Identification of a cellular protein that interacts and synergizes with the RTA (ORF50) protein of Kaposi's sarcoma-associated herpesvirus in transcriptional activation

Lytic reactivation of Kaposi's sarcoma-associated herpesvirus (KSHV), or hu man herpesvirus 8, from latency requires transcriptional transactivation by the viral protein RTA encoded by the ORF50 gene. Very little is known abou t how RTA functions and the cellular factors that may be involved in its tr ansactivation function. Using the yeast two-hybrid system, we have identifi ed a human cellular protein that can interact with KSHV RTA. The cellular p rotein, referred to as the human hypothetical protein MGC2663 by GenBank, i s encoded by human chromosome 19. This protein is 554 amino acids (aa) in s ize and displays sequence similarity with members of the Krueppel-associate d box-zinc finger proteins (KRAB-ZFPs). MGC2663 expression could be detecte d in all primate cell lines tested, and its expression level was neither st imulated nor inhibited by RTA. MGC2663 specifically synergizes with RTA to activate viral transcription, and overexpression of MGC2663 in the presence of RTA further enhances RTA transactivation of several viral promoters tha t were identified as targets for RTA. Coimmunoprecipitation and pull-down a ssays further demonstrated that MGC2663 interacts with RTA both in vivo and in vitro, and the N-terminal 273 aa of KSHV RTA and the potential zinc fin ger domain of MGC2663 are required for their interaction. Our results indic ate that this novel human cellular protein, MGC2663, named K-RBP (KSHV RTA binding protein) due to its RTA binding feature, specifically interacts wit h the KSHV RTA protein and functions as a cellular RTA cofactor to activate viral gene expression. Though its normal cellular function needs to be fur ther studied, K-R-BP may play a significant role in mediating RTA transacti vation in vivo.