A herpesvirus transactivator and cellular POU proteins extensively regulate DNA binding of the host Notch signaling protein RBP-Jκ to the virus genome

• Published in: The Journal of biological chemistry 2019-08, Vol.294 (35), p.13073-13092
• Main Authors: Gonzalez-Lopez, Olga, DeCotiis, Jennifer, Goyeneche, Corey, Mello, Helena, Vicente-Ortiz, Bryan Alexis, Shin, Hye Jin, Driscoll, Kyla E., Du, Peicheng, Palmeri, Diana, Lukac, David M.
• Format: Article
• Language: English
• Subjects: Binding Sites; Cell Line, Tumor; chromatin immunoprecipitation (ChiP); DNA - metabolism; DNA viruses; DNA-binding protein; Gene Regulation; herpesvirus; Herpesvirus 8, Human - genetics; Herpesvirus 8, Human - metabolism; Humans; Immunoglobulin J Recombination Signal Sequence-Binding Protein - metabolism; KSHV; Notch pathway; Oct-1; POU Domain Factors - metabolism; POU2F1; Rta; Trans-Activators - metabolism; tumor virus; viral DNA
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.RA118.007331

Reactivation of Kaposi's sarcoma-associated herpesvirus (KSHV) from latency requires the viral transactivator Rta to contact the host protein Jκ recombination signal-binding protein (RBP-Jκ or CSL). RBP-Jκ normally binds DNA sequence-specifically to determine the transcriptional targets of the Notch-signaling pathway, yet Notch alone cannot reactivate KSHV. We previously showed that Rta stimulates RBP-Jκ DNA binding to the viral genome. On a model viral promoter, this function requires Rta to bind to multiple copies of an Rta DNA motif (called “CANT” or Rta-c) proximal to an RBP-Jκ motif. Here, high-resolution ChIP/deep sequencing from infected primary effusion lymphoma cells revealed that RBP-Jκ binds nearly exclusively to different sets of viral genome sites during latency and reactivation. RBP-Jκ bound DNA frequently, but not exclusively, proximal to Rta bound to single, but not multiple, Rta-c motifs. To discover additional regulators of RBP-Jκ DNA binding, we used bioinformatics to identify cellular DNA-binding protein motifs adjacent to either latent or reactivation-specific RBP-Jκ–binding sites. Many of these cellular factors, including POU class homeobox (POU) proteins, have known Notch or herpesvirus phenotypes. Among a set of Rta- and RBP-Jκ–bound promoters, Rta transactivated only those that also contained POU motifs in conserved positions. On some promoters, POU factors appeared to inhibit RBP-Jκ DNA binding unless Rta bound to a proximal Rta-c motif. Moreover, POU2F1/Oct-1 expression was induced during KSHV reactivation, and POU2F1 knockdown diminished infectious virus production. Our results suggest that Rta and POU proteins broadly regulate DNA binding of RBP-Jκ during KSHV reactivation.