Biomedical subjects

Zhuo Zhang

Publications and source records attributed to Zhuo Zhang.

59 records · Page 4Linked to original sources

PubMed2002

Arsenite-induced Cdc25C degradation is through the KEN-box and ubiquitin-proteasome pathway.

Arsenite is a known human carcinogen that induces tumorigenesis through either a genotoxic or an epigenetic mechanism. In this study, the effect of arsenite on cell cycle regulation and the mechanisms that contribute to this effect were investigated. Treatment of the cells with arsenite suppressed cell proliferation and reduced cell viability in a dose- or time-dependent manner. Analysis of cell cycle profile and cell cycle regulatory proteins indicated that arsenite arrested the cell cycle at G(2)/M phase, partially through induction of cell division cycle 25 (Cdc25) isoform C (Cdc25C) degradation via ubiquitin-proteasome pathways. Mutation of the putative KEN box within the region 151 to 157 of human Cdc25C or treatment of the cells with a peptide competitor encompassing the KEN box partially inhibited arsenite-induced ubiquitination of Cdc25C. Thus, these results indicate that the regulated ubiquitination of Cdc25C may be involved in the arsenite-induced proteolytic down-regulation of Cdc25C activity in the G(2)/M phase of the cell cycle and suggest a link between cell cycle and the carcinogenic effects of arsenite.

Acetylcysteine↗

PubMed2002

Regulation of protein and vesicle trafficking at the apical membrane of epithelial cells.

The characterization of endocytotic and post-endocytotic trafficking pathways at the apical membrane of epithelial cells presents a potential avenue for the identification of targets to modulate the initial stages of absorption and transepithelial transport of macromolecules. In addition, it is becoming increasingly clear that the activity of a number of apical membrane transporters is acutely regulated by vesicular trafficking. The gastric HCl-secreting parietal (oxyntic) cell is a model system to characterize an apical membrane vesicular trafficking pathway and its relationship to the regulation of the function of the gastric proton pump. The subapical tubulovesicular compartment of the parietal cell is highly enriched in the H,K-ATPase and is a key endosomal-like system in the apical membrane recycling pathway. In the process of cataloging the proteins that interact with the H,K-ATPase and tubulovesicles, we have identified novel components that may regulate protein sorting through this compartment and candidate linker proteins between the vesicular trafficking machinery and the cytoskeleton. One protein associated with H,K-ATPase-rich tubulovesicles is the nonreceptor tyrosine kinase c-src, identified by a screen for dynamin-binding proteins. The tyrosine kinase is active, as it can tyrosine-phosphorylate tubulovesicular proteins in vitro. One of the tyrosine-phosphorylated proteins of M(r) 100 kDa may be the H,K-ATPase itself, or a protein in a complex with the H,K-ATPase that is stable to dissociation by nonionic detergents. By virtue of its association with tubulovesicular membranes, c-src may regulate the trafficking and/or activity of the H,K-ATPase. A second protein identified by a screen for dynamin-binding proteins is the protein lasp-1. Lasp-1, through its modular protein structure, may bind to dynamin and to the actin cytoskeleton, thus linking the vesicular trafficking machinery with the cytoskeleton. These two examples illustrate the utility of the parietal cell in the biochemical characterization of components potentially involved in the regulation of apical membrane trafficking pathways.

Animals↗

PubMed2002

Intracellular angiotensin II increases the long isoform of PDGF mRNA in rat hepatoma cells.

Our recent published studies suggest that angiotensin II (AII), generated and retained intracellularly, enhances growth of H4-II-E-C3 rat hepatoma cells, an average of 33%. Proliferation conferred by introduction of a plasmid [ Ang(-S)Exp/pSVL ] encoding a signal sequence-depleted angiotensinogen [Ang(-S)Exp] into these cells (which we have shown possess ACE and renin mRNAs) is mediated, at least in part, by enhanced PDGF-A chain mRNA production and protein secretion. The mitogenic effect is inhibited by losartan suggesting that it involves AII interaction with an AT(1)-like receptor. Introduction of anti-AII antibodies into the medium of these transfected cells has no effect upon growth of the cells, suggesting that AII is retained by the cells and that intracellular AII is growth stimulatory. In the present study, we sought to further characterize the intracellular localization and mode of action of Ang(-S)Exp. Consistent with our expectations, we now show that a fusion product of Ang(-S)Exp with green fluorescent protein [Ang(-S)Exp/EGFP], generated from an expression plasmid, is abundant and primarily cytoplasmic. Wild-type angiotensinogen/EGFP, in contrast, is only detectable following a cold-block (which acts to enhance folding-kinetics and slow secretion) and is largely restricted to the secretory pathway. We further show, using semi-quantitative RT/PCR that the long isoform of PDGF mRNA is elevated in Ang(-S)Exp transfected cells and in AII-treated naive cells but not in losartan-treated Ang(-S)Exp transfected cells. We identify C-terminal amidation recognition sites within the long-form protein (that are not present in the short-form) and show that these cells possess PAM (amidating enzyme precursor) and carboxypeptidase E mRNAs (the corresponding proteins of which are sufficient for amidation). Inhibitors of amidation inhibit growth of naive and Ang(-S)Cntr/ pSVL -transfected cells (2.6-fold for phenylbutenoic acid and 3.5-fold for disulfiram treatment) but more profoundly inhibit growth of Ang(-S)Exp/pSVL -transfected cells (6.7-fold for phenylbutenoic acid and 13-fold for disulfiram). In conclusion, these data confirm that signal sequence-depleted Ang(-S)Exp is retained within cells and is largely cytoplasmic. Because C-terminal amidation is absolutely required for full biological potency of a number of peptide hormones (including oxytocin, gastrin and calcitonin), we postulate that growth effects of both intracellular AII and exogenous AII can be conferred by PDGF long-form, possibly through an amidation-dependent mechanism.

Amides↗

PubMed2002

MAPKs mediate S phase arrest induced by vanadate through a p53-dependent pathway in mouse epidermal C141 cells.

Mitogen-activated protein (MAP) kinases play an important role in mediation of the signal transduction pathway in cellular response to genotoxic stress. Cell growth arrest is considered as an early stage in response to the genotoxic stress. p53 is well-known as a tumor suppression gene involved in both cell growth arrest and apoptosis. The present study investigated the involvement of MAP kinases in vanadate-induced cell growth arrest and the relationship of p53. DNA content analysis showed that vanadate-induced S phase arrest is time- and dose-dependent in p53 wild-type C141 cells but not in p53-deficient C141 cells. Western blotting results indicated that vanadate caused an inactivation of p-cdk2 at Thr160, which is an important kinase for the progression of S phase, and an increase in expression of p21, which is a key for S phase arrest. In p53-deficient cells, vanadate did not induce any observable change in p21 or p-cdk2 level. In addition, vanadate up-regulated phospho-p38 and ERK, two members of MAP kinases. At the same time, vanadate increased the p53 activity as measured by luciferase assay. Addition of PD98059 and SB202190, inhibitors of ERK and p38, respectively, decreased vanadate-induced S phase arrest, reduced p21 levels, restored activation of p-cdk2, and decreased p53 activity. The study demonstrated that vanadate-induced S phase arrest is mediated by both ERK and p38 in a p53-dependent pathway.

Animals↗

PubMed2002

Vanadate induces G2/M phase arrest in p53-deficient mouse embryo fibroblasts.

Vanadium compounds exert potent toxic and carcinogenic effects on a wide variety of biological systems. The mechanisms involved in their toxicity and carcinogenesis require investigation. Cell growth arrest and its regulation are important mechanisms in maintaining genomic stability and integrity in response to environmental stress. The p53 tumor suppressor plays a central role in the regulation of the normal cell cycle. To investigate the role of p53 in vanadate-induced cell growth arrest and its regulation, two cell lines--normal mouse embryo fibroblasts [p53(+/+)] and p53-deficient mouse embryo fibroblasts [p53(-/-)],--were used in this study. Flow cytometry was used to analyze cell growth arrest at G0/G1, S, or G2/M phase. Western blotting analysis was performed to determine several cell growth regulatory proteins. The results showed that in p53(-/-) cells vanadate induced G2/M phase arrest in a dose- and time-dependent manner without alteration of S phase. In p53(+/+) cells, vanadate treatment increased the S phase with no significant change in the G2/M phase. Furthermore, Western blotting results showed that in p53(-/-) cells vanadate caused cdc25C degradation and activation of phospho-cdc2 without alteration of the p21 level. In p53(+/+) cells, vanadate increased the expression of p21 and degraded cdc25A instead of cdc25C without any effect on cdc2. These results demonstrate that vanadate induced G2/M phase arrest in p53-deficient mouse embryo fibroblasts, and promoted S phase entry in p53 wild-type mouse embryo fibroblasts.

Animals↗