Yoon Ki Kim

E2 of Hepatitis C Virus Inhibits Apoptosis

Hepatitis C virus (HCV) is the major causative agent of chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma, and can be involved in very long chronic infections up to 30 years or more. Therefore, it has been speculated that HCV possesses mechanisms capable of modulating host defense systems such as innate and adaptive immunity. To investigate this virus-host interaction, we generated HCV replicons containing various HCV structural proteins and then analyzed the sensitivity of replicon-containing cells to the apoptosis-inducing agent, TRAIL. TRAIL-induced apoptosis was monitored by cleavage of procaspase-3 and procaspase-9 as well as that of their substrate poly(ADP-ribose) polymerase. TRAIL-induced apoptosis was inhibited in cells expressing HCV E2. Moreover, expression of HCV E2 enhanced the colony forming efficiency of replicon-containing cells by 25-fold. Blockage of apoptosis by E2 seems to be related to inhibition of TRAIL-induced cytochrome c release from the mitochondria. Based on these results, we propose that E2 augments persistent HCV infection by blocking host-induced apoptosis of infected cells.

Polypyrimidine tract-binding protein interacts with HnRNP L

Polypyrimidine tract‐binding protein (PTB) is involved in pre‐mRNA splicing and internal ribosomal entry site (IRES)‐dependent translation. In order to identify cellular protein(s) interacting with PTB, we performed a yeast two‐hybrid screening. Heterogeneous nuclear ribonucleoprotein L (hnRNP L) was identified as a PTB‐binding protein. The interaction between PTB and hnRNP L was confirmed in an in vitro binding assay. Both PTB and hnRNP L were found to localize in the nucleoplasm, excepting the nucleoli, in HeLa cells by the green fluorescent protein (GFP)‐fused protein detection method. The N‐terminal half of PTB (aa 1–329) and most of hnRNP L (aa 141–558) is required for the interaction between PTB and hnRNP L.

Continuous heat shock enhances translational initiation directed by internal ribosomal entry site.

Many cellular mRNAs contain internal ribosomal entry sites (IRES) that become functional under conditions of cellular stress, when the rate of protein synthesis for most cellular mRNA is reduced. Internal ribosomal entry increases in response to hypoxia, cell differentiation, apoptosis, gamma irradiation, and heat shock. Heat shock is the principal cellular stress in which general cap-dependent translation is inhibited. On the other hand, heat shock induces the preferential translation of a small class of mRNA, called heat shock protein (HSP) mRNAs, which probably occurs because little or no eIF4F activity is required for their translation. In this study, we found that continuous heat stress enhances expression of the heat shock protein BiP at the level of translation. Interestingly, heat stress also enhanced the viral IRES-dependent translation of encephalomyocarditis virus and hepatitis C virus but not poliovirus. Although several BiP inducers increased BiP protein expression, BiP IRES-dependent translation was enhanced only during heat shock, suggesting that heat shock is a specific inducer for BiP IRES-dependent translation. Taken together, these results indicate that the mechanism of IRES-dependent translation can be used during heat shock and suggest that this translational mechanism may be critical to the survival and proliferation of cells under stress.

La protein is required for efficient translation driven by encephalomyocarditis virus internal ribosomal entry site

Translation of internal ribosomal entry site (IRES)-dependent mRNAs is mediated by RNA-binding proteins as well as canonical translation factors. In order to elucidate the roles of RNA-binding proteins in IRES-dependent translation, the role of polypyrimidine tract-binding protein (PTB) and La protein in encephalomyocarditis virus (EMCV) IRES-dependent translation was investigated. PTB was required for efficient EMCV IRES-driven translation but, intriguingly, an excess of PTB suppressed it. Such a translational suppression by surplus PTB was relieved by addition of La protein. A possible role for La protein in IRES-dependent translation is discussed.