Effects of cyclin-dependent kinase 9 (CDK9) on lytic replication of Kaposi′s sarcoma-associated herpesvirus (KSHV)

Abstract

Objective To investigate the role of cyclin-dependent kinase 9 (CDK9) in regulating lytic replication of Kaposi′s sarcoma-associated herpesvirus (KSHV). Methods Percentages of red fluorescent protein (RFP) positive cells were counted after treating and stimulating iSLK.219 cells with FIT-039, a CDK9 specific inhibitor, and doxycycline (Dox), respectively, or transfecting and stimulating them with CDK9 siRNA (si-CDK9) and Dox, respectively. Quantitative real-time PCR was used to detect the expression of KSHV genes at mRNA level in TREx-K-Rta BCBL-1 cells treated with FIT-039 or transfected with si-CKD9 in the presence of Dox. Chromatin immunoprecipitation (ChIP) assay was used to examine the enrichment of RNA polymerase Ⅱ (RNA Pol Ⅱ) and phosphorylated CTD-S2 of RNA Pol Ⅱ on the PAN promoter in TREx-K-Rta BCBL-1 cells treated with FIT-039. Results Both FIT-039 intervention and CDK9 knockdown dramatically deceased the percentage of RFP-positive iSLK.219 cells and suppressed the expression of ORF50, PAN and K2 in TREx-K-Rta BCBL-1 cells at mRNA level. Furthermore, FIT-039 significantly inhibited the enrichment of RNA Pol Ⅱ and phosphorylated CTD-S2 of RNA Pol Ⅱ on the PAN promoter in TREx-K-Rta BCBL-1 cells. Conclusion CDK9 is involved in regulating the lytic replication of KSHV through promoting the phosphorylation of CTD-S2 of RNA Pol Ⅱ. Key words: Cyclin-dependent kinase 9; Kaposi′s sarcoma-associated herpesvirus; Lytic replication; Gene transcriptional elongation