Essential role for Cdk2 inhibitory phosphorylation during replication stress revealed by a human Cdk2 knockin mutation
| Autor: | Julia M. Sidorova, Jherek Swanger, Bridget T. Hughes, Raymond J. Monnat, Bruce E. Clurman |
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| Rok vydání: | 2013 |
| Předmět: |
DNA Replication
Microfluidics Cyclin A Cell Cycle Proteins Genomic Instability S Phase Cyclin-dependent kinase Cyclin E Humans Gene Knock-In Techniques Phosphorylation RNA Small Interfering Cells Cultured Cyclin-dependent kinase 1 Multidisciplinary biology Cyclin-Dependent Kinase 2 Cyclin-dependent kinase 2 DNA replication Nuclear Proteins Protein-Tyrosine Kinases Biological Sciences Cell cycle Flow Cytometry Molecular biology Electrophoresis Gel Pulsed-Field Cell biology DNA-Binding Proteins enzymes and coenzymes (carbohydrates) S Phase Cell Cycle Checkpoints biology.protein Origin recognition complex RNA Interference biological phenomena cell phenomena and immunity DNA Damage Signal Transduction Transcription Factors |
| Zdroj: | Proceedings of the National Academy of Sciences. 110:8954-8959 |
| ISSN: | 1091-6490 0027-8424 |
| DOI: | 10.1073/pnas.1302927110 |
| Popis: | Cyclin-dependent kinases (Cdks) coordinate cell division, and their activities are tightly controlled. Phosphorylation of threonine 14 (T14) and tyrosine 15 (Y15) inhibits Cdks and regulates their activities in numerous physiologic contexts. Although the roles of Cdk1 inhibitory phosphorylation during mitosis are well described, studies of Cdk2 inhibitory phosphorylation during S phrase have largely been indirect. To specifically study the functions of Cdk2 inhibitory phosphorylation, we used gene targeting to make an endogenous Cdk2 knockin allele in human cells, termed Cdk2AF, which prevents Cdk2 T14 and Y15 phosphorylation. Cdk2AF caused premature S-phase entry, rapid cyclin E degradation, abnormal DNA replication, and genome instability. Cdk2AF cells also exhibited strikingly abnormal responses to replication stress, accumulated irreparable DNA damage, and permanently exited the cell cycle after transient exposure to S-phase inhibitors. Our results reveal the specific and essential roles of Cdk2 inhibitory phosphorylation in the successful execution of the replication stress checkpoint response and in maintaining genome integrity. |
| Databáze: | OpenAIRE |
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