tetracycline and pyrazolanthrone

Hepatitis C virus (HCV) non-structural protein 3 (NS3) has been shown to affect cellular functions and is thought to contribute to the development of HCV-related hepatocarcinogenesis. In this study, we delineated part of the mechanisms whereby NS3 protein stimulates cell growth in liver (HepG2) and non-liver (HeLa) cells. The expression of NS3 protein enhanced cell growth, c-jun NH(2)-terminal kinase (JNK) activation, DNA binding activities of the transcription factors AP-1 and ATF-2, and c-jun expression, but not the activation of extracellular signal-regulated kinase (ERK) or p38(MAPK). Whereas co-expression of NS3 with its cofactor NS4A inhibited NS3-mediated cell growth without to influence NS3-mediated JNK activation, or to affect the basal activities of ERK or p38(MAPK). Pre-treatment of NS3 protein-expressing cells with JNK inhibitor, SP600125, abolished activation of AP-1 and ATF-2 and inhibited c-jun expression and induced cell growth, suggesting that JNK activation is essential for the stimulation of NS3-mediated cell growth.

Topics: Activating Transcription Factor 2; Anthracenes; Base Sequence; Cell Division; Cell Line; Cyclic AMP Response Element-Binding Protein; DNA, Viral; Enzyme Inhibitors; Extracellular Signal-Regulated MAP Kinases; Gene Expression; Genes, Viral; HeLa Cells; Hepacivirus; Humans; JNK Mitogen-Activated Protein Kinases; MAP Kinase Signaling System; p38 Mitogen-Activated Protein Kinases; Tetracycline; Transcription Factor AP-1; Transcription Factors; Viral Nonstructural Proteins

Hsp70 overexpression can protect cells from stress-induced apoptosis. Our previous observation that Hsp70 inhibits cytochrome c release in heat-stressed cells led us to examine events occurring upstream of mitochondrial disruption. In this study we examined the effects of heat shock on the proapoptotic Bcl-2 family member Bax because of its central role in regulating cytochrome c release in stressed cells. We found that heat shock caused a conformational change in Bax that leads to its translocation to mitochondria, stable membrane association, and oligomerization. All of these events were inhibited in cells that had elevated levels of Hsp70. Hsp70 did not physically interact with Bax in control or heat-shocked cells, indicating that Hsp70 acts to suppress signals leading to Bax activation. Hsp70 inhibited stress-induced JNK activation and inhibition of JNK with SP600125 or by expression of a dominant negative mutant of JNK-blocked Bax translocation as effectively as Hsp70 overexpression. Hsp70 did not protect cells expressing a mutant form of Bax that has constitutive membrane insertion capability or cells treated with a small molecule activator of apoptosome formation, indicating that it is unable to prevent cell death after mitochondrial disruption and caspase activation have occurred. These results indicate that Hsp70 blocks heat-induced apoptosis primarily by inhibiting Bax activation and thereby preventing the release of proapoptotic factors from mitochondria. Hsp70, therefore, inhibits events leading up to mitochondrial membrane permeabilization in heat-stressed cells and thereby controls the decision to die but does not interfere with cell death after this event has occurred.

Topics: Anthracenes; Apoptosis; bcl-2-Associated X Protein; Blotting, Western; Caspases; Cell Death; Cell Line, Tumor; Cell Membrane; Cross-Linking Reagents; Cytochromes c; Cytosol; Dimerization; Enzyme Activation; Extracellular Signal-Regulated MAP Kinases; Genes, Dominant; HeLa Cells; Hot Temperature; HSP70 Heat-Shock Proteins; Humans; Immunoprecipitation; MAP Kinase Kinase 4; Microscopy, Fluorescence; Mitochondria; Mutation; Protein Binding; Protein Conformation; Protein Transport; Proto-Oncogene Proteins c-bcl-2; T-Lymphocytes; Temperature; Tetracycline; Time Factors