Repression of CDK1 and other genes with CDE and CHR promoter elements during DNA damage-induced G(2)/M arrest in human cells.
Entry into mitosis is controlled by the cyclin-dependent kinase CDK1 and can be delayed in response to DNA damage. In some systems, such G(2)/M arrest has been shown to reflect the stabilization of inhibitory phosphorylation sites on CDK1. In human cells, full G(2) arrest appears to involve additional mechanisms. We describe here the prolonged (>6 day) downregulation of CDK1 protein and mRNA levels following DNA damage in human cells. This silencing of gene expression is observed in primary human fibroblasts and in two cell lines with functional p53 but not in HeLa cells, where p53 is inactive. Silencing is accompanied by the accumulation of cells in G(2), when CDK1 expression is normally maximal. The response is impaired by mutations in cis-acting elements (CDE and CHR) in the CDK1 promoter, indicating that silencing occurs at the transcriptional level. These elements have previously been implicated in the repression of transcription during G(1) that is normally lifted as cells progress into S and G(2). Interestingly, we find that other genes, including those for CDC25C, cyclin A2, cyclin B1, CENP-A, and topoisomerase IIalpha, that are normally expressed preferentially in G(2) and whose promoter regions include putative CDE and CHR elements are also downregulated in response to DNA damage. These data, together with those of other groups, support the existence of a p53-dependent, DNA damage-activated pathway leading to CHR- and CDE-mediated transcriptional repression of various G(2)-specific genes. This pathway may be required for sustained periods of G(2) arrest following DNA damage.
Badie
C
C
Gene Targeting Group, MRC Clinical Sciences Centre, Imperial College School of Medicine, Hammersmith Hospital, London W12 0NN, United Kingdom.
Itzhaki
J E
JE
Sullivan
M J
MJ
Carpenter
A J
AJ
Porter
A C
AC
D016203:CDC2 Protein Kinase / Q000235:genetics* / Q000378:metabolism; D002453:Cell Cycle / Q000235:genetics* / Q000528:radiation effects; D002460:Cell Line; D004249:DNA Damage / Q000235:genetics* / Q000528:radiation effects; D004261:DNA Replication; D015536:Down-Regulation / Q000528:radiation effects; D005434:Flow Cytometry; D016195:G2 Phase; D005720:Gamma Rays; D015971:Gene Expression Regulation, Enzymologic; D017930:Genes, Reporter; D006801:Humans; D008938:Mitosis; D009154:Mutation; D009687:Nuclear Proteins / Q000032:analysis; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000235:genetics
D009687:Nuclear Proteins; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; D016203:CDC2 Protein Kinase