Involvement of the Cellular Protein SIAH-1 in the Infection Cycle of Kaposi’s Sarcoma-associated Herpesvirus

Kaposi`s sarcoma-associated herpesvirus (KSHV) is an oncogenic human virus which is etiologically linked with Kaposi’s sarcoma, primary effusion and certain forms of Multicentric Castleman`s disease. The open reading frame 45 (ORF45) protein, encoded by the KSHV genome, is an immediate-early lytic protein which functions during the early and late stages of infection and is included in the outer layer of the virion tegument. Our lab has previously identified an interaction between ORF45 and the cellular ubiquitin E3 ligase protein SIAH-1 which targets ubiquitylation and degradation of ORF45.

In the present work we examined the significance of the interaction between ORF45 and SIAH-1 during de novo infection by using recombinant KSHV genomes encoding mCherry-fluorescent protein-tagged wild-type and mutated ORF45 that fails to interact with SIAH-1. We hypothesized that the degradation of ORF45 by SIAH-1 may represent an antiviral cellular response to reduce ORF45 levels or alternatively may promote the dissociation of ORF45 from the capsids to allow progression of the infection cycle. By using fluorescent microscopy and 3-D image analyses, we detected similar dissociation kinetics of wild-type and mutated ORF45 from the capsids upon infection; yet, the mutated ORF45 accumulated to higher levels as compared with wild-type ORF45. Thus, our results indicate that SIAH-1 is not involved in the uncoating of ORF45, but instead functions to mediate the degradation of ORF45 after its release into the cytoplasm. As ORF45 inhibits the interferon response, our results suggest that SIAH-1 reduces ORF45 levels which in-turn increases the cellular anti-viral response.