Role of cdk9 in the Optimization of Expression of the Genes Regulated by ICP22 of Herpes Simplex Virus 1

Abstract

ICP22 is a multifunctional herpes simplex virus 1 (HSV-1) regulatory protein that regulates the accumulation of a subset of late (␥ 2 ) proteins exemplified by U L 38, U L 41, and U S 11. ICP22 binds the cyclin-dependent kinase 9 (cdk9) but not cdk7, and this complex in conjunction with viral protein kinases phosphorylates the carboxyl terminus of RNA polymerase II (Pol II) in vitro. The primary function of cdk9 and its partners, the cyclin T variants, is in the elongation of RNA transcripts, although functions related to the initiation and processing of transcripts have also been reported. We report two series of experiments designed to probe the role of cdk9 in infected cells. In the first, infected cells were treated with 5,6-dichloro-1-␤-D-ribofuranosylbenzimidazole (DRB), a specific inhibitor of cdk9. In cells treated with DRB, the major effect was in the accumulation of viral RNAs and proteins regulated by ICP22. The accumulation of ␣, ␤, or ␥ proteins not regulated by ICP22 was not affected by the drug. The results obtained with DRB were duplicated in cells transfected with small interfering RNA (siRNA) targeting cdk9 mRNAs. Interestingly, DRB and siRNA reduced the levels of ICP22 but not those of other ␣ gene products. In addition, cdk9 and ICP22 appeared to colocalize with RNA Pol II in wild-type-virus-infected cells but not in ⌬U L 13-infected cells. We conclude that cdk9 plays a critical role in the optimization of expression of genes regulated by ICP22 and that one function of cdk9 in HSV-1-infected cells may be to bring ICP22 into the RNA Pol II transcriptional complex.The studies described in this report center on the role of infected cell protein 22 (ICP22), an ␣ (immediate early) protein in viral replication. Mutants lacking ICP22 yield reduced levels of viral progeny in a cell-type-dependent manner (36). The protein appears to perform several functions (24). A key function expressed by the carboxyl-terminal domain (CTD) of ICP22 in conjunction with the viral U L 13 protein kinase is to enhance the synthesis of a subset of late (␥ 2 ) proteins exemplified by the products of the U L 38, U L 41, and U S 11 genes (2,24,31,37). In earlier studies, this laboratory reported that ICP22 and the U L 13 protein kinase mediate the activation of cdc2 and degradation of its partners, cyclins A and B (3, 4). cdc2 and its new partner, the viral DNA polymerase accessory factor (U L 42), bind topoisomerase II␣ in an ICP22-dependent manner (1, 4). In addition, ICP22 and U L 13 mediate an intermediate phosphorylation of the carboxyl terminus of RNA polymerase II (Pol II) in Vero cells (14,18,34,35).Subsequent studies designed to elucidate the interaction of ICP22 with RNA Pol II led to the discovery that ICP22 physically interacts with cyclin-dependent kinase 9 (cdk9) and that the protein complex containing ICP22 and cdk9 phosphorylated the CTD of RNA Pol II in a viral U S 3 protein kinasedependent fashion in vitro (8). These studies also showed that the CTD of RNA Pol II fused to glutathione S-transferase was phos...

CDK9 inhibitor FIT-039 prevents replication of multiple DNA viruses

“…Since CDK9/cyclin T1 complex was reported previously to be required for the replication of other viruses of Herpesviridae origin, we next verified the antiviral effect of FIT-039 on tion through phosphorylation of the CTD of RNA polymerase II and viral proteins, such as the ICP22 of HSV. The ICP22-CDK9 complex was reported previously to phosphorylate the CTD of RNA polymerase II and regulate the transcription of HSV (20,22). A previous study showed that CDK9 was localized to nuclear viral transcriptosomes at the immediate-early stage of the HCMV infection, which allowed the active transcription of HCMV IEGs (14).…”

“…CDK1 was shown to activate the expression of HSV-1 late genes (19), while CDK2 promoted the replication of adenovirus, papillomavirus, and herpes virus DNAs (7). CDK7 and CDK9 were shown to activate viral RNA transcription in HIV and HSV (7,14,15,(20)(21)(22)(23)(24). Feichtinger et al recently described the interaction between CDK9/cyclin T1 and viral mRNA export factor pUL69 in human cytomegalovirus (HCMV), which belongs to the β subgroup of Herpesviridae (20).…”

Flavopiridol induces phosphorylation of AKT in a human glioblastoma cell line, in contrast to siRNA-mediated silencing of Cdk9: Implications for drug design and development

“…[1][2][3][4][5][6][21][22][23][24][25][26][27][28][29][30][31][32][33][34] For instance, the Cdk9-interacting cellular factors comprise MyoD, retinoblastoma protein (pRb), p53, c-Myc, hSPT5 and gp130. 5 In these cases, the Cdk9-related pathway controls the phosphorylation of the aforementioned cellular factors.…”

Comprehensive Characterization of Interaction Complexes of Herpes Simplex Virus Type 1 ICP22, UL3, UL4, and UL20.5

“…It localizes in the nucleus of infected cells and functions as a transcriptional repressor for both viral and cellular genes (5). Furthermore, ICP22 associates with transcriptional complexes containing the viral transactivator protein ICP4, RNA polymerase II (4,9,14,16), and other host factors, including cdc25C (25,26) and cdk9 (7). Previous studies have reported that both UL3 and UL4 are expressed late in infection and are dispensable for viral replication in cultured cells in vitro (2,8,12).…”