Kaposi’s sarcoma-associated herpesvirus vIRF2 protein utilizes an IFN-dependent pathway to regulate viral early gene expression

Abstract

Kaposi’s sarcoma-associated herpesvirus (KSHV; human herpesvirus 8) belongs to the subfamily of Gammaherpesvirinae and is the etiological agent of Kaposi’s sarcoma as well as of two lymphoproliferative diseases: primary effusion lymphoma and multicentric Castleman disease. The KSHV life cycle is divided into a latent and a lytic phase and is highly regulated by viral immunomodulatory proteins which control the host antiviral immune response. Among them is a group of proteins with homology to cellular interferon regulatory factors, the viral interferon regulatory factors 1–4. The KSHV vIRFs are known as inhibitors of cellular interferon signaling and are involved in different oncogenic pathways. Here we characterized the role of the second vIRF protein, vIRF2, during the KSHV life cycle. We found the vIRF2 protein to be expressed in different KSHV positive cells with early lytic kinetics. Importantly, we observed that vIRF2 suppresses the expression of viral early lytic genes in both newly infected and reactivated persistently infected endothelial cells. This vIRF2-dependent regulation of the KSHV life cycle might involve the increased expression of cellular interferon-induced genes such as the IFIT proteins 1, 2 and 3, which antagonize the expression of early KSHV lytic proteins. Our findings suggest a model in which the viral protein vIRF2 allows KSHV to harness an IFN-dependent pathway to regulate KSHV early gene expression.

One Step Ahead: Herpesviruses Light the Way to Understanding Interferon-Stimulated Genes (ISGs)

“…In line with these findings, vIRF2 has been recently described to manipulate the innate immune response. vIRF2 regulates the expression of 51 genes known to be involved in innate or intrinsic defences, boosting the formation of the antiviral cellular state to restrict KSHV early lytic protein expression and promote latency (Koch et al, 2019). This is an intriguing illustration of the fine-tuned balance between herpesviruses and their host, which dictates the outcome of the infection course.…”

USP7-Dependent Regulation of TRAF Activation and Signaling by a Viral Interferon Regulatory Factor Homologue

“…It has been reported very recently (during conclusion of our own studies here) that vIRF-2 inhibits lytic gene expression during lytic reactivation in and de novo infection of endothelial cells and inhibits virus production in the former (39); these are the only published phenotypic studies of vIRF-2. It is notable that in the context of PEL cells, naturally infected with HHV-8, vIRF-2 transcripts and protein have been detected in both latency and lytic infection (24,39,48). We first tested for potential effects of vIRF-2 depletion on the growth of latently infected PEL cells.…”

“…9B). These data demonstrated suppression of lytic replication by vIRF-2, reflective of the effect on lytic replication of vIRF-3 (27), which is, like vIRF-2, expressed in latently infected PEL cells (24,39,48). To investigate the generality of the vIRF-2 lytic phenotype and to facilitate biological analysis of vIRF-2-USP7 interaction specifically, bacmid (BAC16)-based recombinant viruses were generated containing either mutation of the initiator ATG codon (ATG to TTG) or of codon 247 to specify USP7-refractory vIRF-2.S 247 A (Fig.…”

“…1A). To verify interactions of endogenous proteins, a USP7 immunoprecipitation experiment was carried out using extracts of TRExBCBL1-RTA PEL cells, doxycycline inducible for the immediate-early lytic cycle inducer RTA (38); vIRF-2 was identified in the immunoprecipitates from both latently infected and lytically reactivated (doxycyclinetreated) cultures, using vIRF-2-specific antibody (39) for immunoblotting ( Fig. 1B).…”

“…To investigate the generality of the vIRF-2 lytic phenotype and to facilitate biological analysis of vIRF-2-USP7 interaction specifically, bacmid (BAC16)-based recombinant viruses were generated containing either mutation of the initiator ATG codon (ATG to TTG) or of codon 247 to specify USP7-refractory vIRF-2.S 247 A (Fig. 9C); these changes were reverted to wild-type sequences to generate control repaired (R) viruses to ensure that no other phenotypically significant changes occurred during the bacterially based genetic manipulations of the BAC16 genome (39,49) (see Materials and Methods). The gross integrities of each viral genome was checked by restriction analysis (Fig.…”