Abstract
Kaposi’s sarcoma-associated herpesvirus (KSHV) is the causative agent of Kaposi’s sarcoma, primary effusion lymphoma, and multicentric Castleman’s disease. KSHV establishes a life-long infection in its host and alternates between a latent and lytic infection state. During lytic infection, lytic-related genes are expressed in a temporal manner and categorized as immediate early, early, and late gene transcripts. ORF34 is an early-late gene that interacts with several viral transcription-associated factors, however its physiological importance remains poorly understood. Here, we investigated the role of ORF34 during KSHV infection by generating ORF34-deficient KSHV, using a bacterial artificial chromosome system. Our results reveal that ORF34-deficient KSHV exhibited significantly attenuated late gene expression and viral production but did not affect viral DNA replication. ORF34 interacted with transcription factors ORF18, ORF24, ORF31, and ORF66, and a novel ORF34-interaction partner, ORF23. The C-terminal region of ORF34 was important for interaction with ORF24 and viral production. Our data support a model, in which ORF34 serves as a hub for recruiting a viral transcription complex to ORF24 to promote late viral gene expression.
Highlights
Kaposi’s sarcoma-associated herpesvirus (KSHV) belongs to the gamma–herpesvirus family
Our study reveals the importance of ORF34 for KSHV gene expression through the formation of the predicted KSHV-PIC
We revealed that viral production of inducible (Tet-on) RTA/ORF50-expressing Vero (iVero)-ΔORF34 cells are rescued by exogenous full length ORF34 expression but not by the expression of ORF34 C-terminal deletion mutants (Fig. 7)
Summary
Kaposi’s sarcoma-associated herpesvirus (KSHV) belongs to the gamma–herpesvirus family. The alternation of KSHV between lytic replication and latency depends on the IE-gene RTA/ORF50, which triggers transcriptional activation of E genes associated with viral DNA replication[13, 14]. BcRF1, a core factor of EBV-PIC and virus homolog of TATA box binding protein (TBP), is essential for L gene expression[17]. KSHV ORF24 (a homolog of EBV BcRF1) binds to RNAPII22, ORF34 protein[23], and TATT sequences in the L gene promoter (K8.1 and ORF57 promoter)[22, 24]. ORF34 appears to serve as a scaffold for viral TBP ORF24 and PIC components (ORF18, 30, 31, 34, and 66), its functional role during KSHV replication remains unknown. We show that ORF34 binds with ORF23, a novel ORF34-interaction partner, and regulates KSHV L gene expression
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