title: Evidence for a CDK4-dependent checkpoint in a conditional model of cellular senescence.
abstract: Cellular senescence, the stable cell cycle arrest elicited by various forms of stress, is an important facet of tumor suppression. Although much is known about the key players in the implementation of senescence, including the pRb and p53 axes and the cyclin dependent kinase inhibitors p16(INK4a) and p21(CIP1), many details remain unresolved. In studying conditional senescence in human fibroblasts that express a temperature sensitive SV40 large T-antigen (T-Ag), we uncovered an unexpected role for CDK4. At the permissive temperature, where pRb and p53 are functionally compromised by T-Ag, cyclin D-CDK4 complexes are disrupted by the high p16(INK4a) levels and reduced expression of p21(CIP1). In cells arrested at the non-permissive temperature, p21(CIP1) promotes reassembly of cyclin D-CDK4 yet pRb is in a hypo-phosphorylated state, consistent with cell cycle arrest. In exploring whether the reassembled cyclin D-CDK4-p21 complexes are functional, we found that shRNA-mediated knockdown or chemical inhibition of CDK4 prevented the increase in cell size associated with the senescent phenotype by allowing the cells to arrest in G1 rather than G2/M. The data point to a role for CDK4 kinase activity in a G2 checkpoint that contributes to senescence.
authors.affiliation: a Cancer Research-UK London Research Institute ; London , UK.
authors.forename: Sharon
authors.initials: S
authors.lastname: Brookes
authors.forename: Sladjana
authors.initials: S
authors.lastname: Gagrica
authors.forename: Elaine
authors.initials: E
authors.lastname: Sanij
authors.forename: Janice
authors.initials: J
authors.lastname: Rowe
authors.forename: Fiona J
authors.initials: FJ
authors.lastname: Gregory
authors.forename: Eiji
authors.initials: E
authors.lastname: Hara
authors.forename: Gordon
authors.initials: G
authors.lastname: Peters
mesh_terms: D000952:Antigens, Polyomavirus Transforming / Q000235:genetics / Q000378:metabolism; D059447:Cell Cycle Checkpoints / Q000187:drug effects; D002460:Cell Line; D016922:Cellular Senescence; D019938:Cyclin D1 / Q000378:metabolism; D051358:Cyclin-Dependent Kinase 4 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D051361:Cyclin-Dependent Kinase 6 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000378:metabolism; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008954:Models, Biological; D010641:Phenotype; D010879:Piperazines / Q000494:pharmacology; D011725:Pyridines / Q000494:pharmacology; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D016160:Retinoblastoma Protein / Q000378:metabolism; D013696:Temperature; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
chemical_list: D000952:Antigens, Polyomavirus Transforming; D019941:Cyclin-Dependent Kinase Inhibitor p16; D050759:Cyclin-Dependent Kinase Inhibitor p21; D010879:Piperazines; D011725:Pyridines; D034741:RNA, Small Interfering; D016160:Retinoblastoma Protein; D016159:Tumor Suppressor Protein p53; D019938:Cyclin D1; D051358:Cyclin-Dependent Kinase 4; D051361:Cyclin-Dependent Kinase 6; C500026:palbociclib
keywords: BrdU, bromodeoxyuridine; CDK, cyclin dependent kinase; CDK4; FACS, fluorescence actvated cell sorting; HFs, human fibroblasts; PI, propidium iodide; SA-βgal, senescence-associated β-galactosidase activity; SV40 T-antigen; SV40, simian virus 40; TERT, telomerase reverse transcriptase; human fibroblasts; p16INK4a; p21CIP1; p53; pRb, retinoblastoma protein; retinoblastoma protein; senescence; shRNA, short-hairpin RNA; ts, temperature sensitive