title: Molecular Determinants for the Inactivation of the Retinoblastoma Tumor Suppressor by the Viral Cyclin-dependent Kinase UL97.
abstract: The retinoblastoma (Rb) tumor suppressor restricts cell cycle progression by repressing E2F-responsive transcription. Cellular cyclin-dependent kinase (CDK)-mediated Rb inactivation through phosphorylation disrupts Rb-E2F complexes, stimulating transcription. The human cytomegalovirus (HCMV) UL97 protein is a viral CDK (v-CDK) that phosphorylates Rb. Here we show that UL97 phosphorylates 11 of the 16 consensus CDK sites in Rb. A cleft within Rb accommodates peptides with the amino acid sequence LXCXE. UL97 contains three such motifs. We determined that the first LXCXE motif (L1) of UL97 and the Rb cleft enhance UL97-mediated Rb phosphorylation. A UL97 mutant with a non-functional L1 motif (UL97-L1m) displayed significantly reduced Rb phosphorylation at multiple sites. Curiously, however, it efficiently disrupted Rb-E2F complexes but failed to relieve Rb-mediated repression of E2F reporter constructs. The HCMV immediate early 1 protein cooperated with UL97-L1m to inactivate Rb in transfection assays, likely indicating that cells infected with a UL97-L1m mutant virus show no defects in growth or E2F-responsive gene expression because of redundant viral mechanisms to inactivate Rb. Our data suggest that UL97 possesses a mechanism to elicit E2F-dependent gene expression distinct from disruption of Rb-E2F complexes and dependent upon both the L1 motif of UL97 and the cleft region of Rb.
authors.affiliation: From the Institute for Molecular Virology and McArdle Laboratory for Cancer Research, University of Wisconsin-Madison, Madison, Wisconsin 53706 and.
authors.forename: Satoko
authors.initials: S
authors.lastname: Iwahori
authors.affiliation: the Division of Infectious Diseases, Oregon Health and Science University and.
authors.forename: Morgan
authors.initials: M
authors.lastname: Hakki
authors.affiliation: the Division of Infectious Diseases, Oregon Health and Science University and Veterans Affairs Portland Health Care System, Portland, Oregon 97239.
authors.forename: Sunwen
authors.initials: S
authors.lastname: Chou
authors.affiliation: From the Institute for Molecular Virology and McArdle Laboratory for Cancer Research, University of Wisconsin-Madison, Madison, Wisconsin 53706 and rfkalejta@wisc.edu.
authors.forename: Robert F
authors.initials: RF
authors.lastname: Kalejta
mesh_terms: D001665:Binding Sites; D045744:Cell Line, Tumor; D003587:Cytomegalovirus / Q000235:genetics / Q000378:metabolism*; D050687:E2F1 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism* / Q000821:virology; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D054884:Host-Pathogen Interactions; D006801:Humans; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D061251:Primary Cell Culture; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D016160:Retinoblastoma Protein / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D014158:Transcription, Genetic
chemical_list: D050687:E2F1 Transcription Factor; D016160:Retinoblastoma Protein; D008156:Luciferases; D017853:Phosphotransferases (Alcohol Group Acceptor); C075365:ganciclovir kinase
keywords: cancer; cyclin-dependent kinase (CDK); cytomegalovirus; herpesvirus; oncogene; phosphorylation; retinoblastoma protein (pRb, RB); transcription