Hepatitis C Virus Core Protein and Cellular Protein HAX-1 Promote 5-Fluorouracil-Mediated Hepatocyte Growth Inhibition

Abstract

Hepatitis C virus (HCV) often causes chronic infection and may lead to hepatocellular carcinoma (HCC).We have shown previously that HCV core protein has pleiotropic functions, including transcriptional regulation of a number of cellular genes, although the mechanism for gene regulation remains unclear. In this study, a mammalian two-hybrid screen identified a novel binding partner, HS1-associated protein X-1 (HAX-1), for HCV core protein from a human liver cDNA library. An association between HAX-1 and HCV core protein was further verified by confocal microscopy and coimmunoprecipitation in HepG2 cells expressing HCV core or full-length (FL) gene. Both HCV core protein and a chemotherapeutic agent for HCC, 5-flouorouracil (5-FU), are known to modulate p53. We examined here whether an association between core and HAX-1 has any functional relevance to p53 modulation in 5-FU-treated cells. For this, the role of HAX-1 on 5-FU treatment was examined in HepG2 cells expressing HCV core or FL gene using cell proliferation, p53 expression, and caspase activation analysis. Cells expressing HCV-core or FL gene were more susceptible to 5-FU-induced growth inhibition than control cells, whereas cell survival was enhanced after suppression of HAX-1 by small interfering RNA. Further, 5-FU-mediated p53 expression was reduced with concurrent HAX-1 suppression in core-or polyprotein-expressing cells compared to control HepG2 cells, and caspase-2 and -7 activities were diminished. On the other hand, HCV core protein did not play a detectable role in 5-FU-mediated caspase-7 activation in the absence of functional p53 in Hep3B or Huh-7 cells. These observations underscore an association between HCV core and HAX-1, which promotes 5-FU mediated p53-dependent caspase-7 activation and hepatocyte growth inhibition.

The classical swine fever virus N-terminal protease Npro binds to cellular HAX-1

“…N pro may orchestrate this redistribution of HAX-1 to the ER during infection to increase cellular resistance to apoptosis. A similar pattern of co-localization has been reported for other proteins which bind HAX-1 including the Vpr protein of HIV type 1, the K15 protein of KSHV, HCV core protein and PLN (Banerjee et al, 2009;Sharp et al, 2002;Vafiadaki et al, 2007;Yedavalli et al, 2005). Importantly, the redistribution of HAX-1 to the ER in the presence of PLN correlated with stronger resistance to apoptosis (Vafiadaki et al, 2007).…”

“…core protein (Banerjee et al, 2009;Dufva et al, 2001;Forsman et al, 2008;Kawaguchi et al, 2000;Matsuda et al, 2003;Modem & Reddy, 2008;Sharp et al, 2002;Yedavalli et al, 2005). These studies point to virus-mediated antagonism of host-cell apoptotic response.…”

“…Banerjee et al (2009) showed that HepG2 cells expressing HCV core protein were sensitized to undergo apoptosis when treated with the chemotherapeutic agent 5-fluorouracil. They suggested that sensitization is probably linked with the association of HCV core protein and HAX-1, and showed that cell death occurred via a mechanism involving p53 and activation of caspase-2 and -7, in association with HAX-1.…”

Hepatitis C Virus Impairs Natural Killer Cell-Mediated Augmentation of Complement Synthesis

“…Stable transfectants of Huh7.5 cells were maintained in Dulbecco's modified Eagle medium containing 10% fetal calf serum and selection antibiotic. The expression of HCV proteins (core or NS5A) was verified by immunofluorescence or Western blot analysis (18,19). Immortalized human hepatocytes (IHH) were generated and maintained as previously describedsaline (PBS) containing 0.5% bovine serum albumin (BSA) for 30 min at room temperature.…”

Pathogenic mechanisms and clinical implications of congenital neutropenia syndromes

“…In line with an important function for HAX1 in cellular homeostatis is the observation that a number of viruses such as Epstein-Barr virus (EBV), human herpesvirus 8 (HHV-8), HIV and hepatitis C virus (HCV), which partially establish latency, code for HAX-1 interacting proteins, even though the mechanistic consequences of these interactions remain elusive [151][152][153][154][155][156].…”