As thermostable enzymes and organisms are much more needed, researches on heat shock proteins(HSPs) of hyperthermophilic archaea have drawn more concerns. HSPs from hyperthermophilic archaea are concise only with HSP60, sHSP, prefoldin and AAA+proteins, but without HSP100s, HSP90s, HSP70 (DnaK), HSP40 (DnaJ) and GrpE which are common in mesophilic or thermophilic archaea. Accordingly, studies on the structure, function and operation mechanism of these four groups are much more important and meaningful. This review focuses on the recent progress in the researchs on the structure, function, operation mechanism and cooperation of the HSPs from hyperthermophilic archaea. The problems and obfuscations in these HSPs are analyzed, and farther research direction and key points are put out.
Archaea ; classification ; metabolism ; Archaeal Proteins ; metabolism ; Chaperonin 60 ; metabolism ; Heat-Shock Proteins ; genetics ; metabolism ; Molecular Chaperones ; metabolism

Objective: To investigate the expression characteristic of the Daxx gene in the mouse testis and its role in spermatogenesis. METHODS: Real-time PCR, Western blot and immunofluorescence were used in examining the expression characteristics of DAXX in the testis tissue from wild-type, Sertoli cell-specific androgen receptor knockout (SCARKO) and androgen receptor knockout (ARKO) mice at different postnatal weeks . RESULTS: The Daxx gene was highly expressed in the testis tissue and mainly in the nuclei of the wild-type mice at 4 postnatal weeks. Compared with the wild-type, the ARKO mice showed a markedly decreased expression of DAXX (0.299±0.026), which displayed a polar distribution in the spermatogenic cells (0.853±0.058) and exhibited no significant difference in the SCARKO mice (1.000±0.015). CONCLUSIONS The Daxx gene expression is the highest in the middle-stage development of the mouse testis, significantly decreased in ARKO mice as compared with the wild-type, and its location influenced by specific AR knockout in Sertoli cells. DAXX may be involved in the regulation of spermatogenesis in mice.
Animals ; Carrier Proteins ; genetics ; metabolism ; Cell Nucleus ; genetics ; metabolism ; Gene Expression ; Intracellular Signaling Peptides and Proteins ; genetics ; metabolism ; Male ; Mice ; Mice, Knockout ; Molecular Chaperones ; Nuclear Proteins ; genetics ; metabolism ; Receptors, Androgen ; genetics ; Sertoli Cells ; Spermatogenesis ; genetics ; Testis ; metabolism

OBJECTIVETo identify gene differential expression in embryo and adult testes by cDNA microarray techniques.

METHODScDNA probes of embryo and adult testes were used to hybridize the cDNA microarray of human testis, and the clones of differential hybridization were sequenced and analyzed.

RESULTSThe calmegin (CLGN) gene which had been confirmed to be involved in spermatogenesis was found, and it exhibited more than 20-fold difference at expression level between adult and embryo human testes.

CONCLUSIONGenes differential expression in adult and embryo human testes can be identified by cDNA microarray hybridization.

Adult ; Base Sequence ; Calcium-Binding Proteins ; chemistry ; genetics ; Cloning, Molecular ; Gene Expression Regulation, Developmental ; Humans ; Male ; Molecular Chaperones ; chemistry ; genetics ; Molecular Sequence Data ; Nucleic Acid Hybridization ; Oligonucleotide Array Sequence Analysis ; Spermatogenesis ; genetics ; Testis ; embryology ; metabolism

OBJECTIVESTo investigate the function and possible mechanisms of PIAS3 expression on the invasion of TJ905 cells.

METHODSPIAS3 overexpression vectors were constructed and PIAS3 siRNA were chemically synthesized, which were separately transfected into TJ905 cells for upregulation or downregulation of PIAS3 expression levels in TJ905 cells. After that, the invasive effects of TJ905 cells were measured by Transwell assay, and the expression of PIAS3, tissue inhibitor of metalloproteinases (TIMP)3, matrix metalloprotease (MMP)-2, and MMP-9 were identified by Western blot.

RESULTSIn vitro transfection efficiency of plasmids and oligonucleotides were separately 85.3% ± 3.1% and 95.1% ± 2.9%. PIAS3 overexpression plasmid transfection in vitro could effectively improve the expression of PIAS3 protein in TJ905 cells and inhibit the invasion of TJ905 cells (P < 0.05), and cell penetration ratio reduced from 87.9% ± 9.3% to 37.3% ± 7.9% compared with control group, while it upregulated TIMP3 and downregulated MMP-2, MMP-9 protein expression (P < 0.05); PIAS3 siRNA transfection could inhibit the PIAS3 protein expression of TJ905 cells and promote the invasion of TJ905 cells (P < 0.05), and cell penetration ratio increased from 83.9% ± 7.1% to 93.2% ± 3.1% compared with control group, while it downregulated TIMP3 and upregulated MMP-2, MMP-9 protein expression (P < 0.05).

CONCLUSIONPIAS3 expression is closely related to the invasion properties of glioma TJ905 cells.

Cell Line, Tumor ; Genetic Vectors ; Glioma ; metabolism ; pathology ; Humans ; Matrix Metalloproteinase 2 ; metabolism ; Matrix Metalloproteinase 9 ; metabolism ; Molecular Chaperones ; genetics ; metabolism ; Neoplasm Invasiveness ; Protein Inhibitors of Activated STAT ; genetics ; metabolism ; RNA, Small Interfering ; genetics ; Tissue Inhibitor of Metalloproteinase-3 ; metabolism ; Transfection

Calmegin is a testis-specific molecular chaperon playing a key role in spermatogenesis. However, the transcriptional regulatory mechanisms for calmegin expression are entirely unknown. Herein, we revealed that calmegin is transcriptionally regulated by histone deacetylase (HDAC) and CpG methyltransferase. The cDNA microarray analysis of the human fibrosarcoma cells treated with trichostatin A (TSA) showed an increased level of calmegin mRNA. The induction of calmegin mRNA by TSA was added by the treatment with 5-aza-2'-deoxycytidine (5'Aza- dC), implying that epigenetic alterations are involved in the transcriptional repression of the gene. Moreover, chromatin immunoprecipitation assay using an anti-acetyl-histone H3 antibody exhibited that the proximal region (-152~-31) of the calmegin promoter is responsible for HDAC-mediated transcriptional repression of the gene. These results demonstrate that calmegin expression is regulated by HDAC and CpG methyltransferase in a coordinative way.
Animals ; Calcium-Binding Proteins/*genetics ; Cell Line, Tumor ; *Gene Expression Regulation ; Histone Deacetylases/*metabolism ; Humans ; Male ; Methyltransferases/*metabolism ; Mice ; Molecular Chaperones/*genetics ; Organ Specificity ; Promoter Regions (Genetics)/genetics ; Research Support, Non-U.S. Gov't ; Testis/*metabolism ; *Transcription, Genetic

OBJECTIVEPseudomonas aeruginosa is a ubiquitous and opportunistic pathogen that uses the type III secretion system (TTSS) to inject effector proteins directly into the cytosol of target cells to subvert the host cell's functions. Specialized bacterial chaperones are required for effective secretion of some effectors. To identify the chaperone of ExoS, the representative effector secreted by the TTSS of P. aeruginosa, we analyzed the role of a postulated chaperone termed Orf1.

METHODSBy allelic exchange, we constructed the mutant with the deletion of gene Orf1. Analysis of secreted and cell-associated fractions was performed by SDS-PAGE and Western blotting. Using strain expressing in trans Orf1, tagged by V5 polypeptide and histidine, protein-protein interaction was determined by affinity resin pull-down assay in combination with MALDI-TOF. The role of Orf1 in the expression of exoS was evaluated by gene reporter analysis.

RESULTSPull-down assay showed that Orf1 binds to ExoS and ExoT. Secretion profile analysis showed that Orf1 was necessary for the optimal secretion of ExoS and ExoT. However, Orf1 had no effect on the expression of exoS.

CONCLUSIONOrf1 is important for the secretion of ExoS probably by maintaining ExoS in a secretion-competent conformation. We propose to name Orf1 as SpcS for "specific Pseudomonas chaperone for ExoS".

ADP Ribose Transferases ; genetics ; metabolism ; Bacterial Toxins ; genetics ; metabolism ; Base Sequence ; Blotting, Western ; DNA Primers ; Electrophoresis, Polyacrylamide Gel ; Kinetics ; Molecular Chaperones ; genetics ; metabolism ; Protein Binding ; Pseudomonas aeruginosa ; metabolism ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

Dendritic-cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN; CD209) has an important role in mediating adherence of Mycobacteria species, including M. tuberculosis and M. bovis BCG to human dendritic cells and macrophages, in which these bacteria can survive intracellularly. DC-SIGN is a C-type lectin, and interactions with mycobacterial cells are believed to occur via mannosylated structures on the mycobacterial surface. Recent studies suggest more varied modes of binding to multiple mycobacterial ligands. Here we identify, by affinity chromatography and mass-spectrometry, four novel ligands of M. bovis BCG that bind to DC-SIGN. The novel ligands are chaperone protein DnaK, 60 kDa chaperonin-1 (Cpn60.1), glyceraldehyde-3 phosphate dehydrogenase (GAPDH) and lipoprotein lprG. Other published work strongly suggests that these are on the cell surface. Of these ligands, lprG appears to bind DC-SIGN via typical proteinglycan interactions, but DnaK and Cpn60.1 binding do not show evidence of carbohydrate-dependent interactions. LprG was also identified as a ligand for DC-SIGNR (L-SIGN; CD299) and the M. tuberculosis orthologue of lprG has been found previously to interact with human toll-like receptor 2. Collectively, these findings offer new targets for combating mycobacterial adhesion and within-host survival, and reinforce the role of DCSIGN as an important host ligand in mycobacterial infection.
Amino Acid Sequence ; Bacterial Adhesion ; physiology ; Bacterial Proteins ; genetics ; metabolism ; Cell Adhesion Molecules ; genetics ; metabolism ; Chromatography, Affinity ; Dendritic Cells ; metabolism ; microbiology ; Host-Pathogen Interactions ; genetics ; physiology ; Humans ; In Vitro Techniques ; Lectins, C-Type ; genetics ; metabolism ; Ligands ; Macrophages ; metabolism ; microbiology ; Mass Spectrometry ; Membrane Proteins ; genetics ; metabolism ; Models, Biological ; Molecular Chaperones ; genetics ; metabolism ; Molecular Sequence Data ; Mycobacterium bovis ; genetics ; metabolism ; Mycobacterium tuberculosis ; genetics ; metabolism ; pathogenicity ; Pulmonary Surfactant-Associated Protein A ; metabolism ; Receptors, Cell Surface ; genetics ; metabolism

OBJECTIVETo identify the differentially expressed proteins or peptides and potential biomarkers of tumorigenesis for colorectal cancers.

METHODSImmobilized pH gradient two-dimensional gel electrophoresis (2-DE) was used to separate and obtain the differentially expressed protein spots between colorectal cancers and matched normal mucosa. Liquid chromatography/mass spectrometry (LC-MS/MS) was used to characterize these proteins. Selected candidate proteins were further studied by Western blot, semi-quantitative RT-PCR and immunohistochemical staining.

RESULTSThirty-five protein spots showed marked expression changes (more than 5-fold) in colorectal carcinoma compared to normal mucosa. Fifteen proteins were up regulated and 20 were down regulated. Fourteen of these proteins were identified by tandem mass spectrometry, among which secretagogin (SCGN) was down-regulated and glucose-related protein (GRP) 78 was up-regulated in the tumors. The SCGN down-regulation was further supported by Western blot and RT-PCR analyses. Immunohistochemistry revealed that SCGN was strongly expressed in neuroendocrine cells of the colonic crypts and 53 of 54 (98%) neuroendocrine tumors. At protein level, although GRP78 was up regulated in colorectal carcinoma, there was no difference in the mRNA expression level between the tumor and paired normal mucosa.

CONCLUSIONSThe 2-DE combined with MS is a powerful tool for screening potential tumor biomarkers. The differentially expressed candidate proteins identified by 2-DE may be of significance in understanding the tumorigenesis of the colon cancer. SCGN is a potential biomarker for neuroendocrinal differentiation. GRP78 up-regulation in colorectal carcinomas may be related to its post-translational modification.

Biomarkers, Tumor ; genetics ; metabolism ; Calcium-Binding Proteins ; genetics ; metabolism ; Colorectal Neoplasms ; metabolism ; Electrophoresis, Gel, Two-Dimensional ; Gene Expression Profiling ; methods ; Gene Expression Regulation, Neoplastic ; Heat-Shock Proteins ; genetics ; metabolism ; Humans ; Immunohistochemistry ; Molecular Chaperones ; genetics ; metabolism ; Neuroendocrine Cells ; metabolism ; Neuroendocrine Tumors ; metabolism ; Proteomics ; methods ; RNA, Messenger ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Secretagogins

OBJECTIVETo discuss the effect of Yisui Shengxue granules on expression of alpha-hemoglobin stabilizing protein (AHSP) mRNA in different developmental stages mice.

METHODThe total RNAs were extracted from the bone marrow karyocyte of normal adult mice and the karyocyte of fetus liver and fetus spleen in pregnanted mice (pregnanted 21 days) and fetal mice (pregnanted 14 days). The expression level of AHSP mRNA in different developmental stages mice interfered with Yisui Shengxue granules was measured by real-time PCR.

RESULTThe intervention of Yisui Shengxue granules could significantly up-regulated the expression levels of AHSP mRNA in normal adult mice.

CONCLUSIONThe result revealed that one of possible molecular mechanism of the effects caused by Yisui Shengxue granules is that it can promote the AHSP gene expression, reduce the free a-globin deposit, then prevent the poison to erythrocyte and decrease the haemolysis.

Animals ; Blood Proteins ; genetics ; Bone Marrow Cells ; cytology ; drug effects ; metabolism ; Drug Combinations ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; Erythrocytes ; cytology ; drug effects ; metabolism ; Female ; Gene Expression Regulation, Developmental ; drug effects ; Liver ; cytology ; embryology ; metabolism ; Male ; Mice ; Molecular Chaperones ; genetics ; Plants, Medicinal ; chemistry ; Pregnancy ; RNA, Messenger ; biosynthesis ; genetics ; Random Allocation ; Spleen ; cytology ; embryology ; metabolism ; Up-Regulation ; drug effects ; genetics

When we treated rat bone marrow stromal cells (rBMSCs) with neuronal differentiation induction media, typical unfolded protein response (UPR) was observed. BIP/GRP78 protein expression was time-dependently increased, and three branches of UPR were all activated. ATF6 increased the transcription of XBP1 which was successfully spliced by IRE1. PERK was phosphorylated and it was followed by eIF2alpha phosphorylation. Transcription of two downstream targets of eIF2alpha, ATF4 and CHOP/GADD153, were transiently up-regulated with the peak level at 24 h. Immunocytochemical study showed clear coexpression of BIP and ATF4 with NeuN and Map2, respectively. UPR was also observed during the neuronal differentiation of mouse embryonic stem (mES) cells. Finally, chemical endoplasmic reticulum (ER) stress inducers, thapsigargin, tunicamycin, and brefeldin A, dose-dependently increased both mRNA and protein expressions of NF-L, and, its expression was specific to BIP-positive rBMSCs. Our results showing the induction of UPR during neuronal differentiations of rBMSCs and mES cells as well as NF-L expression by ER stress inducers strongly suggest the potential role of UPR in neuronal differentiation.
Activating Transcription Factor 4/genetics/metabolism ; Animals ; Apoptosis/drug effects ; Bone Marrow Cells/*cytology ; Cell Differentiation ; Culture Media/pharmacology ; Embryonic Stem Cells/*cytology ; Endoplasmic Reticulum/genetics/metabolism ; Gene Expression/drug effects ; Heat-Shock Proteins/*genetics/metabolism ; Mice ; Microtubule-Associated Proteins/genetics/metabolism ; Molecular Chaperones/*genetics/metabolism ; Nerve Tissue Proteins/genetics/*metabolism ; Neurofilament Proteins/genetics/metabolism ; Neurons/*cytology ; Nuclear Proteins/genetics/metabolism ; Protein Folding ; Rats ; Stromal Cells