A role of hypoxia-inducible factor 1 alpha in Mouse Gammaherpesvirus 68 (MHV68) lytic replication and reactivation from latency.

Author summary The host oxygen-sensing machinery, including the HIF1 alpha pathway, is important during the viral life cycle of oncogenic gammaherpesviruses such as KSHV and EBV. However, due to the host specificity, the effects of HIF1 alpha in herpes biology is limited to studies with in vitro systems. Here, we study the role of HIF1 alpha using the mouse gammaherpesvirus 68 (MHV68) that readily infects laboratory mice. We demonstrate that MHV68 infection upregulates HIF1 alpha during replication and inactivation of HIF1 alpha transcriptional activity significantly decreased viral gene expression, which results in impaired virus production in vitro. In vivo deletion of HIF1 alpha impaired viral expansion during acute infection and affected reactivation from latency. These results show the importance of the interplay with the oxygen-sensing machinery in gammaherpesvirus infection and pathogenesis, placing the MHV68 mouse model as a unique platform to gain insight into this important aspect of oncogenic gammaherpesviruses biology and to test HIF1 alpha targeted therapeutics. The hypoxia-inducible factor 1 alpha (HIF1 alpha) protein and the hypoxic microenvironment are critical for infection and pathogenesis by the oncogenic gammaherpesviruses (gamma HV), Kaposi' Sarcoma-associated Herpes Virus (KSHV) and Epstein-Barr virus (EBV). However, understanding the role of HIF1 alpha during the virus life cycle and its biological relevance in the context of host pathogenesis has been challenging due to the lack of animal models for human gamma HV. To study the role of HIF1 alpha, we employed the murine gammaherpesvirus 68 (MHV68), a rodent pathogen that readily infects laboratory mice. We show that MHV68 infection induces HIF1 alpha protein and HIF1 alpha-responsive gene expression in permissive cells. siRNA silencing or drug-inhibition of HIF1 alpha reduce virus production due to a global downregulation of viral gene expression. Most notable was the marked decrease in many viral genes bearing hypoxia-responsive elements (HREs) such as the viral G-Protein Coupled Receptor (vGPCR), which is known to activate HIF1 alpha transcriptional activity during KSHV infection. We found that the promoter of MHV68 ORF74 is responsive to HIF1 alpha and MHV-68 RTA. Moreover, Intranasal infection of HIF1 alpha(LoxP/LoxP) mice with MHV68 expressing Cre- recombinase impaired virus expansion during early acute infection and affected lytic reactivation in the splenocytes explanted from mice. Low oxygen concentrations accelerated lytic reactivation and enhanced virus production in MHV68 infected splenocytes. Thus, we conclude that HIF1 alpha plays a critical role in promoting virus replication and reactivation from latency by impacting viral gene expression. Our results highlight the importance of the mutual interactions of the oxygen-sensing machinery and gammaherpesviruses in viral replication and pathogenesis.