Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.5.1.18 (glutathione S-transferase)
22,582 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The cells derived from the human embryo liver tissue were transfected with a plasmid pSV3neo containing both the large and small T-antigen gene of the early region of simian virus 40 (SV40), and two cell strains, OUMS-21 and -22, were obtained. OUMS-22 cells, to date, have reached over 100 population doublings through a culture crisis and are considered to have become an immortal cell line. However, OUMS-21 cells failed to become an immortal cell line. Both OUMS-21 and -22 cells were SV40 T-antigen-positive, epithelial-like, and immunoreactive against an anti-keratin 18 monoclonal antibody but against neither an anti-vimentin nor an anti-von Willebrandt factor VIII monoclonal antibody. The staining pattern of cytokeratin in these cells was similar to that in the differentiated human hepatoblastoma and hepatocellular carcinoma cell lines but not to that in the human cholangiocellular carcinoma cell lines. OUMS-21 and -22 cells expressed neither alpha-fetoprotein nor albumin mRNAs. These cells showed no tyrosine aminotransferase activity. However, both OUMS-21 and -22 cells were sensitive to cytotoxicity of aflatoxin B1, 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole, and benzo[a]pyrene, whereas human embryo lung fibroblasts were insensitive to the cytotoxicity of these carcinogens. These findings suggest that OUMS-21 and -22 cells may arise from undifferentiated liver stem cells or from hepatocytes that lost their ability to express the liver-specific functions prior to immortalization. Both OUMS-21 and -22 cells expressed glutathione S-transferase pi (GST-pi) mRNA. The expression of GST-pi mRNA highly increased in OUMS-22 cells with their immortalization. Karyotypic analysis showed that numerical and structural aberrations of the chromosomes were profound, but neither specific events nor marker chromosomes were found in OUMS-21 and -22 cells. Both OUMS-21 and -22 cells could grow in soft agar, but they were not tumorigenic when transplanted into nude mice.
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PMID:Immortalization of epithelial-like cells from human liver tissue with SV40 T-antigen gene. 768 77

The integrin alpha 6 beta 4 is a major component of hemidesmosomes, in which it is linked to intermediate filaments. Its presence in these structures is dependent on the beta 4 cytoplasmic domain but it is not known whether beta 4 interacts directly with keratin filaments or by interaction with other proteins. In this study, we have investigated the interaction of GST-cyto beta 4A fusion proteins with cellular proteins and demonstrate that a fragment of beta 4A, consisting of the two pairs of fibronectin type III repeats, separated by the connecting segment, forms a specific complex containing a 500-kDa protein that comigrates with HD1, a hemidesmosomal plaque protein. A similar protein was also bound by a glutathione S-transferase fusion protein containing the cytoplasmic domain of a variant beta 4 subunit (beta 4B), in which a stretch of 53 amino acids is inserted in the connecting segment. Subsequent immunoblot analysis revealed that the 500-kDa protein is in fact HD1. In COS-7 cells, which do not express alpha 6 beta 4 or the hemidesmosomal components BP230 and BP180, HD1 is associated with the cytoskeleton, but after transfecting the cells with cDNAs for human alpha 6 and beta 4, it was, instead, colocalized with alpha 6 beta 4 at the basal side of the cells. The organization of the vimentin, keratin, actin, and tubulin cytoskeletal networks was not affected by the expression of alpha 6 beta 4 in COS-7 cells. The localization of HD1 at the basal side of the cells depends on the same region of beta 4 that forms a complex containing HD1 in vitro, since the expression of alpha 6 with a mutant beta 4 subunit that lacks the four fibronectin type III repeats and the connecting segment did not alter the distribution of HD1. The results indicate that for association of alpha 6 beta 4 with HD1, the cytoplasmic domain of beta 4 is essential. We suggest that this association may be crucial for hemidesmosome assembly.
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PMID:Integrin alpha 6 beta 4 forms a complex with the cytoskeletal protein HD1 and induces its redistribution in transfected COS-7 cells. 924 37

Variants of the human ovarian carcinoma cell line, OAW42, exhibiting low-level intrinsic resistance (OAW42-SR) and drug-induced higher-level resistance (OAW42-A1 & OAW42-A), were studied along with a sensitive clonal population (OAW42-S) which was isolated from OAW42-SR. Expression of the MDR-associated protein P-170, the more recently discovered LRP (lung resistance-related protein) and MRP (multidrug resistance-associated protein), topoisomerase II alpha and beta, GST pi and the cytoskeletal proteins, cytokeratin 8 and vimentin, were studied (using immunocytochemistry and Western blotting techniques) in conjunction with drug (doxorubicin) accumulation and subcellular distribution. Expression of mRNA for P-170, MRP, topoisomerase 11 alpha and beta and GST pi was studied using RT-PCR (reverse transcriptase polymerase chain reaction). Results indicate differential co-expression of four MDR-associated parameters (P-170, MRP, LRP and reduced topoisomerase II alpha and beta) in the OAW42-SR and OAW42-A1 variants, whereas resistance in the OAW42-A variant appeared to be mainly P-170 mediated. Comparable amounts of MRP and greater amounts of LRP were detected in the OAW42-S cells compared to the OAW42-SR variant (which showed increased resistance compared to the OAW42-S cells), but all cell lines expressed similar low-level amounts of MRP mRNA (by RT-PCR). GST pi levels did not differ markedly between variants. Increased levels of the cytoskeletal proteins were observed with increasing levels of resistance. The relative resistance of the variants, OAW42-SR and OAW42-A1, compared with OAW42-S was seen to change during increased serial passaging of the cells. There was greater drug accumulation by the sensitive OAW42-S cell line compared with that of the resistant variants, particularly the most highly resistant OAW42-A cells. Both verapamil and cyclosporin A effectively restored the accumulation defects seen in the resistant variants, cyclosporin A being the more effective of the two. Sub-cellular location of drug was predominantly in the nucleus with maximum levels seen in the sensitive OAW42-S variant and minimum levels in the most resistant OAW42-A clone.
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PMID:Co-expression of MDR-associated markers, including P-170, MRP and LRP and cytoskeletal proteins, in three resistant variants of the human ovarian carcinoma cell line, OAW42. 927 50

Active neuronal-glial interaction is important in the maintenance of brain homeostasis and is vital for neuronal survival following brain injury. The time course of post-ischemic astroglial dysfunction and neuronal death was studied in the spontaneously hypertensive rat (SHR) brain following permanent middle cerebral artery occlusion (MCAO). In situ hybridization with 35S-labeled riboprobes for GFAP and GLUT3 was used to monitor mRNA expression in glia and neurons. Astrocytic proteins GFAP, vimentin, S100, Glutathione-S-Transferase Yb (GST Yb) and neuronal protein TG2 were detected by immunofluorescence. Cells were co-stained with in situ end labeling (ISEL) to detect DNA fragmentation, a hallmark of cell death. GFAP mRNA expression declined rapidly in the ischemic region of the cortex and was almost absent by 12 h. Immunohistochemical studies revealed a parallel decline in the corresponding protein: a reduction in GFAP staining was apparent in the infarct after 3 h and by 24 h, there was essentially no remaining GFAP. Three other glial proteins (vimentin, S100 and GST Yb) disappeared from infarct over a similar time course. A few ISEL positive cells were observed in the infarct at 6 h, but maximal detection was not seen until 24-48 h. Most of the ISEL-positive cells were neurons, identified by co-staining with the neuronal marker TG2. Few cells expressing GFAP or other glial markers were positive at any time point. Neuronal GLUT3 mRNA declined more slowly than GFAP mRNA in the ischemic core and disappeared during the period of neuronal death. Concurrent with the loss of GFAP mRNA and protein expression in the infarct, there was a rapid rise in GFAP mRNA in the peri-infarct region of ipsilateral hemisphere and proximal region of the contralateral hemisphere. This was followed by the enhanced GFAP protein expression characteristic of reactive astrocytes, but over a significantly slower time course. These studies show that MCAO leads to a rapid decline of GFAP mRNA and glial proteins, which appears to precede the decline in neuronal mRNA and neuronal death within the infarct. Early astroglial dysfunction may play a critical role in determining the outcome of acute hypoxic-ischemic injury by compromising neuronal-glial interactions.
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PMID:Astrocytic demise precedes delayed neuronal death in focal ischemic rat brain. 1032 Jul 81

The presence of the intermediate filament protein nestin has been the predominant marker used to describe stem and progenitor cells in the mammalian CNS. In this study, a 998-bp fragment in the 3' region of the nestin mRNA was cloned from human fetal brain cells (HFBC). The nucleotide sequence of the cloned cDNA revealed 21 differences with the previously published human nestin sequence, resulting in 17 amino acid changes. A 150-amino-acid fragment derived from the cloned nestin cDNA was coupled to glutathione S-transferase and used as an immunogen to generate a rabbit polyclonal antiserum that selectively detects human nestin. HFBC that proliferated in response to basic fibroblast growth factor incorporated 5-bromo-2'-deoxyuridine into their nuclei and immunostained for nestin, indicating nestin expression in proliferating CNS progenitor cells. In all cell cultures, nestin costained with the neuroepithelial cell marker vimentin. A small subset of nestin-stained cells (1-2%) immunostained with neuronal marker MAP-2 during the first week and after 4 weeks in culture. However, during the first week in culture, approximately 10-30% of the total cell population of HFBC stained for the glial cell marker GFAP, and nearly all coimmunostained for nestin. After 4 weeks in culture, a subset of GFAP-positive cells emerged that no longer costained with nestin. These results describe nestin expression not only in CNS progenitor cells but also in the cells which were in transition from a progenitor stage to glial differentiation. Collectively, these data suggest a differential temporal regulation of nestin expression during glial and neuronal cell differentiation.
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PMID:Coexpression of nestin in neural and glial cells in the developing human CNS defined by a human-specific anti-nestin antibody. 1068 78

Using immobilized GST-Raf-1 as bait, we have isolated the intermediate filament protein vimentin as a Raf-1-associated protein. Vimentin coimmunoprecipitated and colocalized with Raf-1 in fibroblasts. Vimentin was not a Raf-1 substrate, but was phosphorylated by Raf-1-associated vimentin kinases. We provide evidence for at least two Raf-1-associated vimentin kinases and identified one as casein kinase 2. They are regulated by Raf-1, since the activation status of Raf-1 correlated with the phosphorylation of vimentin. Vimentin phosphorylation by Raf-1 preparations interfered with its polymerization in vitro. A subset of tryptic vimentin phosphopeptides induced by Raf-1 in vitro matched the vimentin phosphopeptides isolated from v-raf-transfected cells labeled with orthophosphoric acid, indicating that Raf-1 also induces vimentin phosphorylation in intact cells. In NIH 3T3 fibroblasts, the selective activation of an estrogen-regulated Raf-1 mutant induced a rearrangement and depolymerization of the reticular vimentin scaffold similar to the changes elicited by serum treatment. The rearrangement of the vimentin network occurred independently of the MEK/ERK pathway. These data identify a new branch point in Raf-1 signaling, which links Raf-1 to changes in the cytoskeletal architecture.
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PMID:The Raf-1 kinase associates with vimentin kinases and regulates the structure of vimentin filaments. 1102 85

Polycystin-1, the protein defective in a majority of patients with autosomal dominant polycystic kidney disease, is a ubiquitously expressed multi-span transmembrane protein of unknown function. Subcellular localization studies found this protein to be a component of various cell junctional complexes and to be associated with the cytoskeleton, but the specificity and nature of such associations are not known. To identify proteins that interact with the polycystin-1 C-tail (P1CT), this segment was used as bait in a yeast two-hybrid screening of a kidney epithelial cell library. The intermediate filament (IF) protein vimentin was identified as a strong polycystin-1-interacting partner. Cytokeratins K8 and K18 and desmin were also found to interact with P1CT. These interactions were mediated by coiled-coil motifs in polycystin-1 and IF proteins. Vimentin, cytokeratins K8 and K18, and desmin also bound directly to P1CT in GST pull-down and in in vitro filament assembly assays. Two observations confirmed these interactions in vivo: (i) a cell membrane-anchored form of recombinant P1CT decorated the IF network and was found to associate with the cytoskeleton in detergent-solubilized cells and (ii) endogenous polycystin-1 distributed with IF at desmosomal junctions. Polycystin-1 may utilize this association for structural, storage, or signaling functions.
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PMID:Polycystin-1 interacts with intermediate filaments. 1158 Dec 69

Recently the multiple endocrine neoplasia type 1 (MEN1) tumor suppressor gene was cloned. Its protein product, called menin, has been shown to associate with the AP1 transcription factor JunD and to repress JunD-mediated transcription. However, little is known concerning the regulation of menin. Here we report that menin interacts with the type III intermediate filament (IF) proteins glial fibrillary acidic protein (GFAP) and vimentin. Menin's interaction with these IF proteins was characterized and confirmed both in vitro and in vivo using GST pull-down analysis, co-immunoprecipitation experiments, and immunofluorescence studies. Deletion mutants of GFAP or vimentin involving the head domains of the molecules abolish the interaction with menin. Endogenous menin is colocalized with GFAP and vimentin in glioma cells as determined by confocal microscopy. Furthermore, a tailless GFAP deletion mutant, which disrupts the IF network, results in menin/GFAP/vimentin-containing aggregates. Triple immunofluorescence labeling studies with antibodies against menin, BrdU, and GFAP show that menin and GFAP colocalize in glioma cells at the S-G2 phase of the cell cycle, as measured by BrdU incorporation. Our data suggest that the intermediate filament network interacts with and may serve as a cytoplasmic sequestering network for menin at the S and early G2 phase of the cell cycle.
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PMID:Menin's interaction with glial fibrillary acidic protein and vimentin suggests a role for the intermediate filament network in regulating menin activity. 1216 73

Rab1 GTPases participate in regulating the vesicular transport of ER-Golgi compartments. Recently, GM130, p115, and Golgin-84 were identified as effectors of the active conformation of rab1. Here, we describe a novel protein, MICAL-1b, a splice variant of the MICAL-1a protein. Using the yeast two-hybrid system, we showed that it specifically interacts with rab1 in a nucleotide-dependent manner. The interaction was confirmed by GST pulldown experiments. Cell fractionation revealed that in contrast to the mainly membrane-associated rab1 effector GM130, MICAL-1 displays a predominantly cytosolic localization. We mapped the rab1 interacting domain to the C-terminus of MICAL-1, which also mediates binding to the intermediate filament vimentin. Therefore, the interaction of MICAL-1 and rab1 might provide a link between the Golgi apparatus and the intermediate filament cytoskeleton. We suggest that MICAL-1 isoforms with their multidomain structure are novel rab1 interacting proteins that function as scaffold proteins connecting different components in the cell.
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PMID:MICAL-1 isoforms, novel rab1 interacting proteins. 1278 69

Proteomics is a relatively new approach for understanding the pathology and pathogenesis of various diseases. It has also been used for characterizing the modifications in protein expression during the development of interstitial lung diseases, in lung tumors, or following exposure to exogenous stress signals. We compared the protein composition of primary human lung fibroblasts derived from patients with lung fibrosis and control fibroblasts of unaffected lung tissues. We found a predominant modulation in proteins related to the cytoskeleton, including decreased expression of vimentin and lamin A/C, and increased expression of moesin. Furthermore, we observed lower levels of components of the antioxidative system, such as omega class glutathione S-transferase and an up-regulation of an intracellular chloride channel. In fibroblasts obtained from fibrotic lungs, the expression of a major histocompatibility complex class I C was decreased, and so was the expression of tripeptidyl-peptidase-I-precursor, a collagen-degrading exopeptidase. Our results and the studies reviewed in this paper represent just the beginning of detailed studies that should unravel the relevance and the functional consequences of differential protein expressions in the diseased lung.
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PMID:Clinical proteomics in lung diseases. 1523 23


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