast cell granules

Directory of Open Access Journals (Sweden)

Noriko Ito

1998-01-01

Full Text Available The exposure of the macrophage cell line, J774 to mast cell granules (MCG led to the form ation of altered nuclear transcription factor proteins (NFκBx, which had faster electrophoretic mobility than the p50 homodimer of NF-κB, but retained comparable DNA binding capacity. Antibodies to N-terminal peptides of p50, p52, p65 or c-Rel supershifted only a fraction of NF-κBx. Western blot analyses revealed that nuclear p65 and c-Rel were progressively degraded after exposure to MCG, whereas nuclear p50 appeared to be unaffected. In contrast, cytoplasmic p50, p65, c-Rel as well as IkBα remained intact after MCG treatment, although p52 was clearly degraded. In comparison to J774 cells, incubation of m ouse peritoneal macrophages with MCG resulted in more extensive alterations to NF-κB proteins. The alterations in NF-κB proteins did not affect the expression of inducible nitric oxide synthase (iNOS or TNF-α mRNA in J774 cells. These data indicate that exposure of J774 cells to MCG leads to generation of altered nuclear p52, p65 and c-Rel, which retain intact N-terminal peptides, specific oligonucleotide binding and transactivating activity. On the other hand, in peritoneal macrophages, MCG induce more extensive modifications to NF-κB proteins with associated inhibition of iNOS or TNF-α mRNA expression.

File list: InP.Dig.50.AllAg.Intestines [Chip-atlas[Archive

Lifescience Database Archive (English)

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File list: InP.Prs.50.AllAg.Prostate [Chip-atlas[Archive

Lifescience Database Archive (English)

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File list: InP.Neu.50.AllAg.Cerebellum [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Neu.50.AllAg.Cerebellum mm9 Input control Neural Cerebellum SRX026435,SRX026436...24,SRX685923,SRX022872,SRX150265,SRX022867,SRX019017,SRX685922,SRX150263 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Neu.50.AllAg.Cerebellum.bed ...

In 0.35Ga 0.65P light-emitting diodes grown by gas-source MBE

Science.gov (United States)

Masselink, W. Ted; Zachau, Martin

1993-02-01

This paper describes the growth and optical characteristics of In yGa 1- yP with 0.3Raman spectra of these undoped films and observe strong excitonic luminescence over the entire composition range investigated. The band gap derived from the luminescence excitation spectra corresponds to that of a fully relaxed InGaP film with no residual strain, which is confirmed by the Raman measurements. Light-emitting diodes with peak (300 K) emission centered at less than 590 nm have been fabricated from p-i-n junctions in In 0.35Ga 0.65P. This alloy is close to that with the largest direct band gap in the In yGa 1- y P system and has lattice mismatch from the GaAs substrate of 1%.

A diagnostic model incorporating P50 sensory gating and neuropsychological tests for schizophrenia.

Directory of Open Access Journals (Sweden)

Jia-Chi Shan

Full Text Available OBJECTIVES: Endophenotypes in schizophrenia research is a contemporary approach to studying this heterogeneous mental illness, and several candidate neurophysiological markers (e.g. P50 sensory gating and neuropsychological tests (e.g. Continuous Performance Test (CPT and Wisconsin Card Sorting Test (WCST have been proposed. However, the clinical utility of a single marker appears to be limited. In the present study, we aimed to construct a diagnostic model incorporating P50 sensory gating with other neuropsychological tests in order to improve the clinical utility. METHODS: We recruited clinically stable outpatients meeting DSM-IV criteria of schizophrenia and age- and gender-matched healthy controls. Participants underwent P50 sensory gating experimental sessions and batteries of neuropsychological tests, including CPT, WCST and Wechsler Adult Intelligence Scale Third Edition (WAIS-III. RESULTS: A total of 106 schizophrenia patients and 74 healthy controls were enrolled. Compared with healthy controls, the patient group had significantly a larger S2 amplitude, and thus poorer P50 gating ratio (gating ratio = S2/S1. In addition, schizophrenia patients had a poorer performance on neuropsychological tests. We then developed a diagnostic model by using multivariable logistic regression analysis to differentiate patients from healthy controls. The final model included the following covariates: abnormal P50 gating (defined as P50 gating ratio >0.4, three subscales derived from the WAIS-III (Arithmetic, Block Design, and Performance IQ, sensitivity index from CPT and smoking status. This model had an adequate accuracy (concordant percentage = 90.4%; c-statistic = 0.904; Hosmer-Lemeshow Goodness-of-Fit Test, p = 0.64>0.05. CONCLUSION: To the best of our knowledge, this is the largest study to date using P50 sensory gating in subjects of Chinese ethnicity and the first to use P50 sensory gating along with other neuropsychological tests

On the development of protein pKa calculation algorithms

Science.gov (United States)

Carstensen, Tommy; Farrell, Damien; Huang, Yong; Baker, Nathan A.; Nielsen, Jens Erik

2011-01-01

Protein pKa calculation methods are developed partly to provide fast non-experimental estimates of the ionization constants of protein side chains. However, the most significant reason for developing such methods is that a good pKa calculation method is presumed to provide an accurate physical model of protein electrostatics, which can be applied in methods for drug design, protein design and other structure-based energy calculation methods. We explore the validity of this presumption by simulating the development of a pKa calculation method using artificial experimental data derived from a human-defined physical reality. We examine the ability of an RMSD-guided development protocol to retrieve the correct (artificial) physical reality and find that a rugged optimization landscape and a huge parameter space prevent the identification of the correct physical reality. We examine the importance of the training set in developing pKa calculation methods and investigate the effect of experimental noise on our ability to identify the correct physical reality, and find that both effects have a significant and detrimental impact on the physical reality of the optimal model identified. Our findings are of relevance to all structure-based methods for protein energy calculations and simulation, and have large implications for all types of current pKa calculation methods. Our analysis furthermore suggests that careful and extensive validation on many types of experimental data can go some way in making current models more realistic. PMID:21744393

Interactions involved in pH protection of the alphavirus fusion protein

Energy Technology Data Exchange (ETDEWEB)

Fields, Whitney; Kielian, Margaret, E-mail: margaret.kielian@einstein.yu.edu

2015-12-15

The alphavirus membrane protein E1 mediates low pH-triggered fusion of the viral and endosome membranes during virus entry. During virus biogenesis E1 associates as a heterodimer with the transmembrane protein p62. Late in the secretory pathway, cellular furin cleaves p62 to the mature E2 protein and a peripheral protein E3. E3 remains bound to E2 at low pH, stabilizing the heterodimer and thus protecting E1 from the acidic pH of the secretory pathway. Release of E3 at neutral pH then primes the virus for fusion during entry. Here we used site-directed mutagenesis and revertant analysis to define residues important for the interactions at the E3–E2 interface. Our data identified a key residue, E2 W235, which was required for E1 pH protection and alphavirus production. Our data also suggest additional residues on E3 and E2 that affect their interacting surfaces and thus influence the pH protection of E1 during alphavirus exit.

Characterization of the archaeal ribonuclease P proteins from Pyrococcus horikoshii OT3.

Science.gov (United States)

Terada, Atsushi; Honda, Takashi; Fukuhara, Hideo; Hada, Kazumasa; Kimura, Makoto

2006-08-01

Ribonuclease P (RNase P) is a ribonucleoprotein complex involved in the processing of the 5'-leader sequence of precursor tRNA (pre-tRNA). Our earlier study revealed that RNase P RNA (pRNA) and five proteins (PhoPop5, PhoRpp38, PhoRpp21, PhoRpp29, and PhoRpp30) in the hyperthermophilic archaeon Pyrococcus horikoshii OT3 reconstituted RNase P activity that exhibits enzymatic properties like those of the authentic enzyme. In present study, we investigated involvement of the individual proteins in RNase P activity. Two particles (R-3Ps), in which pRNA was mixed with three proteins, PhoPop5, PhoRpp30, and PhoRpp38 or PhoPop5, PhoRpp30, and PhoRpp21 showed a detectable RNase P activity, and five reconstituted particles (R-4Ps) composed of pRNA and four proteins exhibited RNase P activity, albeit at reduced level compared to that of the reconstituted particle (R-5P) composed of pRNA and five proteins. Time-course analysis of the RNase P activities of R-4Ps indicated that the R-4Ps lacking PhoPop5, PhoRpp21, or PhoRpp30 had virtually reduced activity, while omission of PhoRpp29 or PhoRpp38 had a slight effect on the activity. The results indicate that the proteins contribute to RNase P activity in order of PhoPop5 > PhoRpp30 > PhoRpp21 > PhoRpp29 > PhoRpp38. It was further found that R-4Ps showed a characteristic Mg2+ ion dependency approximately identical to that of R-5P. However, R-4Ps had optimum temperature of around at 55 degrees C which is lower than 70 degrees C for R-5P. Together, it is suggested that the P. horikoshii RNase P proteins are predominantly involved in optimization of the pRNA conformation, though they are individually dispensable for RNase P activity in vitro.

File list: InP.Emb.50.AllAg.Embryonic_palates [Chip-atlas[Archive

Lifescience Database Archive (English)

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File list: InP.Bld.50.AllAg.Polymorphonuclear_leukocytes [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Bld.50.AllAg.Polymorphonuclear_leukocytes hg19 Input control Blood Polymorphonuclear... leukocytes http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Bld.50.AllAg.Polymorphonuclear_leukocytes.bed ...

File list: InP.Adl.50.AllAg.Octopaminergic_neurons [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Adl.50.AllAg.Octopaminergic_neurons dm3 Input control Adult Octopaminergic neurons... http://dbarchive.biosciencedbc.jp/kyushu-u/dm3/assembled/InP.Adl.50.AllAg.Octopaminergic_neurons.bed ...

File list: InP.CDV.50.AllAg.Atrioventicular_canals [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.CDV.50.AllAg.Atrioventicular_canals mm9 Input control Cardiovascular Atrioventicular canals... http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.CDV.50.AllAg.Atrioventicular_canals.bed ...

File list: InP.Oth.50.AllAg.Pancreas_and_brain [Chip-atlas[Archive

Lifescience Database Archive (English)

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File list: InP.NoD.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.NoD.50.Input_control.AllCell dm3 Input control Input control No description htt...p://dbarchive.biosciencedbc.jp/kyushu-u/dm3/assembled/InP.NoD.50.Input_control.AllCell.bed ...

Electrophoretic detection of protein p53 in human leukocytes

International Nuclear Information System (INIS)

Paponov, V.D.; Kupsik, E.G.; Shcheglova, E.G.; Yarullin, N.N.

1986-01-01

The authors have found an acid-soluble protein with mol. wt. of about 53 kD in peripheral blood leukocytes of persons with Down's syndrome. It was present in different quantities in all 20 patients tested, but was virtually not discovered in 12 healthy blood donors. This paper determines the possible identity of this protein with protein p53 from mouse ascites carcinoma by comparing their electrophoretic mobilities, because the accuracy of electrophoretic determination of the molecular weight of proteins is not sufficient to identify them. The paper also describes experiments to detect a protein with electrophoretic mobility identical with that of a protein in the leukocytes of patients with Down's syndrome in leukocytes of patients with leukemia. To discover if protein p53 is involved in cell proliferation, the protein composition of leukocytes from healthy blood donors, cultured in the presence and absence of phytohemagglutinin (PHA), was compared. Increased incorporation of H 3-thymidine by leukocytes of patients with Down's syndrome is explained by the presence of a population of immature leukocytes actively synthesizing DNA in the peripheral blood of these patients, and this can also explain the presence of protein p53 in the leukocytes of these patients

File list: InP.CDV.50.AllAg.Endocardial_cells [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.CDV.50.AllAg.Endocardial_cells hg19 Input control Cardiovascular Endocardial ce...lls http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.CDV.50.AllAg.Endocardial_cells.bed ...

High CpG island methylation of p16 gene and loss of p16 protein ...

Indian Academy of Sciences (India)

SI-JU GAO

The study subjects consisted of 75 healthy controls and 63 ToF ... Additionally, our analysis suggested that CpG island methylation in p16 promoters in ToF ..... reduced p16 protein expression in lung cancer (Kondo et al. 2006). In this context ..... promoter methylation in gastric carcinogenesis: a meta-analysis. Mol. Biol. Rep.

File list: InP.Dig.50.AllAg.Intestinal_adenoma [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Dig.50.AllAg.Intestinal_adenoma mm9 Input control Digestive tract Intestinal ad...enoma http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Dig.50.AllAg.Intestinal_adenoma.bed ...

File list: InP.Epd.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Epd.50.Input_control.AllCell mm9 Input control Input control Epidermis SRX14260...tp://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Epd.50.Input_control.AllCell.bed ...

File list: InP.Emb.50.AllAg.Embryonic_flank [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Emb.50.AllAg.Embryonic_flank mm9 Input control Embryo Embryonic flank SRX804059... http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Emb.50.AllAg.Embryonic_flank.bed ...

File list: InP.Emb.50.AllAg.Embryonic_pancreas [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Emb.50.AllAg.Embryonic_pancreas mm9 Input control Embryo Embryonic pancreas SRX...287026 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Emb.50.AllAg.Embryonic_pancreas.bed ...

Predicting Secretory Proteins with SignalP

DEFF Research Database (Denmark)

Nielsen, Henrik

2017-01-01

SignalP is the currently most widely used program for prediction of signal peptides from amino acid sequences. Proteins with signal peptides are targeted to the secretory pathway, but are not necessarily secreted. After a brief introduction to the biology of signal peptides and the history...

Non-structural proteins P17 and P33 are involved in the assembly of the internal membrane-containing virus PRD1

Energy Technology Data Exchange (ETDEWEB)

Karttunen, Jenni; Mäntynen, Sari [Centre of Excellence in Biological Interactions, Department of Biological and Environmental Science and Nanoscience Center, University of Jyväskylä, P.O. Box 35, 40014 Jyväskylä (Finland); Ihalainen, Teemu O. [Stem Cells in Neurological Applications Group, BioMediTech, University of Tampere, Tampere (Finland); Bamford, Jaana K.H. [Centre of Excellence in Biological Interactions, Department of Biological and Environmental Science and Nanoscience Center, University of Jyväskylä, P.O. Box 35, 40014 Jyväskylä (Finland); Oksanen, Hanna M., E-mail: hanna.oksanen@helsinki.fi [Institute of Biotechnology and Department of Biosciences, University of Helsinki, Biocenter 2, P.O. Box 56 (Viikinkaari 5), FIN-00014 Helsinki (Finland)

2015-08-15

Bacteriophage PRD1, which has been studied intensively at the structural and functional levels, still has some gene products with unknown functions and certain aspects of the PRD1 assembly process have remained unsolved. In this study, we demonstrate that the phage-encoded non-structural proteins P17 and P33, either individually or together, complement the defect in a temperature-sensitive GroES mutant of Escherichia coli for host growth and PRD1 propagation. Confocal microscopy of fluorescent fusion proteins revealed co-localisation between P33 and P17 as well as between P33 and the host chaperonin GroEL. A fluorescence recovery after photobleaching assay demonstrated that the diffusion of the P33 fluorescent fusion protein was substantially slower in E. coli than theoretically calculated, presumably resulting from intermolecular interactions. Our results indicate that P33 and P17 function in procapsid assembly, possibly in association with the host chaperonin complex GroEL/GroES. - Highlights: • Two non-structural proteins of PRD1 are involved in the virus assembly. • P17 and P33 complement the defect in GroES of Escherichia coli. • P33 co-localises with GroEL and P17 in the bacterium. • Slow motion of P33 in the bacterium suggests association with cellular components.

File list: InP.Emb.50.AllAg.Neuronal_cells [Chip-atlas[Archive

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File list: InP.Emb.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Emb.50.Input_control.AllCell dm3 Input control Input control Embryo SRX681824,S...http://dbarchive.biosciencedbc.jp/kyushu-u/dm3/assembled/InP.Emb.50.Input_control.AllCell.bed ...

File list: InP.Liv.50.AllAg.Carcinoma,_Hepatocellular [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Liv.50.AllAg.Carcinoma,_Hepatocellular mm9 Input control Liver Carcinoma, Hepat...ocellular SRX467208 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Liv.50.AllAg.Carcinoma,_Hepatocellular.bed ...

File list: InP.ALL.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

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Characterization of fatty acid binding by the P2 myelin protein

International Nuclear Information System (INIS)

Gudaitis, P.G.; Weise, M.J.

1987-01-01

In recent years, significant sequence homology has been found between the P2 protein of peripheral myelin and intracellular retinoid- and fatty acid-binding proteins. They have found that salt extracts of bovine intradural nerve roots contain the P2 basic protein in association with free fatty acid. Preliminary results from quantitative analyses showed a ratio of 0.4-1.1 fatty acid (mainly oleate and palmitate) per P2 molecule. P2/ligand interactions were partially characterized using ( 3 H)-oleate in gel permeation assays and binding studies using lipidex to separated bound and free fatty acid. Methyloleate was found to displace ( 3 H)-oleate from P2, indicating that ligand binding interactions are predominantly hydrophobic in nature. On the other hand, myristic acid and retinol did not inhibit the binding of oleate to the protein, results consistent with a decided affinity for long chain fatty acids but not for the retinoids. The binding between P2 and oleic acid showed an apparent Kd in the micromolar range, a value comparable to those found for other fatty acid-binding proteins. From these results they conclude that P2 shares not only structural homology with certain fatty acid binding proteins but also an ability to bind long chain fatty acids. Although the significance of these similarities is not yet clear, they may, by analogy, expect P2 to have a role in PNS lipid metabolism

Stability of the Human Hsp90-p50Cdc37 Chaperone Complex against Nucleotides and Hsp90 Inhibitors, and the Influence of Phosphorylation by Casein Kinase 2

Directory of Open Access Journals (Sweden)

Sanne H. Olesen

2015-01-01

Full Text Available The molecular chaperone Hsp90 is regulated by co-chaperones such as p50Cdc37, which recruits a wide selection of client protein kinases. Targeted disruption of the Hsp90-p50Cdc37 complex by protein–protein interaction (PPI inhibitors has emerged as an alternative strategy to treat diseases characterized by aberrant Hsp90 activity. Using isothermal microcalorimetry, ELISA and GST-pull down assays we evaluated reported Hsp90 inhibitors and nucleotides for their ability to inhibit formation of the human Hsp90β-p50Cdc37 complex, reconstituted in vitro from full-length proteins. Hsp90 inhibitors, including the proposed PPI inhibitors gedunin and H2-gamendazole, did not affect the interaction of Hsp90 with p50Cdc37 in vitro. Phosphorylation of Hsp90 and p50Cdc37 by casein kinase 2 (CK2 did not alter the thermodynamic signature of complex formation. However, the phosphorylated complex was vulnerable to disruption by ADP (IC50 = 32 µM, while ATP, AMPPNP and Hsp90 inhibitors remained largely ineffective. The differential inhibitory activity of ADP suggests that phosphorylation by CK2 primes the complex for dissociation in response to a drop in ATP/ADP levels. The approach applied herein provides robust assays for a comprehensive biochemical evaluation of potential effectors of the Hsp90-p50Cdc37 complex, such as phosphorylation by a kinase or the interaction with small molecule ligands.

Characterization of protein kinase CK2 protein subunits and p53 in F9 teratocarcinoma cells in the absence and presence of cisplatin

DEFF Research Database (Denmark)

Küpper, M; Köster, M; Schmidt-Spaniol, I

1994-01-01

cell extracts treated with and without cisplatin were analyzed by ion exchange chromatography for protein kinase CK2 alpha/beta subunits and p53 distribution. The following results were obtained: (a) in crude extracts of cisplatin-treated cells, CK2 activity was sometimes reduced by as much as 50%; (b......The effect of cis-diaminedichloroplatinum(II) (cisplatin) on the induction of p53 and protein kinase CK2 activity was studied in the mouse teratocarcinoma cell line F9. Treatment of the cells with the chemotherapeutic agent cisplatin led to the detection of p53 3 h after addition of the drug. F9...... by immunostaining, we have detected, at a concentration of approximately 200 mM NaCl, a protein of approximately 46 kDa which reacted with the CK2 alpha-specific antibody. This fraction was devoid of CK2 activity; and (d) cisplatin-treated cells exhibited p53 protein, which was mostly eluting ahead but also partly...

File list: InP.Bld.50.AllAg.Lymphoid_cells [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Bld.50.AllAg.Lymphoid_cells mm9 Input control Blood Lymphoid cells SRX021894,SR...X971601 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Bld.50.AllAg.Lymphoid_cells.bed ...

File list: InP.Bld.50.AllAg.Plasma_Cells [Chip-atlas[Archive

Lifescience Database Archive (English)

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File list: InP.Emb.50.AllAg.embryonic_skin [Chip-atlas[Archive

Lifescience Database Archive (English)

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File list: InP.Lar.50.AllAg.Larval_brain [Chip-atlas[Archive

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File list: InP.Plc.50.Input_control.AllCell [Chip-atlas[Archive

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File list: InP.YSt.50.Input_control.AllCell [Chip-atlas[Archive

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File list: InP.ALL.50.Input_control.AllCell [Chip-atlas[Archive

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Full Text Available InP.ALL.50.Input_control.AllCell dm3 Input control Input control All cell types SRX...948 http://dbarchive.biosciencedbc.jp/kyushu-u/dm3/assembled/InP.ALL.50.Input_control.AllCell.bed ...

File list: InP.Emb.50.AllAg.Embryonic_eye [Chip-atlas[Archive

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Full Text Available InP.Emb.50.AllAg.Embryonic_eye mm9 Input control Embryo Embryonic eye SRX804057,SRX...804055 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Emb.50.AllAg.Embryonic_eye.bed ...

File list: InP.Bld.50.AllAg.Kidney_Cortex [Chip-atlas[Archive

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Full Text Available InP.Bld.50.AllAg.Kidney_Cortex hg19 Input control Blood Kidney Cortex SRX1318623,SR...X1318622 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Bld.50.AllAg.Kidney_Cortex.bed ...

Mice Deficient in NF-κB p50 and p52 or RANK Have Defective Growth Plate Formation and Post-natal Dwarfism.

Science.gov (United States)

Xing, Lianping; Chen, Di; Boyce, Brendan F

2013-12-01

NF-κBp50/p52 double knockout (dKO) and RANK KO mice have no osteoclasts and develop severe osteopetrosis associated with dwarfism. In contrast, Op/Op mice, which form few osteoclasts, and Src KO mice, which have osteoclasts with defective resorptive function, are osteopetrotic, but they are not dwarfed. Here, we compared the morphologic features of long bones from p50/p52 dKO, RANK KO, Op/Op and Src KO mice to attempt to explain the differences in their long bone lengths. We found that growth plates in p50/p52 dKO and RANK KO mice are significantly thicker than those in WT mice due to a 2-3-fold increase in the hypertrophic chondrocyte zone associated with normal a proliferative chondrocyte zone. This growth plate abnormality disappears when animals become older, but their dwarfism persists. Op/Op or Src KO mice have relatively normal growth plate morphology. In-situ hybridization study of long bones from p50/p52 dKO mice showed marked thickening of the growth plate region containing type 10 collagen-expressing chondrocytes. Treatment of micro-mass chondrocyte cultures with RANKL did not affect expression levels of type 2 collagen and Sox9, markers for proliferative chondrocytes, but RANKL reduced the number of type 10 collagen-expressing hypertrophic chondrocytes. Thus, RANK/NF-κB signaling plays a regulatory role in post-natal endochondral ossification that maintains hypertrophic conversion and prevents dwarfism in normal mice.

Diagnostic significance of pleural fluid pH and pCO2

Directory of Open Access Journals (Sweden)

K.E. Sobhey

2015-10-01

Results: We conducted this study on 50 patients with pleural effusions of different causes. The patients were classified into 5 groups according to the cause. For all the patients, measurement of pleural pH, pCO2, pO2, HCO3, protein, LDH, glucose and WBC was done. We observed lowest pH in complicated parapneumonic effusion (empyema 6.80 ± 0.15 and highest pH was observed in transudative effusion 7.47 ± 0.07. Tuberculous effusion has pH lower than pH of malignant effusion 7.17 ± 0.017 and 7.39 ± 0.08, respectively. Post pleurodesis malignant effusion has pH lower than pH of malignant effusion 7.28 ± 0.17 and 7.39 ± 0.08, respectively. There is a strong inverse correlation between pH and pCO2, WBC, LDH and protein (r = −0.813 and p < 0.001, (r = −0.796 and p, 0.001, (r = −0.829 and p, 0.001 and (r = −.837 and p, 0.001, respectively. While there is a weak correlation between pH and glucose of pleural fluid (r = 0.249 and p = 0.066. The highest increase of PNL numbers was in empyema (20169 ± 8094.8 cells/cc.The highest increase of lymphocytes was in malignant effusions (4285.00 ± 2948.20 cells/cc and tuberculous effusion (3977.7 ± 3169 cells/cc.

Preliminary crystallographic studies of yeast mitochondrial peripheral membrane protein Tim44p

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Josyula, Ratnakar [Department of Cell Biology, Center for Biophysical Sciences and Engineering, University of Alabama at Birmingham (United States); Jin, Zhongmin [SER-CAT, APS, Argonne National Laboratory, 9700 South Cass Avenue, Argonne, IL 60439 (United States); McCombs, Deborah; DeLucas, Lawrence [Center for Biophysical Sciences and Engineering, University of Alabama at Birmingham (United States); Sha, Bingdong, E-mail: bdsha@uab.edu [Department of Cell Biology, Center for Biophysical Sciences and Engineering, University of Alabama at Birmingham (United States)

2006-02-01

Tim44p is an essential mitochondrial peripheral membrane protein. To investigate the mechanism by which Tim44p functions in the TIM23 translocon to deliver the mitochondrial protein precursors, the yeast Tim44p has been crystallized. Protein translocations across mitochondrial membranes play critical roles in mitochondrion biogenesis. Protein transport from the cell cytosol to the mitochondrial matrix is carried out by the translocase of the outer membrane (TOM) complex and the translocase of the inner membrane (TIM) complexes. Tim44p is an essential mitochondrial peripheral membrane protein and a major component of the TIM23 translocon. To investigate the mechanism by which Tim44p functions in the TIM23 translocon to deliver the mitochondrial protein precursors, the yeast Tim44p was crystallized. The crystals diffract to 3.2 Å using a synchrotron X-ray source and belong to space group P6{sub 3}22, with unit-cell parameters a = 124.25, c = 77.83 Å. There is one Tim44p molecule in one asymmetric unit, which corresponds to a solvent content of approximately 43%. Structure determination by MAD methods is under way.

File list: InP.Neu.50.AllAg.Cortical_neuron [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Neu.50.AllAg.Cortical_neuron mm9 Input control Neural Cortical neuron SRX804268...76,SRX671651,SRX671655 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Neu.50.AllAg.Cortical_neuron.bed ...

File list: InP.Myo.50.Input_control.AllCell [Chip-atlas[Archive

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Full Text Available InP.Myo.50.Input_control.AllCell mm9 Input control Input control Muscle SRX262224,S...62124,SRX022849 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Myo.50.Input_control.AllCell.bed ...

File list: InP.ALL.50.Input_control.AllCell [Chip-atlas[Archive

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Full Text Available InP.ALL.50.Input_control.AllCell mm9 Input control Input control All cell types SRX...7314,ERX807291 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.ALL.50.Input_control.AllCell.bed ...

File list: InP.Bld.50.AllAg.CCRF-CEM [Chip-atlas[Archive

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Full Text Available InP.Bld.50.AllAg.CCRF-CEM hg19 Input control Blood CCRF-CEM SRX107305,SRX107290,SRX...728791 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Bld.50.AllAg.CCRF-CEM.bed ...

File list: InP.Adl.50.AllAg.Egg_chamber [Chip-atlas[Archive

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Full Text Available InP.Adl.50.AllAg.Egg_chamber dm3 Input control Adult Egg chamber SRX706812,SRX70681...6,SRX706813 http://dbarchive.biosciencedbc.jp/kyushu-u/dm3/assembled/InP.Adl.50.AllAg.Egg_chamber.bed ...

File list: InP.Prs.50.AllAg.Prostate_cancer [Chip-atlas[Archive

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Full Text Available InP.Prs.50.AllAg.Prostate_cancer hg19 Input control Prostate Prostate cancer SRX861...688,SRX861686,SRX861689 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Prs.50.AllAg.Prostate_cancer.bed ...

File list: InP.Bld.50.Input_control.AllCell [Chip-atlas[Archive

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Full Text Available InP.Bld.50.Input_control.AllCell mm9 Input control Input control Blood SRX1287947,S...,SRX120230,SRX021428 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Bld.50.Input_control.AllCell.bed ...

File list: InP.CDV.50.AllAg.Carotid_Arteries [Chip-atlas[Archive

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Full Text Available InP.CDV.50.AllAg.Carotid_Arteries hg19 Input control Cardiovascular Carotid Arterie...s DRX021452,DRX021453,DRX021454 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.CDV.50.AllAg.Carotid_Arteries.bed ...

Vorinostat enhances protein stability of p27 and p21 through negative regulation of Skp2 and Cks1 in human breast cancer cells.

Science.gov (United States)

Uehara, Norihisa; Yoshizawa, Katsuhiko; Tsubura, Airo

2012-07-01

Vorinostat is a histone deacetylase inhibitor that blocks cancer cell proliferation through the regulation of cyclin-dependent kinase inhibitors. We, herein, examined the involvement of S-phase kinase-associated protein 2 (Skp2) and cyclin-dependent kinase subunit 1 (Cks1), the components of the SCFSkp2-Cks1 (Skp1/Cul1/F-box protein) ubiquitin ligase complex, in the regulation of p27 and p21 during vorinostat-induced growth arrest of MDA-MB-231 and MCF-7 human breast cancer cells. Vorinostat significantly reduced BrdU incorporation in MDA-MB-231 and MCF-7 cells, which was associated with increased p27 and p21 protein levels without concomitant induction of p27 mRNA. Vorinostat-induced accumulation of p27 and p21 proteins was inversely correlated with the mRNA and protein levels of Skp2 and Cks1. Cycloheximide chase analysis revealed that vorinostat increased the half-life of p27 and p21 proteins. The accumulation of p27 and p21 proteins was attenuated by forced expression of Skp2 and Cks1, which conferred resistance to the vorinostat-induced S-phase reduction. These results suggest that vorinostat-induced growth arrest may be in part due to the enhanced protein stability of p27 and p21 through the downregulation of Skp2 and Cks1.

P-proteins in Arabidopsis are heteromeric structures involved in rapid sieve tube sealing.

Science.gov (United States)

Jekat, Stephan B; Ernst, Antonia M; von Bohl, Andreas; Zielonka, Sascia; Twyman, Richard M; Noll, Gundula A; Prüfer, Dirk

2013-01-01

Structural phloem proteins (P-proteins) are characteristic components of the sieve elements in all dicotyledonous and many monocotyledonous angiosperms. Tobacco P-proteins were recently confirmed to be encoded by the widespread sieve element occlusion (SEO) gene family, and tobacco SEO proteins were shown to be directly involved in sieve tube sealing thus preventing the loss of photosynthate. Analysis of the two Arabidopsis SEO proteins (AtSEOa and AtSEOb) indicated that the corresponding P-protein subunits do not act in a redundant manner. However, there are still pending questions regarding the interaction properties and specific functions of AtSEOa and AtSEOb as well as the general function of structural P-proteins in Arabidopsis. In this study, we characterized the Arabidopsis P-proteins in more detail. We used in planta bimolecular fluorescence complementation assays to confirm the predicted heteromeric interactions between AtSEOa and AtSEOb. Arabidopsis mutants depleted for one or both AtSEO proteins lacked the typical P-protein structures normally found in sieve elements, underlining the identity of AtSEO proteins as P-proteins and furthermore providing the means to determine the role of Arabidopsis P-proteins in sieve tube sealing. We therefore developed an assay based on phloem exudation. Mutants with reduced AtSEO expression levels lost twice as much photosynthate following injury as comparable wild-type plants, confirming that Arabidopsis P-proteins are indeed involved in sieve tube sealing.

P-proteins in Arabidopsis are heteromeric structures involved in rapid sieve tube sealing

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Stephan B Jekat

2013-07-01

Full Text Available Structural phloem proteins (P-proteins are characteristic components of the sieve elements in all dicotyledonous and many monocotyledonous angiosperms. Tobacco P-proteins were recently evidenced to be encoded by the widespread SEO gene family, and tobacco SEO proteins were shown to be directly involved in sieve tube sealing thus preventing the loss of photosynthate. Analysis of the two Arabidopsis SEO proteins (AtSEOa and AtSEOb indicated that the corresponding P-protein subunits do not act in a redundant manner. However, there are still pending questions regarding the interaction properties and specific functions of AtSEOa and AtSEOb as well as the general function of structural P-proteins in Arabidopsis. In this study, we characterized the Arabidopsis P-proteins in more detail. We used in planta bimolecular fluorescence complementation assays to confirm the predicted heteromeric interactions between AtSEOa and AtSEOb. Arabidopsis mutants depleted for one or both AtSEO proteins lacked the typical P-protein structures normally found in sieve elements, underlining the identity of AtSEO proteins as P-proteins and furthermore providing the means to determine the role of Arabidopsis P-proteins in sieve tube sealing. We therefore developed an assay based on phloem exudation. Mutants with reduced AtSEO expression levels lost twice as much photosynthate following injury as comparable wild-type plants, confirming that Arabidopsis P-proteins are indeed involved in sieve tube sealing. 

Preliminary X-ray crystallographic studies of yeast mitochondrial protein Tom70p

Energy Technology Data Exchange (ETDEWEB)

Wu, Yunkun [Department of Cell Biology, Center for Biophysical Sciences and Engineering, University of Alabama at Birmingham (United States); McCombs, Debbie; Nagy, Lisa; DeLucas, Lawrence [Center for Biophysical Sciences and Engineering, University of Alabama at Birmingham (United States); Sha, Bingdong, E-mail: bdsha@uab.edu [Department of Cell Biology, Center for Biophysical Sciences and Engineering, University of Alabama at Birmingham (United States)

2006-03-01

Tom70p is an important translocase of the outer membrane complex member and a major surface receptor of the protein-translocation machinery in the outer mitochondrial membrane. To investigate the mechanism by which Tom70p functions to deliver the mitochondrial protein precursors, the cytosolic fragment of yeast Tom70p (cTom70p) has been crystallized. Protein translocations across mitochondrial membranes play critical roles in mitochondrion biogenesis. Protein transport from the cell cytosol to the mitochondrial matrix is carried out by the translocase of the outer membrane (TOM) complex and the translocase of the inner membrane (TIM) complexes. Tom70p is an important TOM-complex member and a major surface receptor of the protein-translocation machinery in the outer mitochondrial membrane. To investigate the mechanism by which Tom70p functions to deliver the mitochondrial protein precursors, the cytosolic fragment of yeast Tom70p (cTom70p) was crystallized. The crystals diffract to 3.2 Å using a synchrotron X-ray source and belong to space group P2{sub 1}, with unit-cell parameters a = 44.89, b = 168.78, c = 83.41 Å, α = 90.00, β = 102.74, γ = 90.00°. There are two Tom70p molecules in one asymmetric unit, which corresponds to a solvent content of approximately 51%. Structure determination by MAD methods is under way.

Sacarose e pH na germinação in vitro de grãos de pólen de citros Sucrose and pH on in vitro pollen germination of citros

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Leila Aparecida Salles

2006-02-01

Full Text Available Objetivou-se avaliar o efeito da sacarose e pH na germinação in vitro de grãos de pólen das cultivares Valência, Natal e Pêra. Para testar o efeito da sacarose, os grãos de pólen foram distribuídos uniformemente em placas de Petri contendo meio de cultura básico constituído de 10 gL-1 de ágar, 800 mgL-1 de nitrato de cálcio e 200 mgL-1 de ácido bórico, acrescido de sacarose (0, 50, 100, 150 e 200 gL-1. Para verificação do pH satisfatório, os grãos de pólen foram inoculados em meio de cultura contendo 10 gL-1 de ágar e 800 mgL-1 de nitrato de cálcio, 200 mgL-1 de ácido bórico,100 gL-1 de sacarose e pH de 3,5; 4,0; 4,5; 5,0; 5,5; 6,0; 6,5. Após inoculação, os grãos de pólen foram incubados em BOD a 25ºC por 12 horas. A porcentagem de germinação foi obtida com auxílio de microscópio óptico com objetiva de 10 X. Para todas as cultivares estudadas, a maior porcentagem de germinação foi obtida com 100 gL-1 de sacarose e o maior número de grãos de pólen germinados foi verificado em pH 6,5, sendo observado que maiores valores de pH aumentaram também a quantidade de polens estourados para as cultivares Natal e Pêra e diminuíram para Valência.This work was carried out in order to evaluate the effect of sucrose and pH on in vitro pollen germination of cultivars Valencia, Natal and Pera (Citrus sinensis Osbeck. Pollens were distributed in Petri dishes containing culture medium with 10 gL-1 of agar 800 mgL-1 calcium nitrate, 200 mgL-1 boric acid, added of sucrose (0, 50, 100, 150 and 200 gL-1 In order to verify the satisfactory pH, the pollen were inoculated in culture middle containing 10 agar gL-1 and 800 mgL-1 calcium nitrate, 200 mgL-1 acid bórico,100 sucrose gL-1 and pH values of 3,5; 4,0; 4,5; 5,0; 5,5; 6,0; 6,5. After inoculation, pollen were incubated at 25ºC for 12 hours B.O.D. The largest germination percentage was obtained with an aid of optical microscope with lens of 10 X. For all the studied

File list: InP.Kid.50.AllAg.Nephrectomy_sample [Chip-atlas[Archive

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Full Text Available InP.Kid.50.AllAg.Nephrectomy_sample hg19 Input control Kidney Nephrectomy sample SR...84,SRX1037589,SRX1037590,SRX1037583 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Kid.50.AllAg.Nephrectomy_sample.bed ...

File list: InP.Dig.50.AllAg.Colon_cancer [Chip-atlas[Archive

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Full Text Available InP.Dig.50.AllAg.Colon_cancer hg19 Input control Digestive tract Colon cancer SRX12...155774,SRX124693,SRX124698 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Dig.50.AllAg.Colon_cancer.bed ...

File list: InP.Emb.50.Input_control.AllCell [Chip-atlas[Archive

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File list: InP.Brs.50.AllAg.Mammary_cells [Chip-atlas[Archive

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File list: InP.Gon.50.Input_control.AllCell [Chip-atlas[Archive

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File list: InP.Utr.50.Input_control.AllCell [Chip-atlas[Archive

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File list: InP.Dig.50.Input_control.AllCell [Chip-atlas[Archive

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File list: InP.Emb.50.AllAg.Embryonic_limb [Chip-atlas[Archive

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Andrographolide inhibits nuclear factor-κB activation through JNK-Akt-p65 signaling cascade in tumor necrosis factor-α-stimulated vascular smooth muscle cells.

Science.gov (United States)

Chen, Yu-Ying; Hsu, Ming-Jen; Hsieh, Cheng-Ying; Lee, Lin-Wen; Chen, Zhih-Cherng; Sheu, Joen-Rong

2014-01-01

Critical vascular inflammation leads to vascular dysfunction and cardiovascular diseases, including abdominal aortic aneurysms, hypertension, and atherosclerosis. Andrographolide is the most active and critical constituent isolated from the leaves of Andrographis paniculata, a herbal medicine widely used for treating anti-inflammation in Asia. In this study, we investigated the mechanisms of the inhibitory effects of andrographolide in vascular smooth muscle cells (VSMCs) exposed to a proinflammatory stimulus, tumor necrosis factor-α (TNF-α). Treating TNF-α-stimulated VSMCs with andrographolide suppressed the expression of inducible nitric oxide synthase in a concentration-dependent manner. A reduction in TNF-α-induced c-Jun N-terminal kinase (JNK), Akt, and p65 phosphorylation was observed in andrographolide-treated VSMCs. However, andrographolide affected neither IκBα degradation nor p38 mitogen-activated protein kinase or extracellular signal-regulated kinase 1/2 phosphorylation under these conditions. Both treatment with LY294002, a phosphatidylinositol 3-kinase/Akt inhibitor, and treatment with SP600125, a JNK inhibitor, markedly reversed the andrographolide-mediated inhibition of p65 phosphorylation. In addition, LY294002 and SP600125 both diminished Akt phosphorylation, whereas LY294002 had no effects on JNK phosphorylation. These results collectively suggest that therapeutic interventions using andrographolide can benefit the treatment of vascular inflammatory diseases, and andrographolide-mediated inhibition of NF-κB activity in TNF-α-stimulated VSMCs occurs through the JNK-Akt-p65 signaling cascade, an IκBα-independent mechanism.

Andrographolide Inhibits Nuclear Factor-κB Activation through JNK-Akt-p65 Signaling Cascade in Tumor Necrosis Factor-α-Stimulated Vascular Smooth Muscle Cells

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Yu-Ying Chen

2014-01-01

Full Text Available Critical vascular inflammation leads to vascular dysfunction and cardiovascular diseases, including abdominal aortic aneurysms, hypertension, and atherosclerosis. Andrographolide is the most active and critical constituent isolated from the leaves of Andrographis paniculata, a herbal medicine widely used for treating anti-inflammation in Asia. In this study, we investigated the mechanisms of the inhibitory effects of andrographolide in vascular smooth muscle cells (VSMCs exposed to a proinflammatory stimulus, tumor necrosis factor-α (TNF-α. Treating TNF-α-stimulated VSMCs with andrographolide suppressed the expression of inducible nitric oxide synthase in a concentration-dependent manner. A reduction in TNF-α-induced c-Jun N-terminal kinase (JNK, Akt, and p65 phosphorylation was observed in andrographolide-treated VSMCs. However, andrographolide affected neither IκBα degradation nor p38 mitogen-activated protein kinase or extracellular signal-regulated kinase 1/2 phosphorylation under these conditions. Both treatment with LY294002, a phosphatidylinositol 3-kinase/Akt inhibitor, and treatment with SP600125, a JNK inhibitor, markedly reversed the andrographolide-mediated inhibition of p65 phosphorylation. In addition, LY294002 and SP600125 both diminished Akt phosphorylation, whereas LY294002 had no effects on JNK phosphorylation. These results collectively suggest that therapeutic interventions using andrographolide can benefit the treatment of vascular inflammatory diseases, and andrographolide-mediated inhibition of NF-κB activity in TNF-α-stimulated VSMCs occurs through the JNK-Akt-p65 signaling cascade, an IκBα-independent mechanism.

Phosphorylation and activation of p42 and p44 mitogen-activated protein kinase are required for the P2 purinoceptor stimulation of endothelial prostacyclin production.

Science.gov (United States)

Patel, V; Brown, C; Goodwin, A; Wilkie, N; Boarder, M R

1996-11-15

Extracellular ATP and ADP, released from platelets and other sites stimulate the endothelial production of prostacyclin (PGI2) by acting on G-protein-coupled P2Y2 and P2Y2 purinoceptors, contributing to the maintenance of a non-thrombogenic surface. The mechanism, widely described as being dependent on elevated cytosolic [Ca2+], also requires protein tyrosine phosphorylation. Here we show that activation of both these P2 receptor types leads to the tyrosine phosphorylation and activation of both the p42 and p44 forms of mitogen-activated protein kinase (MAPK). 2-Methylthio-ATP and UTP, selectively activating P2Y1 and P2Y2 purinoceptors respectively, and ATP, a non-selective agonist at these two receptors, stimulate the tyrosine phosphorylation of both p42mapk and p44mapk, as revealed by Western blots with an antiserum specific for the tyrosine-phosphorylated forms of the enzymes. By using separation on Resource Q columns, peptide kinase activity associated with the phosphorylated MAPK enzymes distributes into two peaks, one mainly p42mapk and one mainly p44mapk, both of which are stimulated by ATP with respect to kinase activity and phospho-MAPK immunoreactivity. Stimulation of P2Y1 or P2Y2 purinoceptors leads to a severalfold increase in PGI2 efflux; this was blocked in a dose-dependent manner by the selective MAPK kinase inhibitor PD98059. This drug also blocked the agonist-stimulated increase in phospho-MAPK immunoreactivity for both p42mapk and p44mapk but left the phospholipase C response to P2 agonists essentially unchanged. Olomoucine has been reported to inhibit p44mapk activity. Here we show that in the same concentration range olomoucine inhibits activity in both peaks from the Resource Q column and also the agonist stimulation of 6-keto-PGF1, but has no effect on agonist-stimulated phospho-MAPK immunoreactivity. These results provide direct evidence for the involvement of p42 and p44 MAPK in the PGI2 response of intact endothelial cells: we have shown

Triage of oxidation-prone proteins by Sqstm1/p62 within the mitochondria

Energy Technology Data Exchange (ETDEWEB)

Lee, Minjung [Department of Molecular Cell Biology, Sungkyunkwan University School of Medicine and Samsung Biomedical Research Institute, Suwon-Si, Kyonggi-Do (Korea, Republic of); Shin, Jaekyoon, E-mail: jkshin@med.skku.ac.kr [Department of Molecular Cell Biology, Sungkyunkwan University School of Medicine and Samsung Biomedical Research Institute, Suwon-Si, Kyonggi-Do (Korea, Republic of)

2011-09-16

Highlights: {yields} The mitochondrion contains its own protein quality control system. {yields} p62 localizes within the mitochondria and forms mega-dalton sized complexes. {yields} p62 interacts with oxidation-prone proteins and the proteins of quality control. {yields} In vitro delivery of p62 improves mitochondrial functions. {yields} p62 is implicated as a participant in mitochondrial protein quality control. -- Abstract: As the mitochondrion is vulnerable to oxidative stress, cells have evolved several strategies to maintain mitochondrial integrity, including mitochondrial protein quality control mechanisms and autophagic removal of damaged mitochondria. Involvement of an autophagy adaptor, Sqstm1/p62, in the latter process has been recently described. In the present study, we provide evidence that a portion of p62 directly localizes within the mitochondria and supports stable electron transport by forming heterogeneous protein complexes. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF) of mitochondrial proteins co-purified with p62 revealed that p62 interacts with several oxidation-prone proteins, including a few components of the electron transport chain complexes, as well as multiple chaperone molecules and redox regulatory enzymes. Accordingly, p62-deficient mitochondria exhibited compromised electron transport, and the compromised function was partially restored by in vitro delivery of p62. These results suggest that p62 plays an additional role in maintaining mitochondrial integrity at the vicinity of target machineries through its function in relation to protein quality control.

File list: InP.Oth.50.AllAg.Olfactory_epithelium [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Oth.50.AllAg.Olfactory_epithelium mm9 Input control Others Olfactory epithelium... SRX112965,SRX185883,SRX143806,SRX378545,SRX378544 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Oth.50.AllAg.Olfactory_epithelium.bed ...

File list: InP.Dig.50.AllAg.Intestinal_villus [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Dig.50.AllAg.Intestinal_villus mm9 Input control Digestive tract Intestinal vil...lus SRX112512,SRX028556,SRX1141901,SRX193725 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Dig.50.AllAg.Intestinal_villus.bed ...

File list: InP.Adl.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Adl.50.Input_control.AllCell dm3 Input control Input control Adult SRX181432,SR...15625,SRX507386,SRX390503,SRX016141 http://dbarchive.biosciencedbc.jp/kyushu-u/dm3/assembled/InP.Adl.50.Input_control.AllCell.bed ...

File list: InP.Neu.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Neu.50.Input_control.AllCell mm9 Input control Input control Neural SRX109476,S...X685922,SRX145797,SRX150263,SRX150261 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Neu.50.Input_control.AllCell.bed ...

File list: InP.Oth.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Oth.50.Input_control.AllCell hg19 Input control Input control Others SRX253236,...RX188949,SRX189956,SRX080334,SRX080429 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Oth.50.Input_control.AllCell.bed ...

File list: InP.Bld.50.AllAg.CD20+ [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Bld.50.AllAg.CD20+ hg19 Input control Blood CD20+ SRX186673,SRX199873,SRX189978...,SRX199853,SRX189980 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Bld.50.AllAg.CD20+.bed ...

Narrowband image and the p53 protein immunoexpression in patients with ulcerative colitis and dysplasia

International Nuclear Information System (INIS)

Chao González, Lissette

2012-01-01

Patients with pancolitis and long-standing ulcerative colitis are at increased risk of developing colorectal cancer, so it is advisable to colonoscopic surveillance. The objective of this study was to identify the endoscopic visualization system of imaging with narrowband and overexpression of the p53 protein as procedures useful for the research of Dysplasia in patients with ulcerative colitis and pancolitis, of eight or more years of evolution. A prospective, descriptive study was performed on 50 patients. The Fisher exact probability test was used for the statistical study and of square Chi, with a level of significance α = 0.05. Shown with narrow-band image increases the likelihood of finding suggestive areas of Dysplasia, reduces the amount of biopsy and gets a higher proportion of diagnoses of Dysplasia in fewer samples (70.4%). The overexpression of the p53 protein was associated with the presence of dysplasia (80.0%) p < 0.001 and is immunoexpress in samples with a high degree of severity of dysplasia and the low grade. Concluded that imaging with narrowband system and overexpression of the p53 protein are procedures useful for the research of Dysplasia in these patients. (author)

File list: InP.Oth.50.AllAg.Tumour_tissues [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Oth.50.AllAg.Tumour_tissues hg19 Input control Others Tumour tissues SRX371456,...0,SRX371458,SRX371461,SRX371455,SRX371451,SRX371462 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Oth.50.AllAg.Tumour_tissues.bed ...

File list: InP.Pup.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Pup.50.Input_control.AllCell dm3 Input control Input control Pupae SRX016163,SR...X016167,SRX016169,SRX016164,SRX016165,SRX016168 http://dbarchive.biosciencedbc.jp/kyushu-u/dm3/assembled/InP.Pup.50.Input_control.AllCell.bed ...

File list: InP.Brs.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Brs.50.Input_control.AllCell mm9 Input control Input control Breast SRX483143,S...SRX403484,SRX1078982,SRX031066,SRX031214,SRX697687 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Brs.50.Input_control.AllCell.bed ...

Neutron scattering studies on protein dynamics using the human myelin peripheral membrane protein P2

Directory of Open Access Journals (Sweden)

Laulumaa Saara

2015-01-01

Full Text Available Myelin is a multilayered proteolipid membrane structure surrounding selected axons in the vertebrate nervous system, which allows the rapid saltatory conduction of nerve impulses. Deficits in myelin formation and maintenance may lead to chronic neurological disease. P2 is an abundant myelin protein from peripheral nerves, binding between two apposing lipid bilayers. We studied the dynamics of the human myelin protein P2 and its mutated P38G variant in hydrated powders using elastic incoherent neutron scattering. The local harmonic vibrations at low temperatures were very similar for both samples, but the mutant protein had increased flexibility and softness close to physiological temperatures. The results indicate that a drastic mutation of proline to glycine at a functional site can affect protein dynamics, and in the case of P2, they may explain functional differences between the two proteins.

Neutron scattering studies on protein dynamics using the human myelin peripheral membrane protein P2

Science.gov (United States)

Laulumaa, Saara; Kursula, Petri; Natali, Francesca

2015-01-01

Myelin is a multilayered proteolipid membrane structure surrounding selected axons in the vertebrate nervous system, which allows the rapid saltatory conduction of nerve impulses. Deficits in myelin formation and maintenance may lead to chronic neurological disease. P2 is an abundant myelin protein from peripheral nerves, binding between two apposing lipid bilayers. We studied the dynamics of the human myelin protein P2 and its mutated P38G variant in hydrated powders using elastic incoherent neutron scattering. The local harmonic vibrations at low temperatures were very similar for both samples, but the mutant protein had increased flexibility and softness close to physiological temperatures. The results indicate that a drastic mutation of proline to glycine at a functional site can affect protein dynamics, and in the case of P2, they may explain functional differences between the two proteins.

Solution structure of the Equine Infectious Anemia Virus p9 protein: a rationalization of its different ALIX binding requirements compared to the analogous HIV-p6 protein

Directory of Open Access Journals (Sweden)

Henklein Peter

2009-12-01

Full Text Available Abstract Background The equine infection anemia virus (EIAV p9 Gag protein contains the late (L- domain required for efficient virus release of nascent virions from the cell membrane of infected cell. Results In the present study the p9 protein and N- and C-terminal fragments (residues 1-21 and 22-51, respectively were chemically synthesized and used for structural analyses. Circular dichroism and 1H-NMR spectroscopy provide the first molecular insight into the secondary structure and folding of this 51-amino acid protein under different solution conditions. Qualitative 1H-chemical shift and NOE data indicate that in a pure aqueous environment p9 favors an unstructured state. In its most structured state under hydrophobic conditions, p9 adopts a stable helical structure within the C-terminus. Quantitative NOE data further revealed that this α-helix extends from Ser-27 to Ser-48, while the N-terminal residues remain unstructured. The structural elements identified for p9 differ substantially from that of the functional homologous HIV-1 p6 protein. Conclusions These structural differences are discussed in the context of the different types of L-domains regulating distinct cellular pathways in virus budding. EIAV p9 mediates virus release by recruiting the ALG2-interacting protein X (ALIX via the YPDL-motif to the site of virus budding, the counterpart of the YPXnL-motif found in p6. However, p6 contains an additional PTAP L-domain that promotes HIV-1 release by binding to the tumor susceptibility gene 101 (Tsg101. The notion that structures found in p9 differ form that of p6 further support the idea that different mechanisms regulate binding of ALIX to primary versus secondary L-domains types.

Interactions between whey proteins and salivary proteins as related to astringency of whey protein beverages at low pH.

Science.gov (United States)

Ye, A; Streicher, C; Singh, H

2011-12-01

Whey protein beverages have been shown to be astringent at low pH. In the present study, the interactions between model whey proteins (β-lactoglobulin and lactoferrin) and human saliva in the pH range from 7 to 2 were investigated using particle size, turbidity, and ζ-potential measurements and sodium dodecyl sulfate-PAGE. The correlation between the sensory results of astringency and the physicochemical data was discussed. Strong interactions between β-lactoglobulin and salivary proteins led to an increase in the particle size and turbidity of mixtures of both unheated and heated β-lactoglobulin and human saliva at pH ∼3.4. However, the large particle size and high turbidity that occurred at pH 2.0 were the result of aggregation of human salivary proteins. The intense astringency in whey protein beverages may result from these increases in particle size and turbidity at these pH values and from the aggregation and precipitation of human salivary proteins alone at pH salivary proteins in the interaction is a key factor in the perception of astringency in whey protein beverages. At any pH, the increases in particle size and turbidity were much smaller in mixtures of lactoferrin and saliva, which suggests that aggregation and precipitation may not be the only mechanism linked to the perception of astringency in whey protein. Copyright © 2011 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

The need to accessorize: Molecular roles of HTLV-1 p30 and HTLV-2 p28 accessory proteins in the viral life cycle

Directory of Open Access Journals (Sweden)

Rajaneesh eAnupam

2013-09-01

Full Text Available Extensive studies of HTLV-1 and HTLV-2 over the last three decades have provided detailed knowledge on viral transformation, host-viral interactions and pathogenesis. HTLV-1 is the etiological agent of adult T cell leukemia (ATL and multiple neurodegenerative and inflammatory diseases while HTLV-2 disease association remains elusive, with few infected individuals displaying neurodegenerative diseases similar to HTLV-1. The HTLV group of oncoretroviruses has a genome that encodes structural and enzymatic proteins Gag, Pro and Env, regulatory proteins Tax and Rex, and several accessory proteins from the pX region. Of these proteins, HTLV-1 p30 and HTLV-2 p28 are encoded by the open reading frame (ORF II of the pX region. Like most other accessory proteins, p30 and p28 are dispensable for in vitro viral replication and transformation but are required for efficient viral replication and persistence in vivo. Both p30 and p28 regulate viral gene expression at the post-transcriptional level whereas p30 can also function at the transcriptional level. Recently, several reports have implicated p30 and p28 in multiple cellular processes, which provide novel insight into HTLV spread and survival and ultimately pathogenesis. In this review we summarize and compare what is known about p30 and p28, highlighting their roles in viral replication and viral pathogenesis.

File list: InP.Emb.50.AllAg.Embryonic_testis [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Emb.50.AllAg.Embryonic_testis mm9 Input control Embryo Embryonic testis SRX1491...74,SRX149168,SRX149172,SRX1175150,SRX149166,SRX957136,SRX1175149 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Emb.50.AllAg.Embryonic_testis.bed ...

File list: InP.PSC.50.AllAg.Embryoid_Bodies [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.PSC.50.AllAg.Embryoid_Bodies mm9 Input control Pluripotent stem cell Embryoid Bodie...s SRX109460,SRX109458,SRX212083,SRX109456,SRX385954,SRX026526 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.PSC.50.AllAg.Embryoid_Bodies.bed ...

File list: InP.Pan.50.AllAg.PANC-1 [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Pan.50.AllAg.PANC-1 hg19 Input control Pancreas PANC-1 SRX190309,SRX150696,SRX1...99860,SRX101486,SRX190029,SRX190252,SRX644404 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Pan.50.AllAg.PANC-1.bed ...

File list: InP.Emb.50.AllAg.Embryobic_liver [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Emb.50.AllAg.Embryobic_liver mm9 Input control Embryo Embryobic liver SRX112953...,SRX143845,SRX185833,SRX377699,SRX377697,SRX377701,SRX863175 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Emb.50.AllAg.Embryobic_liver.bed ...

File list: InP.PSC.50.AllAg.EpiLC [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.PSC.50.AllAg.EpiLC mm9 Input control Pluripotent stem cell EpiLC SRX823839,SRX5...5,SRX823838,SRX823831,SRX823826,SRX590324 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.PSC.50.AllAg.EpiLC.bed ...

File list: InP.Bld.50.AllAg.Th17 [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Bld.50.AllAg.Th17 mm9 Input control Blood Th17 SRX288138,SRX495621,SRX288137,SR...X220933,SRX172342,SRX1328057,SRX1328056 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Bld.50.AllAg.Th17.bed ...

File list: InP.Dig.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Dig.50.Input_control.AllCell hg19 Input control Input control Digestive tract S...RX155744,SRX612781,SRX543681,SRX101310,SRX648244,SRX863785 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Dig.50.Input_control.AllCell.bed ...

File list: InP.ALL.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.ALL.50.Input_control.AllCell hg19 Input control Input control All cell types SR...038682,SRX389283,SRX863785,SRX1029468,SRX155724,SRX038907 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.ALL.50.Input_control.AllCell.bed ...

File list: InP.Lng.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Lng.50.Input_control.AllCell hg19 Input control Input control Lung SRX502813,SR...89952,SRX080360,SRX734236,SRX038683,SRX093320,SRX038682 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Lng.50.Input_control.AllCell.bed ...

Footprinting analysis of interactions between the largest eukaryotic RNase P/MRP protein Pop1 and RNase P/MRP RNA components.

Science.gov (United States)

Fagerlund, Robert D; Perederina, Anna; Berezin, Igor; Krasilnikov, Andrey S

2015-09-01

Ribonuclease (RNase) P and RNase MRP are closely related catalytic ribonucleoproteins involved in the metabolism of a wide range of RNA molecules, including tRNA, rRNA, and some mRNAs. The catalytic RNA component of eukaryotic RNase P retains the core elements of the bacterial RNase P ribozyme; however, the peripheral RNA elements responsible for the stabilization of the global architecture are largely absent in the eukaryotic enzyme. At the same time, the protein makeup of eukaryotic RNase P is considerably more complex than that of the bacterial RNase P. RNase MRP, an essential and ubiquitous eukaryotic enzyme, has a structural organization resembling that of eukaryotic RNase P, and the two enzymes share most of their protein components. Here, we present the results of the analysis of interactions between the largest protein component of yeast RNases P/MRP, Pop1, and the RNA moieties of the enzymes, discuss structural implications of the results, and suggest that Pop1 plays the role of a scaffold for the stabilization of the global architecture of eukaryotic RNase P RNA, substituting for the network of RNA-RNA tertiary interactions that maintain the global RNA structure in bacterial RNase P. © 2015 Fagerlund et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

Protein mobilities and P-selectin storage in Weibel–Palade bodies

OpenAIRE

Kiskin, Nikolai I.; Hellen, Nicola; Babich, Victor; Hewlett, Lindsay; Knipe, Laura; Hannah, Matthew J.; Carter, Tom

2010-01-01

Using fluorescence recovery after photobleaching (FRAP) we measured the mobilities of EGFP-tagged soluble secretory proteins in the endoplasmic reticulum (ER) and in individual Weibel–Palade bodies (WPBs) at early (immature) and late (mature) stages in their biogenesis. Membrane proteins (P-selectin, CD63, Rab27a) were also studied in individual WPBs. In the ER, soluble secretory proteins were mobile; however, following insertion into immature WPBs larger molecules (VWF, Proregion, tPA) and P...

Mice Deficient in NF-κB p50 and p52 or RANK Have Defective Growth Plate Formation and Post-natal Dwarfism

OpenAIRE

Xing, Lianping; Chen, Di; Boyce, Brendan F.

2013-01-01

NF-κBp50/p52 double knockout (dKO) and RANK KO mice have no osteoclasts and develop severe osteopetrosis associated with dwarfism. In contrast, Op/Op mice, which form few osteoclasts, and Src KO mice, which have osteoclasts with defective resorptive function, are osteopetrotic, but they are not dwarfed. Here, we compared the morphologic features of long bones from p50/p52 dKO, RANK KO, Op/Op and Src KO mice to attempt to explain the differences in their long bone lengths. We found that growth...

Hemoglobin P50 During a Simulated Ascent of Mt. Everest, Operation Everest II

Science.gov (United States)

2007-01-01

standard HbP50 would in- crease with increasing altitude [on the basis of increased levels of 2,3- diphosphoglycerate , which rose from 1.7 0.3 mmol/L...the re- sult of an increase in 2,3- diphosphoglycerate [2,3-DPG] levels induced by hypoxia (Chanutin and Curnish, 1967; Lenfant et al., 1969; Boushel... diphosphoglycerate of rats and guinea pigs. Respir. Physiol. 11:135. Bencowitz H.Z., Wagner P.D., and West J.B. (1982). Effect of change on P50 on

File list: InP.Emb.50.AllAg.Embryonic_heart [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Emb.50.AllAg.Embryonic_heart mm9 Input control Embryo Embryonic heart SRX143735...RX377685,SRX377687,SRX967654,SRX077933,SRX377683,SRX967652,SRX244285 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Emb.50.AllAg.Embryonic_heart.bed ...

File list: InP.Prs.50.AllAg.PC-3 [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Prs.50.AllAg.PC-3 hg19 Input control Prostate PC-3 SRX539667,SRX084520,SRX43319...9,SRX433195,SRX1021636,SRX1021637,SRX181768,SRX540851 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Prs.50.AllAg.PC-3.bed ...

File list: InP.Gon.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Gon.50.Input_control.AllCell hg19 Input control Input control Gonad SRX1002699,...SRX663452,SRX1002710,SRX1002709,SRX1091823,SRX663439,SRX663445,SRX663442 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Gon.50.Input_control.AllCell.bed ...

File list: InP.Adl.50.AllAg.Germline_containing_young_adult [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Adl.50.AllAg.Germline_containing_young_adult ce10 Input control Adult Germline containing young adult...p/kyushu-u/ce10/assembled/InP.Adl.50.AllAg.Germline_containing_young_adult.bed ...

Graphical analysis of pH-dependent properties of proteins predicted using PROPKA.

Science.gov (United States)

Rostkowski, Michał; Olsson, Mats H M; Søndergaard, Chresten R; Jensen, Jan H

2011-01-26

Charge states of ionizable residues in proteins determine their pH-dependent properties through their pKa values. Thus, various theoretical methods to determine ionization constants of residues in biological systems have been developed. One of the more widely used approaches for predicting pKa values in proteins is the PROPKA program, which provides convenient structural rationalization of the predicted pKa values without any additional calculations. The PROPKA Graphical User Interface (GUI) is a new tool for studying the pH-dependent properties of proteins such as charge and stabilization energy. It facilitates a quantitative analysis of pKa values of ionizable residues together with their structural determinants by providing a direct link between the pKa data, predicted by the PROPKA calculations, and the structure via the Visual Molecular Dynamics (VMD) program. The GUI also calculates contributions to the pH-dependent unfolding free energy at a given pH for each ionizable group in the protein. Moreover, the PROPKA-computed pKa values or energy contributions of the ionizable residues in question can be displayed interactively. The PROPKA GUI can also be used for comparing pH-dependent properties of more than one structure at the same time. The GUI considerably extends the analysis and validation possibilities of the PROPKA approach. The PROPKA GUI can conveniently be used to investigate ionizable groups, and their interactions, of residues with significantly perturbed pKa values or residues that contribute to the stabilization energy the most. Charge-dependent properties can be studied either for a single protein or simultaneously with other homologous structures, which makes it a helpful tool, for instance, in protein design studies or structure-based function predictions. The GUI is implemented as a Tcl/Tk plug-in for VMD, and can be obtained online at http://propka.ki.ku.dk/~luca/wiki/index.php/GUI_Web.

Structure of Pfu Pop5, an archaeal RNase P protein.

Science.gov (United States)

Wilson, Ross C; Bohlen, Christopher J; Foster, Mark P; Bell, Charles E

2006-01-24

We have used NMR spectroscopy and x-ray crystallography to determine the three-dimensional structure of PF1378 (Pfu Pop5), one of four protein subunits of archaeal RNase P that shares a homolog in the eukaryotic enzyme. RNase P is an essential and ubiquitous ribonucleoprotein enzyme required for maturation of tRNA. In bacteria, the enzyme's RNA subunit is responsible for cleaving the single-stranded 5' leader sequence of precursor tRNA molecules (pre-tRNA), whereas the protein subunit assists in substrate binding. Although in bacteria the RNase P holoenzyme consists of one large catalytic RNA and one small protein subunit, in archaea and eukarya the enzyme contains several (> or =4) protein subunits, each of which lacks sequence similarity to the bacterial protein. The functional role of the proteins is poorly understood, as is the increased complexity in comparison to the bacterial enzyme. Pfu Pop5 has been directly implicated in catalysis by the observation that it pairs with PF1914 (Pfu Rpp30) to functionally reconstitute the catalytic domain of the RNA subunit. The protein adopts an alpha-beta sandwich fold highly homologous to the single-stranded RNA binding RRM domain. Furthermore, the three-dimensional arrangement of Pfu Pop5's structural elements is remarkably similar to that of the bacterial protein subunit. NMR spectra have been used to map the interaction of Pop5 with Pfu Rpp30. The data presented permit tantalizing hypotheses regarding the role of this protein subunit shared by archaeal and eukaryotic RNase P.

File list: InP.Neu.50.AllAg.Adult_brains [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Neu.50.AllAg.Adult_brains hg19 Input control Neural Adult brains SRX643470,SRX6...43464,SRX643462,SRX643465,SRX643469,SRX643463,SRX643466,SRX643468,SRX643467 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Neu.50.AllAg.Adult_brains.bed ...

File list: InP.Kid.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Kid.50.Input_control.AllCell mm9 Input control Input control Kidney SRX804277,S...RX286402,SRX143808,SRX804276,SRX286403,SRX062965,SRX085456,SRX804278,SRX1050552 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Kid.50.Input_control.AllCell.bed ...

File list: InP.CDV.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.CDV.50.Input_control.AllCell mm9 Input control Input control Cardiovascular SRX...74,SRX517514,SRX373580,SRX320036,SRX320037,SRX1121694,SRX275462,SRX275461,SRX066549 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.CDV.50.Input_control.AllCell.bed ...

File list: InP.Kid.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Kid.50.Input_control.AllCell hg19 Input control Input control Kidney SRX973437,...,SRX114492,SRX170378,SRX691805,SRX080441,SRX1293074,SRX326417,SRX684265,SRX684263 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Kid.50.Input_control.AllCell.bed ...

Swift Detection of a 65 Day X-Ray Period from the Ultraluminous Pulsar NGC 7793 P13

Energy Technology Data Exchange (ETDEWEB)

Hu, Chin-Ping; Ng, C.-Y. [Department of Physics, The University of Hong Kong, Pokfulam Road (Hong Kong); Li, K. L. [Department of Physics and Astronomy, Michigan State University, East Lansing, MI 48824 (United States); Kong, Albert K. H. [Institute of Astronomy and Department of Physics, National Tsing Hua University, Hsinchu 30013, Taiwan (China); Lin, Lupin Chun-Che, E-mail: cphu@hku.hk, E-mail: liliray@pa.msu.edu [Institute of Astronomy and Astrophysics, Academia Sinica, Taipei 10617, Taiwan (China)

2017-01-20

NGC 7793 P13 is an ultraluminous X-ray source harboring an accreting pulsar. We report on the detection of a ∼65 day period X-ray modulation with Swift observations in this system. The modulation period found in the X-ray band is P = 65.05 ± 0.10 days and the profile is asymmetric with a fast rise and a slower decay. On the other hand, the u -band light curve collected by Swift UVOT confirmed an optical modulation with a period of P = 64.24 ± 0.13 days. We explored the phase evolution of the X-ray and optical periodicities and propose two solutions. A superorbital modulation with a period of ∼2700–4700 days probably caused by the precession of a warped accretion disk is necessary to interpret the phase drift of the optical data. We further discuss the implication if this ∼65 day periodicity is caused by the superorbital modulation. Estimated from the relationship between the spin-orbital and orbital-superorbital periods of known disk-fed high-mass X-ray binaries, the orbital period of P13 is roughly estimated as 3–7 days. In this case, an unknown mechanism with a much longer timescale is needed to interpret the phase drift. Further studies on the stability of these two periodicities with a long-term monitoring could help us to probe their physical origins.

Major membrane surface proteins of Mycoplasma hyopneumoniae selectively modified by covalently bound lipid

International Nuclear Information System (INIS)

Wise, K.S.; Kim, M.F.

1987-01-01

Surface protein antigens of Mycoplasma hyopneumoniae were identified by direct antibody-surface binding or by radioimmunoprecipitation of surface 125 I-labeled proteins with a series of monoclonal antibodies (MAbs). Radioimmunoprecipitation of TX-114-phase proteins from cells labeled with [ 35 S] methionine, 14 C-amino acids, or [ 3 H] palmitic acid showed that proteins p65, p50, and p44 were abundant and (with one other hydrophobic protein, p60) were selectively labeled with lipid. Alkaline hydroxylamine treatment of labeled proteins indicated linkage of lipids by amide or stable O-linked ester bonds. Proteins p65, p50, and p44 were highly immunogenic in the natural host as measured by immunoblots of TX-114-phase proteins with antisera from swine inoculated with whole organisms. These proteins were antigenically and structurally unrelated, since hyperimmune mouse antibodies to individual gel-purified proteins were monospecific and gave distinct proteolytic epitope maps. Intraspecies size variants of one surface antigen of M. hyopneumoniae were revealed by a MAb to p70 (defined in strain J, ATCC 25934), which recognized a large p73 component on strain VPP11 (ATCC 25617). In addition, MAb to internal, aqueous-phase protein p82 of strain J failed to bind an analogous antigen in strain VPP11

Major membrane surface proteins of Mycoplasma hyopneumoniae selectively modified by covalently bound lipid

Energy Technology Data Exchange (ETDEWEB)

Wise K.S.; Kim, M.F.

1987-12-01

Surface protein antigens of Mycoplasma hyopneumoniae were identified by direct antibody-surface binding or by radioimmunoprecipitation of surface /sup 125/I-labeled proteins with a series of monoclonal antibodies (MAbs). Radioimmunoprecipitation of TX-114-phase proteins from cells labeled with (/sup 35/S) methionine, /sup 14/C-amino acids, or (/sup 3/H) palmitic acid showed that proteins p65, p50, and p44 were abundant and (with one other hydrophobic protein, p60) were selectively labeled with lipid. Alkaline hydroxylamine treatment of labeled proteins indicated linkage of lipids by amide or stable O-linked ester bonds. Proteins p65, p50, and p44 were highly immunogenic in the natural host as measured by immunoblots of TX-114-phase proteins with antisera from swine inoculated with whole organisms. These proteins were antigenically and structurally unrelated, since hyperimmune mouse antibodies to individual gel-purified proteins were monospecific and gave distinct proteolytic epitope maps. Intraspecies size variants of one surface antigen of M. hyopneumoniae were revealed by a MAb to p70 (defined in strain J, ATCC 25934), which recognized a large p73 component on strain VPP11 (ATCC 25617). In addition, MAb to internal, aqueous-phase protein p82 of strain J failed to bind an analogous antigen in strain VPP11.

Cystoviral polymerase complex protein P7 uses its acidic C-terminal tail to regulate the RNA-directed RNA polymerase P2.

Science.gov (United States)

Alphonse, Sébastien; Arnold, Jamie J; Bhattacharya, Shibani; Wang, Hsin; Kloss, Brian; Cameron, Craig E; Ghose, Ranajeet

2014-07-15

In bacteriophages of the cystovirus family, the polymerase complex (PX) encodes a 75-kDa RNA-directed RNA polymerase (P2) that transcribes the double-stranded RNA genome. Also a constituent of the PX is the essential protein P7 that, in addition to accelerating PX assembly and facilitating genome packaging, plays a regulatory role in transcription. Deletion of P7 from the PX leads to aberrant plus-strand synthesis suggesting its influence on the transcriptase activity of P2. Here, using solution NMR techniques and the P2 and P7 proteins from cystovirus ϕ12, we demonstrate their largely electrostatic interaction in vitro. Chemical shift perturbations on P7 in the presence of P2 suggest that this interaction involves the dynamic C-terminal tail of P7, more specifically an acidic cluster therein. Patterns of chemical shift changes induced on P2 by the P7 C-terminus resemble those seen in the presence of single-stranded RNA suggesting similarities in binding. This association between P2 and P7 reduces the affinity of the former toward template RNA and results in its decreased activity both in de novo RNA synthesis and in extending a short primer. Given the presence of C-terminal acidic tracts on all cystoviral P7 proteins, the electrostatic nature of the P2/P7 interaction is likely conserved within the family and could constitute a mechanism through which P7 regulates transcription in cystoviruses. Copyright © 2014 Elsevier Ltd. All rights reserved.

Correlation between expression of p53, p21/WAF1, and MDM2 proteins and their prognostic significance in primary hepatocellular carcinoma

Directory of Open Access Journals (Sweden)

Fu Jia

2009-12-01

Full Text Available Abstract Background Tumor Protein p53 (p53, cyclin-dependent kinase inhibitor 1A (p21/WAF1, and murine double minute 2 (MDM2 participate in the regulation of cell growth. Altered expression of these gene products has been found in malignant tumors and has been associated with poor prognosis. Our aim was to investigate the expression of the 3 proteins in hepatocellular carcinoma (HCC and their prognostic significance. Methods We examined p53, p21/WAF1, and MDM2 expression in 181 pairs of HCC tissues and the adjacent hepatic tissues by performing immunohistochemistry and examined the expression of the 3 proteins in 7 pairs of HCC tissues and the adjacent hepatic tissues by using western blot analysis. Results The expression of p53, p21/WAF1, and MDM2 in the HCC tissues was significantly higher than those in the adjacent hepatic tissues (P P = 0.008. A statistical correlation was observed between expression of p53 and p21/WAF1 (R = 0.380, P = 0.000, p53 and MDM2 (R = 0.299, P = 0.000, p21/WAF1 and MDM2 (R = 0.285, P = 0.000 in 181 liver tissues adjacent to the tumor. Patients with a low pathologic grade HCC (I+II had a higher tendency to express p53 on tumor cells than the patients with high pathologic grade HCC (III+IV (P = 0.007. Survival analysis showed that positive p21/WAF1 expression or/and negative MDM2 expression in HCC was a predictor of better survival of patients after tumor resection (P Conclusions The proteins p53, p21/WAF1, and MDM2 were overexpressed in all the HCC cases in this study, and p53 and p21/WAF1 overexpression were positively correlated. The expression of p21/WAF1 and MDM2 can be considered as 2 useful indicators for predicting the prognosis of HCC.

File list: InP.Adl.50.AllAg.Young_adult [Chip-atlas[Archive

Lifescience Database Archive (English)

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Structure and stability insights into tumour suppressor p53 evolutionary related proteins.

Directory of Open Access Journals (Sweden)

Bruno Pagano

Full Text Available The p53 family of genes and their protein products, namely, p53, p63 and p73, have over one billion years of evolutionary history. Advances in computational biology and genomics are enabling studies of the complexities of the molecular evolution of p53 protein family to decipher the underpinnings of key biological conditions spanning from cancer through to various metabolic and developmental disorders and facilitate the design of personalised medicines. However, a complete understanding of the inherent nature of the thermodynamic and structural stability of the p53 protein family is still lacking. This is due, to a degree, to the lack of comprehensive structural information for a large number of homologous proteins and to an incomplete knowledge of the intrinsic factors responsible for their stability and how these might influence function. Here we investigate the thermal stability, secondary structure and folding properties of the DNA-binding domains (DBDs of a range of proteins from the p53 family using biophysical methods. While the N- and the C-terminal domains of the p53 family show sequence diversity and are normally targets for post-translational modifications and alternative splicing, the central DBD is highly conserved. Together with data obtained from Molecular Dynamics simulations in solution and with structure based homology modelling, our results provide further insights into the molecular properties of evolutionary related p53 proteins. We identify some marked structural differences within the p53 family, which could account for the divergence in biological functions as well as the subtleties manifested in the oligomerization properties of this family.

Expression of MDM2 mRNA, MDM2, P53 and P16 Proteins in Urothelial Lesions in the View of the WHO 4th Edition Guidelines as A Molecular Insight towards Personalized Medicine

Directory of Open Access Journals (Sweden)

Olfat Hammam

2017-08-01

Full Text Available AIM: Here we imposed a multimarker molecular panel composed of P53, MDM2 protein & mRNA & P16 with the identification of sensitive and specific cut offs among the Egyptian urothelial carcinomas bilharzial or not emphasize the pathological and molecular classifications, pathways and prognosis as a privilege for adjuvant therapy. METHODS: Three hundred and ten urothelial lesions were pathologically evaluated and grouped as follows: 50 chronic cystitis as benign, 240 urothelial carcinomas and 20 normal bladder tissue as a control. Immunohistochemistry for MDM Protein, P16 & p53 and In Situ Hybridization for MDM2mRNA were done. RESULTS: MDM2mRNA overexpression correlated with low grade low stage non invasive tumors, while P53 > 40% & p16 40% & P16 10% from high grade, high stage invasive urothelial carcinomas (with p53 > 40, p16 40 & p16 < 10%, together with the histopathological features can distinguish in situ urothelial lesions from dysplastic and atypical lesions.

Immunohistochemical positive stained p53 protein in bladder transitional cell carcinoma

Directory of Open Access Journals (Sweden)

Halimi Monireh

2009-04-01

Full Text Available Background: Molecular genetics and immunopathologic analysis of bladder cancer have shown some abnormalities in a number of genes and proteins that have been implicated in the development and progression of such tumors, mainly in the p53 pathway. Aims: To investigate the rate of positively stained p53 protein in patients with urothelial papillary carcinoma of the bladder (UCB by immunohistochemistry and its relationship with tumor grade, gender and age of the patients. Settings and Design: During the present cross-sectional study, 100 paraffin-embedded specimens of UCB, which were provided from biopsies of the bladder by transurethral access, were immunohistochemically stained and studied for p53 protein from May 2006 to May 2007 in our referral center pathology laboratory. Materials and Methods: First, 4 µm slices of paraffin sections were provided and then stained by the avidin-biotin peroxidase method. The rate of positively stained p53 protein (defined as positive nuclear staining in over 10% of the cells was assessed. This rate was also estimated and compared between grades, genders and age-related groups (< 70 years, ≥70 years. Statistical Analysis: The χ2 , Fisher′s exact test and Mann-Whitney U test were used for comparing. Results: The overall rate of positively stained specimens was 11% for nuclear p53 protein. This rate was significantly higher in females (10/29 vs. 1/71; P < 0.001; odds ratio [OR]: 0.23; 95% confidence interval [CI]: 4.43-306.08, patients with 70 or older than 70 years (8/42 vs. 3/58; P = 0.04; OR: 0.55; 95% CI: 1.07-17.39 and in high-grade tumors (10/58 vs. 1/42; P = 0.02; OR: 0.59; 95% CI: 0.01-0.95. Conclusions: The rate of positively stained p53 protein for UCB was lower in our population. This rate was also higher in females, patients with 70 or older than 70 years and high grade of UCB.

File list: InP.Prs.50.AllAg.Prostate_cancer_cells [Chip-atlas[Archive

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File list: InP.Adp.50.AllAg.Adipose_Tissue,_White [Chip-atlas[Archive

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File list: InP.Neu.50.AllAg.Cerebellar_granule_neurons [Chip-atlas[Archive

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Full Text Available InP.Neu.50.AllAg.Cerebellar_granule_neurons mm9 Input control Neural Cerebellar granule neurons... http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Neu.50.AllAg.Cerebellar_granule_neurons.bed ...

Generation of a selectively cytotoxic fusion protein against p53 mutated cancers

International Nuclear Information System (INIS)

Kousparou, Christina A; Yiacoumi, Efthymia; Deonarain, Mahendra P; Epenetos, Agamemnon A

2012-01-01

A significant number of cancers are caused by defects in p21 causing functional defects in p21 or p53 tumour-suppressor proteins. This has led to many therapeutic approaches including restoration by gene therapy with wild-type p53 or p21 using viral or liposomal vectors, which have toxicity or side-effect limitations. We set out to develop a safer, novel fusion protein which has the ability to reconstitute cancer cell lines with active p21 by protein transduction. The fusion protein was produced from the cell-translocating peptide Antennapedia (Antp) and wild-type, full-length p21 (Antp-p21). This was expressed and refolded from E. coli and tested on a variety of cell lines and tumours (in a BALB/c nude xenograft model) with differing p21 or p53 status. Antp-p21 penetrated and killed cancer cells that do not express wild type p53 or p21. This included cells that were matched to cogenic parental cell lines. Antp-p21 killed cancer cells selectively that were malignant as a result of mutations or nuclear exclusion of the p53 and p21 genes and over-expression of MDM2. Non-specific toxicity was excluded by showing that Antp-p21 penetrated but did not kill p53- or p21- wild-type cells. Antp-p21 was not immunogenic in normal New Zealand White rabbits. Recombinant Antp peptide alone was not cytotoxic, showing that killing was due to the transduction of the p21 component of Antp-p21. Antp-p21 was shown to penetrate cancer cells engrafted in vivo and resulted in tumour eradication when administered with conventionally-used chemotherapeutic agents, which alone were unable to produce such an effect. Antp-p21 may represent a new and promising targeted therapy for patients with p53-associated cancers supporting the concept that rational design of therapies directed against specific cancer mutations will play a part in the future of medical oncology

Generation of a selectively cytotoxic fusion protein against p53 mutated cancers

Directory of Open Access Journals (Sweden)

Kousparou Christina A

2012-08-01

Full Text Available Abstract Background A significant number of cancers are caused by defects in p21 causing functional defects in p21 or p53 tumour-suppressor proteins. This has led to many therapeutic approaches including restoration by gene therapy with wild-type p53 or p21 using viral or liposomal vectors, which have toxicity or side-effect limitations. We set out to develop a safer, novel fusion protein which has the ability to reconstitute cancer cell lines with active p21 by protein transduction. Methods The fusion protein was produced from the cell-translocating peptide Antennapedia (Antp and wild-type, full-length p21 (Antp-p21. This was expressed and refolded from E. coli and tested on a variety of cell lines and tumours (in a BALB/c nude xenograft model with differing p21 or p53 status. Results Antp-p21 penetrated and killed cancer cells that do not express wild type p53 or p21. This included cells that were matched to cogenic parental cell lines. Antp-p21 killed cancer cells selectively that were malignant as a result of mutations or nuclear exclusion of the p53 and p21 genes and over-expression of MDM2. Non-specific toxicity was excluded by showing that Antp-p21 penetrated but did not kill p53- or p21- wild-type cells. Antp-p21 was not immunogenic in normal New Zealand White rabbits. Recombinant Antp peptide alone was not cytotoxic, showing that killing was due to the transduction of the p21 component of Antp-p21. Antp-p21 was shown to penetrate cancer cells engrafted in vivo and resulted in tumour eradication when administered with conventionally-used chemotherapeutic agents, which alone were unable to produce such an effect. Conclusions Antp-p21 may represent a new and promising targeted therapy for patients with p53-associated cancers supporting the concept that rational design of therapies directed against specific cancer mutations will play a part in the future of medical oncology.

Effect of nitrogen form and pH of nutrient solution on the changes in pH and EC of spinach rhizosphere in hydroponic culture

OpenAIRE

M. Parsazadeh; N. Najafi

2011-01-01

In this study, the effect of nitrate to ammonium ratio and pH of nutrient solution on the changes in pH and EC of rhizosphere during spinach growth period in perlite culture, under greenhouse conditions, was investigated. A split factorial experiment in a completely randomized design with four replications was conducted with three factors including nutrient solution’s pH in three levels (4.5, 6.5 and 8), nitrate to ammonium ratio of nutrient solution in five levels (100:0, 75:25, 50:50, 25:75...

Exploration of auditory P50 gating in schizophrenia by way of difference waves

DEFF Research Database (Denmark)

Arnfred, Sidse M

2006-01-01

potentials but here this method along with low frequency filtering is applied exploratory on auditory P50 gating data, previously analyzed in the standard format (reported in Am J Psychiatry 2003, 160:2236-8). The exploration was motivated by the observation during visual peak detection that the AEP waveform......Electroencephalographic measures of information processing encompass both mid-latency evoked potentials like the pre-attentive auditory P50 potential and a host of later more cognitive components like P300 and N400.Difference waves have mostly been employed in studies of later event related...

File list: InP.CDV.50.AllAg.Brachiocephalic_endothelial_cells [Chip-atlas[Archive

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Rice black-streaked dwarf virus P6 self-interacts to form punctate, viroplasm-like structures in the cytoplasm and recruits viroplasm-associated protein P9-1

Directory of Open Access Journals (Sweden)

Yu Jialin

2011-01-01

Full Text Available Abstract Background Rice black-streaked dwarf virus (RBSDV, a member of the genus Fijivirus within the family Reoviridae, can infect several graminaceous plant species including rice, maize and wheat, and is transmitted by planthoppers. Although several RBSDV proteins have been studied in detail, functions of the nonstructural protein P6 are still largely unknown. Results In the current study, we employed yeast two-hybrid assays, bimolecular fluorescence complementation and subcellular localization experiments to show that P6 can self-interact to form punctate, cytoplasmic viroplasm-like structures (VLS when expressed alone in plant cells. The region from residues 395 to 659 is necessary for P6 self-interaction, whereas two polypeptides (residues 580-620 and 615-655 are involved in the subcellular localization of P6. Furthermore, P6 strongly interacts with the viroplasm-associated protein P9-1 and recruits P9-1 to localize in VLS. The P6 395-659 region is also important for the P6-P9-1 interaction, and deleting any region of P9-1 abolishes this heterologous interaction. Conclusions RBSDV P6 protein has an intrinsic ability to self-interact and forms VLS without other RBSDV proteins or RNAs. P6 recruits P9-1 to VLS by direct protein-protein interaction. This is the first report on the functionality of RBSDV P6 protein. P6 may be involved in the process of viroplasm nucleation and virus morphogenesis.

Structural basis for the ligand-binding specificity of fatty acid-binding proteins (pFABP4 and pFABP5) in gentoo penguin.

Science.gov (United States)

Lee, Chang Woo; Kim, Jung Eun; Do, Hackwon; Kim, Ryeo-Ok; Lee, Sung Gu; Park, Hyun Ho; Chang, Jeong Ho; Yim, Joung Han; Park, Hyun; Kim, Il-Chan; Lee, Jun Hyuck

2015-09-11

Fatty acid-binding proteins (FABPs) are involved in transporting hydrophobic fatty acids between various aqueous compartments of the cell by directly binding ligands inside their β-barrel cavities. Here, we report the crystal structures of ligand-unbound pFABP4, linoleate-bound pFABP4, and palmitate-bound pFABP5, obtained from gentoo penguin (Pygoscelis papua), at a resolution of 2.1 Å, 2.2 Å, and 2.3 Å, respectively. The pFABP4 and pFABP5 proteins have a canonical β-barrel structure with two short α-helices that form a cap region and fatty acid ligand binding sites in the hydrophobic cavity within the β-barrel structure. Linoleate-bound pFABP4 and palmitate-bound pFABP5 possess different ligand-binding modes and a unique ligand-binding pocket due to several sequence dissimilarities (A76/L78, T30/M32, underlining indicates pFABP4 residues) between the two proteins. Structural comparison revealed significantly different conformational changes in the β3-β4 loop region (residues 57-62) as well as the flipped Phe60 residue of pFABP5 than that in pFABP4 (the corresponding residue is Phe58). A ligand-binding study using fluorophore displacement assays shows that pFABP4 has a relatively strong affinity for linoleate as compared to pFABP5. In contrast, pFABP5 exhibits higher affinity for palmitate than that for pFABP4. In conclusion, our high-resolution structures and ligand-binding studies provide useful insights into the ligand-binding preferences of pFABPs based on key protein-ligand interactions. Copyright © 2015 Elsevier Inc. All rights reserved.

pH controlled gating of toxic protein pores by dendrimers

Science.gov (United States)

Mandal, Taraknath; Kanchi, Subbarao; Ayappa, K. G.; Maiti, Prabal K.

2016-06-01

Designing effective nanoscale blockers for membrane inserted pores formed by pore forming toxins, which are expressed by several virulent bacterial strains, on a target cell membrane is a challenging and active area of research. Here we demonstrate that PAMAM dendrimers can act as effective pH controlled gating devices once the pore has been formed. We have used fully atomistic molecular dynamics (MD) simulations to characterize the cytolysin A (ClyA) protein pores modified with fifth generation (G5) PAMAM dendrimers. Our results show that the PAMAM dendrimer, in either its protonated (P) or non-protonated (NP) states can spontaneously enter the protein lumen. Protonated dendrimers interact strongly with the negatively charged protein pore lumen. As a consequence, P dendrimers assume a more expanded configuration efficiently blocking the pore when compared with the more compact configuration adopted by the neutral NP dendrimers creating a greater void space for the passage of water and ions. To quantify the effective blockage of the protein pore, we have calculated the pore conductance as well as the residence times by applying a weak force on the ions/water. Ionic currents are reduced by 91% for the P dendrimers and 31% for the NP dendrimers. The preferential binding of Cl- counter ions to the P dendrimer creates a zone of high Cl- concentration in the vicinity of the internalized dendrimer and a high concentration of K+ ions in the transmembrane region of the pore lumen. In addition to steric effects, this induced charge segregation for the P dendrimer effectively blocks ionic transport through the pore. Our investigation shows that the bio-compatible PAMAM dendrimers can potentially be used to develop therapeutic protocols based on the pH sensitive gating of pores formed by pore forming toxins to mitigate bacterial infections.Designing effective nanoscale blockers for membrane inserted pores formed by pore forming toxins, which are expressed by several virulent

Analysis of proteins of mouse sarcoma pseudotype viruses: type-specific radioimmunoassays for ecotropic virus p30's

International Nuclear Information System (INIS)

Kennel, S.J.; Tennant, R.W.

1979-01-01

Murine sarcoma virus pseudotypes were prepared by infection of nonproducer cells (A1-2), which were transformed by the Gazdar strain of mouse sarcoma virus, with Gross (N-tropic), WN1802B (B-tropic), or Moloney (NB-tropic) viruses. The respective host range pseudotype sarcoma viruses were defined by the tritration characteristics on cells with the appropriate Fv-1 genotype. Proteins from virus progeny were analyzed by sodium dodecyl sulfate--polyacrylamide gel electrophoresis. Bands present in both the 65,000- and the 10,000- to 20,000-molecular-weight regions of the gel distinguished the pseudotype viruses from their respective helpers. Furthermore, two protein bands were noted in the p30 region of murine sarcoma virus (Gross), one corresponding to Gross virus p30, and another of slightly slower mobility. However, since the mobility of the putative sarcoma p30 is nearly indentical to that of WN1802B, its presence could not be established by sodium dodecyl sulfate--polyacrylamide gel electrophoresis. Type-specific radioimmunossays for Gross virus p30 and for WN1802B p30 were applied for analysis of pseudotype preparations, and among several ecotropic viruses tested, only the homologous virus scored in the respective assay. By use of these assays, pseudotype viruses were found to contain only 8 to 48% helper-specific p30's; the remainder is presumably derived from the sarcoma virus

Kinetics of protein adsorption/desorption mediated by pH-responsive polymer layer

International Nuclear Information System (INIS)

Su Xiao-Hang; Lei Qun-Li; Ren Chun-Lai

2015-01-01

We propose a new way of regulating protein adsorption by using a pH-responsive polymer. According to the theoretical results obtained from the molecular theory and kinetic approaches, both thermodynamics and kinetics of protein adsorption are verified to be well controlled by the solution pH. The kinetics and the amount of adsorbed proteins at equilibrium are greatly increased when the solution environment changes from acid to neutral. The reason is that the increased pH promotes the dissociation of the weak polyelectrolyte, resulting in more charged monomers and more stretched chains. Thus the steric repulsion within the polymer layer is weakened, which effectively lowers the barrier felt by the protein during the process of adsorption. Interestingly, we also find that the kinetics of protein desorption is almost unchanged with the variation of pH. It is because although the barrier formed by the polymer layer changes along with the change of pH, the potential at contact with the surface varies equally. Our results may provide useful insights into controllable protein adsorption/desorption in practical applications. (paper)

File list: InP.Brs.50.AllAg.Breast_cancer_cells [Chip-atlas[Archive

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File list: InP.Gon.50.AllAg.Testicular_somatic_cells [Chip-atlas[Archive

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File list: InP.PSC.50.AllAg.mESCs,_differentiated [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.PSC.50.AllAg.mESCs,_differentiated mm9 Input control Pluripotent stem cell mESCs, differentia...,SRX388447 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.PSC.50.AllAg.mESCs,_differentiated.bed ...

File list: InP.EmF.50.Input_control.AllCell [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.EmF.50.Input_control.AllCell mm9 Input control Input control Embryonic fibrobla...363,SRX115361 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.EmF.50.Input_control.AllCell.bed ...

Alfalfa mosaic virus replicase proteins, P1 and P2, localize to the tonoplast in the presence of virus RNA

International Nuclear Information System (INIS)

Ibrahim, Amr; Hutchens, Heather M.; Howard Berg, R.; Sue Loesch-Fries, L.

2012-01-01

To identify the virus components important for assembly of the Alfalfa mosaic virus replicase complex, we used live cell imaging of Arabidopsis thaliana protoplasts that expressed various virus cDNAs encoding native and GFP-fusion proteins of P1 and P2 replicase proteins and full-length virus RNAs. Expression of P1-GFP alone resulted in fluorescent vesicle-like bodies in the cytoplasm that colocalized with FM4-64, an endocytic marker, and RFP-AtVSR2, RabF2a/Rha1-mCherry, and RabF2b/Ara7-mCherry, all of which localize to multivesicular bodies (MVBs), which are also called prevacuolar compartments, that mediate traffic to the lytic vacuole. GFP-P2 was driven from the cytosol to MVBs when expressed with P1 indicating that P1 recruited GFP-P2. P1-GFP localized on the tonoplast, which surrounds the vacuole, in the presence of infectious virus RNA, replication competent RNA2, or P2 and replication competent RNA1 or RNA3. This suggests that a functional replication complex containing P1, P2, and a full-length AMV RNA assembles on MVBs to traffic to the tonoplast.

Alfalfa mosaic virus replicase proteins, P1 and P2, localize to the tonoplast in the presence of virus RNA

Energy Technology Data Exchange (ETDEWEB)

Ibrahim, Amr [Department of Botany and Plant Pathology, Purdue University, West Lafayette, IN 47907 (United States); Present address: Genomics Facility, Agricultural Genetic Engineering Research Institute, Agricultural Research Center, Giza 12619 (Egypt); Hutchens, Heather M. [Department of Botany and Plant Pathology, Purdue University, West Lafayette, IN 47907 (United States); Howard Berg, R. [Integrated Microscopy Facility, Donald Danforth Plant Science Center, Saint Louis, MO 63132 (United States); Sue Loesch-Fries, L., E-mail: loeschfr@purdue.edu [Department of Botany and Plant Pathology, Purdue University, West Lafayette, IN 47907 (United States)

2012-11-25

To identify the virus components important for assembly of the Alfalfa mosaic virus replicase complex, we used live cell imaging of Arabidopsis thaliana protoplasts that expressed various virus cDNAs encoding native and GFP-fusion proteins of P1 and P2 replicase proteins and full-length virus RNAs. Expression of P1-GFP alone resulted in fluorescent vesicle-like bodies in the cytoplasm that colocalized with FM4-64, an endocytic marker, and RFP-AtVSR2, RabF2a/Rha1-mCherry, and RabF2b/Ara7-mCherry, all of which localize to multivesicular bodies (MVBs), which are also called prevacuolar compartments, that mediate traffic to the lytic vacuole. GFP-P2 was driven from the cytosol to MVBs when expressed with P1 indicating that P1 recruited GFP-P2. P1-GFP localized on the tonoplast, which surrounds the vacuole, in the presence of infectious virus RNA, replication competent RNA2, or P2 and replication competent RNA1 or RNA3. This suggests that a functional replication complex containing P1, P2, and a full-length AMV RNA assembles on MVBs to traffic to the tonoplast.

Cross reaction between P-61 sunflower seedlings oleosomal protein ...

African Journals Online (AJOL)

A true triacylglycerol lipase was detected in germinating sunflower (Helianthus annuus L.) seedlings associated to oleosomes. This enzyme that has not yet been identified was partially purified as shown by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS PAGE) (10%); two protein bands, P-61 and P-66 ...

Novel insights into the role of NF-κB p50 in astrocyte-mediated fate specification of adult neural progenitor cells

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Valeria Bortolotto

2017-01-01

Full Text Available Within the CNS nuclear factor-kappa B (NF-κB transcription factors are involved in a wide range of functions both in homeostasis and in pathology. Over the years, our and other groups produced a vast array of information on the complex involvement of NF-κB proteins in different aspects of postnatal neurogenesis. In particular, several extracellular signals and membrane receptors have been identified as being able to affect neural progenitor cells (NPC and their progeny via NF-κB activation. A crucial role in the regulation of neuronal fate specification in adult hippocampal NPC is played by the NF-κB p50 subunit. NF-κB p50KO mice display a remarkable reduction in adult hippocampal neurogenesis which correlates with a selective defect in hippocampal-dependent short-term memory. Moreover absence of NF-κB p50 can profoundly affect the in vitro proneurogenic response of adult hippocampal NPC (ahNPC to several endogenous signals and drugs. Herein we briefly review the current knowledge on the pivotal role of NF-κB p50 in the regulation of adult hippocampal neurogenesis. In addition we discuss more recent data that further extend the relevance of NF-κB p50 to novel astroglia-derived signals which can influence neuronal specification of ahNPC and to astrocyte-NPC cross-talk.

DNA-binding proteins regulating pIP501 transfer and replication

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Elisabeth Grohmann

2016-08-01

Full Text Available pIP501 is a Gram-positive broad-host-range model plasmid intensively used for studying plasmid replication and conjugative transfer. It is a multiple antibiotic resistance plasmid frequently found in clinical Enterococcus faecalis and Enterococcus faecium isolates. Replication of pIP501 proceeds unidirectionally by a theta mechanism. The minimal replicon of pIP501 is composed of the repR gene encoding the essential rate-limiting replication initiator protein RepR and the origin of replication, oriR, located downstream of repR. RepR is similar to RepE of related streptococcal plasmid pAMβ1, which has been shown to possess RNase activity cleaving free RNA molecules in close proximity of the initiation site of DNA synthesis. Replication of pIP501 is controlled by the concerted action of a small protein, CopR, and an antisense RNA, RNAIII. CopR has a dual role: It acts as transcriptional repressor at the repR promoter and prevents convergent transcription of RNAIII and repR mRNA (RNAII, thereby indirectly increasing RNAIII synthesis. CopR binds asymmetrically as a dimer at two consecutive binding sites upstream of and overlapping with the repR promoter. RNAIII induces transcriptional attenuation within the leader region of the repR mRNA (RNAII. Deletion of either control component causes a 10- to 20-fold increase of plasmid copy number, while simultaneous deletions have no additional effect. Conjugative transfer of pIP501 depends on a type IV secretion system (T4SS encoded in a single operon. Its transfer host-range is considerably broad, as it has been transferred to virtually all Gram-positive bacteria including filamentous streptomycetes and even the Gram-negative Escherichia coli. Expression of the 15 genes encoding the T4SS is tightly controlled by binding of the relaxase TraA, the transfer initiator protein, to the operon promoter, which overlaps with the origin of transfer (oriT. The T4SS operon encodes the DNA-binding proteins TraJ (VirD4

Effect of initial protein concentration and pH on in vitro gastric digestion of heated whey proteins.

Science.gov (United States)

Zhang, Sha; Vardhanabhuti, Bongkosh

2014-02-15

The in vitro digestion of heated whey protein aggregates having different structure and physicochemical properties was evaluated under simulated gastric conditions. Aggregates were formed by heating whey protein isolates (WPI) at 3-9% w/w initial protein concentration and pH 3.0-7.0. Results showed that high protein concentration led to formation of larger WPI aggregates with fewer remaining monomers. Aggregates formed at high protein concentrations showed slower degradation rate compared to those formed at low protein concentration. The effect of initial protein concentration on peptide release pattern was not apparent. Heating pH was a significant factor affecting digestion pattern. At pH above the isoelectric point, the majority of the proteins involved in the aggregation, and aggregates formed at pH 6.0 were more susceptible to pepsin digestion than at pH 7.0. At acidic conditions, only small amount of proteins was involved in the aggregation and heated aggregates were easily digested by pepsin, while the remaining unaggregated proteins were very resistant to gastric digestion. The potential physiological implication of these results on satiety was discussed. Copyright © 2013 Elsevier Ltd. All rights reserved.

p53 Protein interacts specifically with the meiosis-specific mammalian RecA-like protein DMC1 in meiosis.

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Habu, Toshiyuki; Wakabayashi, Nobunao; Yoshida, Kayo; Yomogida, Kenntaro; Nishimune, Yoshitake; Morita, Takashi

2004-06-01

The tumor suppressor protein p53 is specifically expressed during meiosis in spermatocytes. Subsets of p53 knockout mice exhibit testicular giant cell degenerative syndrome, which suggests p53 may be associated with meiotic cell cycle and/or DNA metabolism. Here, we show that p53 binds to the mouse meiosis-specific RecA-like protein Mus musculus DMC1 (MmDMC1). The C-terminal domain (amino acid 234-340) of MmDMC1 binds to DNA-binding domain of p53 protein. p53 might be involved in homologous recombination and/or checkpoint function by directly binding to DMC1 protein to repress genomic instability in meiotic germ cells.

Simultaneous Hypoxia and Low Extracellular pH Suppress Overall Metabolic Rate and Protein Synthesis In Vitro.

Science.gov (United States)

Sørensen, Brita Singers; Busk, Morten; Overgaard, Jens; Horsman, Michael R; Alsner, Jan

2015-01-01

The tumor microenvironment is characterized by regions of hypoxia and acidosis which are linked to poor prognosis. This occurs due to an aberrant vasculature as well as high rates of glycolysis and lactate production in tumor cells even in the presence of oxygen (the Warburg effect), which weakens the spatial linkage between hypoxia and acidosis. Five different human squamous cell carcinoma cell lines (SiHa, FaDuDD, UTSCC5, UTSCC14 and UTSCC15) were treated with hypoxia, acidosis (pH 6.3), or a combination, and gene expression analyzed using microarray. SiHa and FaDuDD were chosen for further characterization of cell energetics and protein synthesis. Total cellular ATP turnover and relative glycolytic dependency was determined by simultaneous measurements of oxygen consumption and lactate synthesis rates and total protein synthesis was determined by autoradiographic quantification of the incorporation of 35S-labelled methionine and cysteine into protein. Microarray analysis allowed differentiation between genes induced at low oxygen only at normal extracellular pH (pHe), genes induced at low oxygen at both normal and low pHe, and genes induced at low pHe independent of oxygen concentration. Several genes were found to be upregulated by acidosis independent of oxygenation. Acidosis resulted in a more wide-scale change in gene expression profiles than hypoxia including upregulation of genes involved in the translation process, for example Eukaryotic translation initiation factor 4A, isoform 2 (EIF4A2), and Ribosomal protein L37 (RPL37). Acidosis suppressed overall ATP turnover and protein synthesis by 50%. Protein synthesis, but not total ATP production, was also suppressed under hypoxic conditions. A dramatic decrease in ATP turnover (SiHa) and protein synthesis (both cell lines) was observed when hypoxia and low pHe were combined. We demonstrate here that the influence of hypoxia and acidosis causes different responses, both in gene expression and in de novo

File list: InP.Adp.50.AllAg.Adipose_progenitor_cells [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Adp.50.AllAg.Adipose_progenitor_cells mm9 Input control Adipocyte Adipose progeni...tor cells SRX127370,SRX127367 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Adp.50.AllAg.Adipose_progenitor_cells.bed ...

Profile of NF-κBp(65/NFκBp50) among prostate specific antigen sera levels in prostatic pathologies.

Science.gov (United States)

Bouraoui, Y; Ben Jemaa, A; Rodriguez, G; Ben Rais, N; Fraile, B; Paniagua, R; Sellemi, S; Royuela, M; Oueslati, R

2012-10-01

The aim of this work was to characterise the immunoexpression of NF-κB (p50/p65) in human prostatic pathologies and to study its profiles of activation among sera prostate specific antigen antigen (PSA) according the three groups: 0-4ng/mL, 4-20ng/mL and >20ng/mL. Twenty-four men with benign prostate hyperplasia (BPH); 19 men with prostate cancer (PC) and five men with normal prostates (NP). Immunohistochemical and western blot analysis was performed. Serum levels of PSA were assayed by immulite autoanalyser. In BPH and PC samples, immunoexpressions were observed for NF-κBp65 and NF-κBp50; while in NP samples, only were detected NF-κBp50. PC samples showed immunoreactions to NF-κBp65 and NF-κBp50 more intense (respectively 24.18±0.67 and 28.23±2.01) than that observed in BPH samples (respectively18.46±2.04 and 18.66±1.59) with special localisation in the nucleus. Different profiles of NF-κBp65 immunoexpressions were observed and BPH patients with sera PSA levels between 0-4ng/mL presented a significant weak percentage compared to BPH patients with sera PSA levels between 4-20ng/mL and >20ng/mL. No immunoreactions to NF-κBp65 were observed in PC patients with sera PSA levels between 4-20ng/mL. The sensibility of both NF-κB and PSA to inflammation allowed confirming the relationship between these two molecules and its involvement in prostatic diseases progression (inflammatory and neoplasic). Copyright © 2011 Elsevier Masson SAS. All rights reserved.

File list: InP.Bld.50.AllAg.Naive_T_cells [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Bld.50.AllAg.Naive_T_cells hg19 Input control Blood Naive T cells SRX1425815,SR...X1425816,SRX1425814 http://dbarchive.biosciencedbc.jp/kyushu-u/hg19/assembled/InP.Bld.50.AllAg.Naive_T_cells.bed ...

File list: InP.Emb.50.AllAg.Mitotic_cycle_14 [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Emb.50.AllAg.Mitotic_cycle_14 dm3 Input control Embryo Mitotic cycle 14 SRX6451...40,SRX750075,SRX645139 http://dbarchive.biosciencedbc.jp/kyushu-u/dm3/assembled/InP.Emb.50.AllAg.Mitotic_cycle_14.bed ...

File list: InP.Neu.50.AllAg.Induced_neural_progenitors [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.Neu.50.AllAg.Induced_neural_progenitors mm9 Input control Neural Induced neural... progenitors SRX323563,SRX323574,SRX667380,SRX668238 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.Neu.50.AllAg.Induced_neural_progenitors.bed ...

Expression of pH-sensitive green fluorescent protein in Arabidopsis thaliana

Science.gov (United States)

Moseyko, N.; Feldman, L. J.

2001-01-01

This is the first report on using green fluorescent protein (GFP) as a pH reporter in plants. Proton fluxes and pH regulation play important roles in plant cellular activity and therefore, it would be extremely helpful to have a plant gene reporter system for rapid, non-invasive visualization of intracellular pH changes. In order to develop such a system, we constructed three vectors for transient and stable transformation of plant cells with a pH-sensitive derivative of green fluorescent protein. Using these vectors, transgenic Arabidopsis thaliana and tobacco plants were produced. Here the application of pH-sensitive GFP technology in plants is described and, for the first time, the visualization of pH gradients between different developmental compartments in intact whole-root tissues of A. thaliana is reported. The utility of pH-sensitive GFP in revealing rapid, environmentally induced changes in cytoplasmic pH in roots is also demonstrated.

Insertion of the LINE-1 element in the C-MYC gene and immunoreactivity of C-MYC, p53, p21 and p27 proteins in different morphological patterns of the canine TVT

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C.R.O. Lima

2016-06-01

Full Text Available ABSTRACT The canine transmissible venereal tumor (TVT affects the external genitalia of dogs by the natural transplant of viable tumor cells. Thus, this research aimed to diagnose and characterize TVT morphological patterns, identify the insertion of the LINE-1 element in C-MYC gene, by means of the polymerase chain reaction (PCR, and evaluate the immunohistochemical expression of C-MYC, p53, p21 and p27 proteins. The relationship between C-MYC and p53 proteins and their interference on the expression of p21 and p27 were also studied. For that, 20 samples of naturally occurring TVT were used, subjected to cytopathological, histopathological and immunohistochemical analysis, and to molecular diagnosis of neoplasia. The increased tissue expression and the correlation among C-MYC, p53, p21 and p27 proteins indicate reduction and/or loss of their functionality in the TVT microenvironment, with consequent apoptotic suppression, maintenance of cell growth and progression of neoplasia.

A genome-wide association study identifies protein quantitative trait loci (pQTLs.

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David Melzer

2008-05-01

Full Text Available There is considerable evidence that human genetic variation influences gene expression. Genome-wide studies have revealed that mRNA levels are associated with genetic variation in or close to the gene coding for those mRNA transcripts - cis effects, and elsewhere in the genome - trans effects. The role of genetic variation in determining protein levels has not been systematically assessed. Using a genome-wide association approach we show that common genetic variation influences levels of clinically relevant proteins in human serum and plasma. We evaluated the role of 496,032 polymorphisms on levels of 42 proteins measured in 1200 fasting individuals from the population based InCHIANTI study. Proteins included insulin, several interleukins, adipokines, chemokines, and liver function markers that are implicated in many common diseases including metabolic, inflammatory, and infectious conditions. We identified eight Cis effects, including variants in or near the IL6R (p = 1.8x10(-57, CCL4L1 (p = 3.9x10(-21, IL18 (p = 6.8x10(-13, LPA (p = 4.4x10(-10, GGT1 (p = 1.5x10(-7, SHBG (p = 3.1x10(-7, CRP (p = 6.4x10(-6 and IL1RN (p = 7.3x10(-6 genes, all associated with their respective protein products with effect sizes ranging from 0.19 to 0.69 standard deviations per allele. Mechanisms implicated include altered rates of cleavage of bound to unbound soluble receptor (IL6R, altered secretion rates of different sized proteins (LPA, variation in gene copy number (CCL4L1 and altered transcription (GGT1. We identified one novel trans effect that was an association between ABO blood group and tumour necrosis factor alpha (TNF-alpha levels (p = 6.8x10(-40, but this finding was not present when TNF-alpha was measured using a different assay , or in a second study, suggesting an assay-specific association. Our results show that protein levels share some of the features of the genetics of gene expression. These include the presence of strong genetic effects in cis

Enhancement of uranium loading on ion exchange resin from carbonate leachate by lowering pH from 8 to 6.5

International Nuclear Information System (INIS)

Otto, J.B.

1984-01-01

This paper discusses a laboratory study that shows the saturation ion-exchange loading of uranium from carbonate leachate can be doubled by lowering the pH of the leachate from 8 to 6.5. Small column and batch resin loading tests using Dowex 21K ion-exchange resin are described. The leachate contained 3,300 ppm chloride, 2,400 ppm carbonate, and 220 ppm U 3 O 8 , and had a pH of 8. Even at this rather mild salinity the saturation ion-exchange loading was found to be only about 3 to 4 lbm U 3 O 8 /cu ft resin (48 to 64 g/dm 3 ) because of competition with the chloride ion for exchange sites on the anionic resin. Lowering the pH of the leachate to 6.5 by CO 2 gas addition, however, increased loading to about 8 lbm U 3 O 8 /cu ft resin (128 g/dm 3 ). The pH-lowering effect worked especially well at relatively high salt concentration. The same leachate, with its chloride content increased to 12,000 ppm, loaded only 0.5 lbm U 3 O 8 /cu ft resin (8 g/dm 3 ) at pH 8 but loaded 5.5 lbm U 3 O 8 /cu ft resin (88 g/dm 3 ) at pH 6.5

Bicarbonate uptake via an anion exchange protein is the main mechanism of inorganic carbon acquisition by the giant kelp Macrocystis pyrifera (Laminariales, Phaeophyceae) under variable pH.

Science.gov (United States)

Fernández, Pamela A; Hurd, Catriona L; Roleda, Michael Y

2014-12-01

Macrocystis pyrifera is a widely distributed, highly productive, seaweed. It is known to use bicarbonate (HCO3 (-) ) from seawater in photosynthesis and the main mechanism of utilization is attributed to the external catalyzed dehydration of HCO3 (-) by the surface-bound enzyme carbonic anhydrase (CAext ). Here, we examined other putative HCO3 (-) uptake mechanisms in M. pyrifera under pHT 9.00 (HCO3 (-) : CO2  = 940:1) and pHT 7.65 (HCO3 (-) : CO2  = 51:1). Rates of photosynthesis, and internal CA (CAint ) and CAext activity were measured following the application of AZ which inhibits CAext , and DIDS which inhibits a different HCO3 (-) uptake system, via an anion exchange (AE) protein. We found that the main mechanism of HCO3 (-) uptake by M. pyrifera is via an AE protein, regardless of the HCO3 (-) : CO2 ratio, with CAext making little contribution. Inhibiting the AE protein led to a 55%-65% decrease in photosynthetic rates. Inhibiting both the AE protein and CAext at pHT 9.00 led to 80%-100% inhibition of photosynthesis, whereas at pHT 7.65, passive CO2 diffusion supported 33% of photosynthesis. CAint was active at pHT 7.65 and 9.00, and activity was always higher than CAext , because of its role in dehydrating HCO3 (-) to supply CO2 to RuBisCO. Interestingly, the main mechanism of HCO3 (-) uptake in M. pyrifera was different than that in other Laminariales studied (CAext -catalyzed reaction) and we suggest that species-specific knowledge of carbon uptake mechanisms is required in order to elucidate how seaweeds might respond to future changes in HCO3 (-) :CO2 due to ocean acidification. © 2014 Phycological Society of America.

In Silico Characterization and Structural Modeling of Dermacentor andersoni p36 Immunosuppressive Protein

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Martin Omulindi Oyugi

2018-01-01

Full Text Available Ticks cause approximately $17–19 billion economic losses to the livestock industry globally. Development of recombinant antitick vaccine is greatly hindered by insufficient knowledge and understanding of proteins expressed by ticks. Ticks secrete immunosuppressant proteins that modulate the host’s immune system during blood feeding; these molecules could be a target for antivector vaccine development. Recombinant p36, a 36 kDa immunosuppressor from the saliva of female Dermacentor andersoni, suppresses T-lymphocytes proliferation in vitro. To identify potential unique structural and dynamic properties responsible for the immunosuppressive function of p36 proteins, this study utilized bioinformatic tool to characterize and model structure of D. andersoni p36 protein. Evaluation of p36 protein family as suitable vaccine antigens predicted a p36 homolog in Rhipicephalus appendiculatus, the tick vector of East Coast fever, with an antigenicity score of 0.7701 that compares well with that of Bm86 (0.7681, the protein antigen that constitute commercial tick vaccine Tickgard™. Ab initio modeling of the D. andersoni p36 protein yielded a 3D structure that predicted conserved antigenic region, which has potential of binding immunomodulating ligands including glycerol and lactose, found located within exposed loop, suggesting a likely role in immunosuppressive function of tick p36 proteins. Laboratory confirmation of these preliminary results is necessary in future studies.

Kinetics of protein adsorption/desorption mediated by pH-responsive polymer layer

Science.gov (United States)

Su, Xiao-Hang; Lei, Qun-Li; Ren, Chun-Lai

2015-11-01

We propose a new way of regulating protein adsorption by using a pH-responsive polymer. According to the theoretical results obtained from the molecular theory and kinetic approaches, both thermodynamics and kinetics of protein adsorption are verified to be well controlled by the solution pH. The kinetics and the amount of adsorbed proteins at equilibrium are greatly increased when the solution environment changes from acid to neutral. The reason is that the increased pH promotes the dissociation of the weak polyelectrolyte, resulting in more charged monomers and more stretched chains. Thus the steric repulsion within the polymer layer is weakened, which effectively lowers the barrier felt by the protein during the process of adsorption. Interestingly, we also find that the kinetics of protein desorption is almost unchanged with the variation of pH. It is because although the barrier formed by the polymer layer changes along with the change of pH, the potential at contact with the surface varies equally. Our results may provide useful insights into controllable protein adsorption/desorption in practical applications. Project supported by the National Natural Science Foundation of China (Grant Nos. 21274062, 11474155, and 91027040).

Computational Studies of pH Sensing Design Principles in Proteins

Science.gov (United States)

Garrido Ruiz, Diego

Changes in pH are important regulatory signals for biological function, under physiological and pathological conditions. Recent advances in computer simulations strategies have made the exploration of the effects of charge titrations on protein function possible. In this work, I make use of these strategies to investigate the thermodynamic coupling between conformation and protonation states that give rise to pH-dependent function. As motivation for the rest of the work, I start by presenting a collaborative investigation on a pH-sensing mutant of the EGFR tyrosine kinase common to a set of distinct cancers. From then, I reduce the complexity of the systems under study to build models where exact enumeration of states is possible to inquire about the nature of the couplings between protonation states and conformation. Finally, I discuss detailed simulations of pH-sensing proteins for which I use the expectations and insights generated with simple models to identify and interpret couplings of interest for pH-dependent behavior.

Heterologous protein secretion in Lactobacilli with modified pSIP vectors.

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Ingrid Lea Karlskås

Full Text Available We describe new variants of the modular pSIP-vectors for inducible gene expression and protein secretion in lactobacilli. The basic functionality of the pSIP system was tested in Lactobacillus strains representing 14 species using pSIP411, which harbors the broad-host-range Lactococcus lactis SH71rep replicon and a β-glucuronidase encoding reporter gene. In 10 species, the inducible gene expression system was functional. Based on these results, three pSIP vectors with different signal peptides were modified by replacing their narrow-host-range L. plantarum 256rep replicon with SH71rep and transformed into strains of five different species of Lactobacillus. All recombinant strains secreted the target protein NucA, albeit with varying production levels and secretion efficiencies. The Lp_3050 derived signal peptide generally resulted in the highest levels of secreted NucA. These modified pSIP vectors are useful tools for engineering a wide variety of Lactobacillus species.

Phage Lambda P Protein: Trans-Activation, Inhibition Phenotypes and their Suppression

Science.gov (United States)

Hayes, Sidney; Erker, Craig; Horbay, Monique A.; Marciniuk, Kristen; Wang, Wen; Hayes, Connie

2013-01-01

The initiation of bacteriophage λ replication depends upon interactions between the oriλ DNA site, phage proteins O and P, and E. coli host replication proteins. P exhibits a high affinity for DnaB, the major replicative helicase for unwinding double stranded DNA. The concept of P-lethality relates to the hypothesis that P can sequester DnaB and in turn prevent cellular replication initiation from oriC. Alternatively, it was suggested that P-lethality does not involve an interaction between P and DnaB, but is targeted to DnaA. P-lethality is assessed by examining host cells for transformation by ColE1-type plasmids that can express P, and the absence of transformants is attributed to a lethal effect of P expression. The plasmid we employed enabled conditional expression of P, where under permissive conditions, cells were efficiently transformed. We observed that ColE1 replication and plasmid establishment upon transformation is extremely sensitive to P, and distinguish this effect from P-lethality directed to cells. We show that alleles of dnaB protect the variant cells from P expression. P-dependent cellular filamentation arose in ΔrecA or lexA[Ind-] cells, defective for SOS induction. Replication propagation and restart could represent additional targets for P interference of E. coli replication, beyond the oriC-dependent initiation step. PMID:23389467

Carprofen induction of p75NTR-dependent apoptosis via the p38 mitogen-activated protein kinase pathway in prostate cancer cells.

Science.gov (United States)

Khwaja, Fatima S; Quann, Emily J; Pattabiraman, Nagarajan; Wynne, Shehla; Djakiew, Daniel

2008-11-01

The p75 neurotrophin receptor (p75(NTR)) functions as a tumor suppressor in prostate epithelial cells, where its expression declines with progression to malignant cancer. Previously, we showed that treatment with R-flurbiprofen or ibuprofen induced p75(NTR) expression in several prostate cancer cell lines leading to p75(NTR)-mediated decreased survival. Using the 2-phenyl propionic acid moiety of these profens as a pharmacophore, we screened an in silico database of 30 million compounds and identified carprofen as having an order of magnitude greater activity for induction of p75(NTR) levels and inhibition of cell survival. Prostate (PC-3 and DU-145) and bladder (T24) cancer cells were more sensitive to carprofen induction of p75(NTR)-associated loss of survival than breast (MCF-7) and fibroblast (3T3) cells. Transfection of prostate cell lines with a dominant-negative form of p75(NTR) before carprofen treatment partially rescued cell survival, showing a cause-and-effect relationship between carprofen induction of p75(NTR) levels and inhibition of survival. Carprofen induced apoptotic nuclear fragmentation in prostate but not in MCF-7 and 3T3 cells. Furthermore, small interfering RNA knockdown of the p38 mitogen-activated protein kinase (MAPK) protein prevented induction of p75(NTR) by carprofen in both prostate cell lines. Carprofen treatment induced phosphorylation of p38 MAPK as early as within 1 min. Expression of a dominant-negative form of MK2, the kinase downstream of p38 MAPK frequently associated with signaling cascades leading to apoptosis, prevented carprofen induction of the p75(NTR) protein. Collectively, we identify carprofen as a highly potent profen capable of inducing p75(NTR)-dependent apoptosis via the p38 MAPK pathway in prostate cancer cells.

pKa predictions for proteins, RNAs, and DNAs with the Gaussian dielectric function using DelPhi pKa.

Science.gov (United States)

Wang, Lin; Li, Lin; Alexov, Emil

2015-12-01

We developed a Poisson-Boltzmann based approach to calculate the pKa values of protein ionizable residues (Glu, Asp, His, Lys and Arg), nucleotides of RNA and single stranded DNA. Two novel features were utilized: the dielectric properties of the macromolecules and water phase were modeled via the smooth Gaussian-based dielectric function in DelPhi and the corresponding electrostatic energies were calculated without defining the molecular surface. We tested the algorithm by calculating pKa values for more than 300 residues from 32 proteins from the PPD dataset and achieved an overall RMSD of 0.77. Particularly, the RMSD of 0.55 was achieved for surface residues, while the RMSD of 1.1 for buried residues. The approach was also found capable of capturing the large pKa shifts of various single point mutations in staphylococcal nuclease (SNase) from pKa-cooperative dataset, resulting in an overall RMSD of 1.6 for this set of pKa's. Investigations showed that predictions for most of buried mutant residues of SNase could be improved by using higher dielectric constant values. Furthermore, an option to generate different hydrogen positions also improves pKa predictions for buried carboxyl residues. Finally, the pKa calculations on two RNAs demonstrated the capability of this approach for other types of biomolecules. © 2015 Wiley Periodicals, Inc.

The L polymerase protein of parainfluenza virus 3 forms an oligomer and can interact with the heterologous Sendai virus L, P and C proteins

International Nuclear Information System (INIS)

Smallwood, Sherin; Moyer, Sue A.

2004-01-01

We recently showed that the L protein of Sendai virus is present as an oligomer in the active P-L polymerase complex [Smallwood et al., Virology 304 (2002) 235]. We now demonstrate using two different epitope tags that the L protein of a second respirovirus, human parainfluenza type 3 virus (PIV3), also forms an L-L complex. L oligomerization requires the coexpression of the differentially epitope tagged L proteins. By exploiting a series of C-terminal truncations the L-L binding site maps to the N-terminal half of L. There is some complex formation between the heterologous PIV3 and Sendai L and P proteins; however, the heterologous L protein does not function in transcription of either the PIV3 or Sendai template. The PIV3 C protein binds PIV3 L and inhibits RNA synthesis in vitro and in vivo. Significant homology exists between the C proteins of PIV3 and Sendai and complex formation occurs between the PIV3 and Sendai heterologous C and L proteins. In addition, the heterologous C proteins can inhibit transcription at ∼50% of the level of the homologous protein. These data suggest that while the C proteins may be functionally somewhat interchangeable, the L and P proteins are specific for each virus

pH induced protein-scaffold biosynthesis of tunable shape gold nanoparticles

International Nuclear Information System (INIS)

Zhang Xiaorong; He Xiaoxiao; Wang Kemin; Ren Fang; Qin Zhihe

2011-01-01

In this paper, a pH-inductive protein-scaffold biosynthesis of shape-tunable crystalline gold nanoparticles at room temperature has been developed. By simple manipulation of the reaction solution's pH, anisotropic gold nanoparticles including spheres, triangles and cubes could be produced by incubating an aqueous solution of sodium tetrachloroaurate with Dolichomitriopsis diversiformis biomasses after immersion in ultrapure Millipore water overnight. A moss protein with molecular weight of about 71 kDa and pI of 4.9 was the primary biomolecule involved in the biosynthesis of gold nanoparticles. The secondary configuration of the proteins by CD spectrum implied that the moss protein could display different secondary configurations including random coil, α-helix and intermediate conformations between random coil and α-helix for the experimental pH solution. The growth process of gold nanoparticles further showed that the moss protein with different configurations provided the template scaffold for the shape-controlled biosynthesis of gold nanoparticles. The constrained shape of the gold nanoparticles, however, disappeared in boiled moss extract. The gold nanoparticles with designed morphology were successfully reconstructed using the moss protein purified from the gold nanoparticles. Structural characterizations by SEM, TEM and SAED showed that the triangular and cubic gold nanoparticles were single crystalline.

The Polerovirus silencing suppressor P0 targets ARGONAUTE proteins for degradation.

Science.gov (United States)

Baumberger, Nicolas; Tsai, Ching-Hsui; Lie, Miranda; Havecker, Ericka; Baulcombe, David C

2007-09-18

Plant and animal viruses encode suppressor proteins of an adaptive immunity mechanism in which viral double-stranded RNA is processed into 21-25 nt short interfering (si)RNAs. The siRNAs guide ARGONAUTE (AGO) proteins so that they target viral RNA. Most viral suppressors bind long dsRNA or siRNAs and thereby prevent production of siRNA or binding of siRNA to AGO. The one exception is the 2b suppressor of Cucumoviruses that binds to and inhibits AGO1. Here we describe a novel suppressor mechanism in which a Polerovirus-encoded F box protein (P0) targets the PAZ motif and its adjacent upstream sequence in AGO1 and mediates its degradation. F box proteins are components of E3 ubiquitin ligase complexes that add polyubiquitin tracts on selected lysine residues and thereby mark a protein for proteasome-mediated degradation. With P0, however, the targeted degradation of AGO is insensitive to inhibition of the proteasome, indicating that the proteasome is not involved. We also show that P0 does not block a mobile signal of silencing, indicating that the signal molecule does not have AGO protein components. The ability of P0 to block silencing without affecting signal movement may contribute to the phloem restriction of viruses in the Polerovirus group.

Exploration of somatosensory P50 gating in schizophrenia spectrum patients

DEFF Research Database (Denmark)

Arnfred, Sidse M; Chen, Andrew C N

2004-01-01

, male, schizophrenia spectrum patients (seven schizophrenic and five schizotypal personality disorder patients) and 14 age-matched healthy men participated in recordings of pair-wise presented auditory and median nerve stimuli. The patients had smaller amplitudes of the SEP P50 at the first stimulus...

Towards the structure of yeast and mammalian P4-ATPases

DEFF Research Database (Denmark)

Lyons, Joseph; Laban, Milena; Mikkelsen, Stine

2017-01-01

P4-ATPases are members of the P-type ATPase superfamily that drive the inward translocation (flipping) of lipids within the membrane. These lipid flippase largely function as binary complexes with an auxiliary protein from the CDC50 family. The bulk of our knowledge has derived genetic and bioche......P4-ATPases are members of the P-type ATPase superfamily that drive the inward translocation (flipping) of lipids within the membrane. These lipid flippase largely function as binary complexes with an auxiliary protein from the CDC50 family. The bulk of our knowledge has derived genetic...... a basis for the analysis of reported mutagenesis data, we aim to solve the first molecular structures of the PS transporting P4-ATPases using electron microscopy. To date, negative stain EM analysis, on detergent, amphipol and saposin-lipoprotein nanoparticle (Salipro) reconstituted of both Drs2p/CDC50p...... and bATP8A2/CDC50A, has yielded comparable low-resolution envelopes of these two transporters, highlighting the bulk architecture of the complex. Current efforts and progress on the functional characterization and cryo-EM analysis of both lipid transporters reconstituted in Salipro are described...

File list: InP.PSC.50.AllAg.iPSC_intermediates [Chip-atlas[Archive

Lifescience Database Archive (English)

Full Text Available InP.PSC.50.AllAg.iPSC_intermediates mm9 Input control Pluripotent stem cell iPSC intermediates...178448,SRX1178449,SRX1184108,SRX1178444,SRX897944,SRX1178447 http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/InP.PSC.50.AllAg.iPSC_intermediates.bed ...

P-body proteins regulate transcriptional rewiring to promote DNA replication stress resistance.