Upton, Jason W. a, b, Linda F. van Dyck and Samuel H. Speck,a, 2005, Virology 341:271-283.
aCenter for Emerging Infectious Diseases, Yerkes National Primate Research Center, Emory University School of Medicine, 954 Gatewood Road, NE Atlanta, GA 30329, USA bGraduate Program in Molecular Cell Biology, Washington University School of Medicine, St. Louis, MO 63110, USA cDepartment of Microbiology, University of Colorado Health Sciences Center, Denver, CO 80262, USA
All known γ2-herpesviruses encode a cyclin homolog with significant homology to mammalian D-type cyclins. The murine gammaherpesvirus 68 (γHV68) viral cyclin (v-cyclin) has been shown to be oncogenic when expression is targeted to thymocytes in transgenic mice and to be critical for virus reactivation from latency. Here, we investigate the interaction of the γHV68 v-cyclin with cellular cyclin-dependent kinases (cdks). We show that, in contrast to the Kaposi's sarcoma-associated herpesvirus (KSHV) v-cyclin, the γHV68 v-cyclin preferentially interacts with cdk2 and cdc2 but does not interact with either cdk4 or cdk6. Mutation of conserved residues, predicted to be involved in cdk binding based on the γHV68 v-cyclin:cdk2 crystal structure, resulted in the loss of both cdk binding and the ability to mediate phosphorylation of substrates. Like the KSHV v-cyclin, the γHV68 v-cyclin appears to confer expanded substrate specificity to the cellular cdk binding partners. As expected, the γHV68 v-cyclin:cdk complexes are able to target phosphorylation of histone H1, the retinoblastoma protein (pRb), and p27Kip1 as assessed using in vitro kinase assays. Notably, hyperphosphorylation of pRb was observed during wt γHV68 replication in serum-starved murine fibroblasts, but not in cells that were either mock-infected or infected with a v-cyclin null γHV68. In addition, infection of serum-starved murine fibroblasts also results in a v-cyclin-dependent increase in cdk2-associated kinase activity and a concomitant decrease in the levels of p27Kip1. Taken together, the latter studies served to validate the results of the in vitro kinase assays. Finally, in vitro kinase assays revealed that the γHV68 v-cyclin:cdk complexes can also phosphorylate p21Cip1, Bcl-2, and p53. The latter suggests that, at least in vitro, the γHV68 v-cyclin exhibits functional characteristics of both cyclin E and cyclin A.
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