Quantitative analysis of the bidirectional viral G-protein-coupled receptor and lytic latency-associated nuclear antigen promoter of Kaposi's sarcoma-associated herpesvirus.

ABSTRACT: Kaposi's sarcoma-associated herpesvirus (KSHV) establishes sustained latent persistence in susceptible cells. This is dependent on the latency-associated nuclear antigen (LANA). Understanding how LANA transcription is regulated thus aids our fundamental understanding of KSHV biology. Two hundred ninety-four base pairs are sufficient to regulate LANA transcription in response to the viral RTA protein and RBPj?. The same region controls K14/viral G-protein-coupled receptor (vGPCR) transcription in the opposite direction. We used a quantitative analysis in conjunction with specific nucleotide substitutions and defined gain-of-function and loss-of-function RTA mutants to dissect this region. We used a bidirectional reporter driving red and green luciferase to study the LANApi and K14p promoters simultaneously. This established that LANApi/K14p functions as a canonical bidirectional promoter. Both were TATA dependent. K14p was favored by ?50-fold in this context. Eliminating the distal LANApi TATA box increased maximal output and lowered the induction threshold (T) of K14p even further. Two RBPj? binding sites were independently required; however, at high concentrations of RTA, direct interactions with an RTA-responsive element (RRE) could complement the loss of one RBPj? binding site. Intracellular Notch (ICN) was no longer able to activate RBPj? in the viral context. This suggests a model whereby KSHV alters ICN-RBPj? gene regulation. When the architecture of this pair of head-to-head RBPj? binding sites is changed, the sites now respond exclusively to the viral transactivator RTA and no longer to the host mediator ICN.

SUBMITTER: Hilton IB 

PROVIDER: S-EPMC3446609 | biostudies-literature | 2012 Sep

REPOSITORIES: biostudies-literature