Abstract

The interplay between influenza virus and host factors to support the viral life cycle is well documented. Influenza A virus (IAV) proteins interact with an array of cellular proteins and hijack host pathways which are at the helm of cellular responses to facilitate virus invasion. The multifaceted nature of the ubiquitination pathway for protein regulation makes it a vulnerable target of many viruses including IAV. To this end we conducted a yeast two-hybrid screen to search for cellular ubiquitin ligases important for influenza virus replication. We identified host protein, RING finger protein 43 (RNF43), a RING-type E3 ubiquitin ligase, as a novel interactor of nucleoprotein (NP) of IAV and an essential partner to induce NP-driven p53-mediated apoptosis in IAV-infected cells. In this study, we demonstrate that IAV leads to attenuation of RNF43 transcripts and hence its respective protein levels in the cellular milieu whereas in RNF43 depleted cells, viral replication was escalated several folds. Moreover, RNF43 polyubiquitinates p53 which further leads to its destabilization resulting in a decrease in induction of the p53 apoptotic pathway, a hitherto unknown process targeted by NP for p53 stabilization and accumulation. Collectively, these results conclude that NP targets RNF43 to modulate p53 ubiquitination levels and hence causes p53 stabilization which is conducive to an enhanced apoptosis level in the host cells. In conclusion, our study unravels a novel strategy adopted by IAV for utilizing the much conserved ubiquitin proteasomal pathway.

Highlights

  • The evolution of host–microbe interaction is mediated through the orchestration of different viral and host signaling pathways

  • RNF43 is a recently identified member of the RING finger family of ubiquitin ligases and has been implicated to be overexpressed in human colorectal and hepatocellular carcinomas with anti-apoptotic and growth-promoting effects that interacts with HAP95 and NEDL1, an upstream effector of p53.15–19 A crucial mediator of cellular apoptosis, p53 is present in a latent form in unstressed cells and is regulated through various posttranslational modifications like phosphorylation, ubiquitination, sumolyation, neddylation, methylation, acetylation and glycosylation of p53 polypeptide.[20,21,22,23]

  • Accumulation of p53 in Influenza A virus (IAV)-infected cells has been demonstrated,[24,25,26] it is still not clear whether IAV-induced accumulation of p53 is correlated with its activation and consecutive transactivation of its target genes that could in turn induce apoptosis in infected cells that is considered to be a peculiar feature of IAV pathogenesis.[27,28,29,30,31]

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Summary

Introduction

The evolution of host–microbe interaction is mediated through the orchestration of different viral and host signaling pathways. RNF43 is a recently identified member of the RING finger family of ubiquitin ligases and has been implicated to be overexpressed in human colorectal and hepatocellular carcinomas with anti-apoptotic and growth-promoting effects that interacts with HAP95 and NEDL1, an upstream effector of p53.15–19 A crucial mediator of cellular apoptosis, p53 is present in a latent form in unstressed cells and is regulated through various posttranslational modifications like phosphorylation, ubiquitination, sumolyation, neddylation, methylation, acetylation and glycosylation of p53 polypeptide.[20,21,22,23] accumulation of p53 in IAV-infected cells has been demonstrated,[24,25,26] it is still not clear whether IAV-induced accumulation of p53 is correlated with its activation and consecutive transactivation of its target genes that could in turn induce apoptosis in infected cells that is considered to be a peculiar feature of IAV pathogenesis.[27,28,29,30,31]. We have shown that IAV attenuates RNF43 on interacting with NP stabilizing p53 protein and induces p53 signaling and apoptosis in the host cells

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