Promoter- and Cell-Specific Transcriptional Transactivation by the Kaposi's Sarcoma-Associated Herpesvirus ORF57/Mta Protein

Abstract

The Kaposi's sarcoma-associated herpesvirus (KSHV) Mta protein, encoded by open reading frame 57, is a transactivator of gene expression that is essential for productive viral replication. Previous studies have suggested both transcriptional and posttranscriptional roles for Mta, but little is known regarding Mta's transcriptional function. In this study, we demonstrate that Mta cooperates with the KSHV lytic switch protein, Rta, to reactivate KSHV from latency, but Mta has little effect on reactivation when expressed alone. We demonstrate that the Mta and Rta proteins are expressed with similar but distinct kinetics during KSHV reactivation. In single-cell analyses, Mta expression coincides tightly with progression to full viral reactivation. We demonstrate with promoter reporter assays that while Rta activates transcription in all cell lines tested, Mta's ability to transactivate promoters, either alone or synergistically with Rta, is cell and promoter specific. In particular, Mta robustly transactivates the nut-1/PAN promoter independently of Rta in 293 and Akata-31 cells. Using nuclear run-on assays, we demonstrate that Mta stimulates transcriptional initiation in 293 cells. Rta and Mta physically interact in infected cell extracts, and this interaction requires the intact leucine repeat and central region of Rta in vitro. We demonstrate that Mta also binds to the nut-1/PAN promoter DNA in vitro and in infected cells. An Mta mutant with a lesion in a putative A/T hook domain is altered in DNA binding and debilitated in transactivation. We propose that one molecular mechanism of Mtamediated transactivation is a direct effect on transcription by direct and indirect promoter association.Reactivation of Kaposi's-sarcoma associated herpesvirus (KSHV) (also known as human herpesvirus 8) from latency is a crucial step in Kaposi's sarcoma development (7,11,27,43,65,76,81,87,104). It is likely that reactivation of KSHV contributes to Kaposi's sarcoma development by facilitating dissemination of the virus from its B-cell reservoir and permitting the expression of lytic cycle genes with direct roles in cancer progression. Most of the candidate pathogenic genes of KSHV (encoding proteins with growth-deregulatory and immunomodulatory functions) are expressed in the delayed early class of the lytic gene expression program (8,14,17,25,26,28,30,41,46,49,75,86,94). Therefore, a complete understanding of KSHV pathogenesis demands elucidation of the mechanisms that regulate viral reactivation and progression through the lytic cycle.The members of the Herpesviridae all encode multiple lytic cycle transactivators that cooperate to promote viral replication. For herpes simplex virus type 1 (HSV-1) infection, the two essential transactivating proteins are called infected cell protein 4 (ICP4) and ICP27. ICP4 is a transcriptional transactivator necessary for activation of early and late genes (80, 103). ICP27 is a posttranscriptional and transcriptional activator that stimulates the switch from early-to late-gene expression (66,67,...

Spironolactone blocks Epstein–Barr virus production by inhibiting EBV SM protein function

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Rhesus Monkey Rhadinovirus ORF57 Induces gH and gL Glycoprotein Expression through Posttranscriptional Accumulation of Target mRNAs

“…The transcriptional activation of these promoters could be accomplished by ORF57 protein alone, or it could be further enhanced by the cooperation with the ORF50 gene product (also known as RTA) (20,28,33). However, as the name MTA (mRNA transcript accumulation) in KSHV implies, a posttranscriptional mechanism of activation by ORF57 orthologues has been studied and documented most extensively (4,14,21,29).…”

“…KSHV ORF57 activates the de novo expression of viral genes in a promoter-specific manner, including genes for oriLyt (L), kaposin, ORF57, and thymidine kinase (TK) (33). The transcriptional activation of these promoters could be accomplished by ORF57 protein alone, or it could be further enhanced by the cooperation with the ORF50 gene product (also known as RTA) (20,28,33).…”

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