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Molecular Mechanisms of the DYRK1A-regulated DNA Repair
Polina Bukina, Dept. of Biology, with Dr. Sarah Golding, Dept. of Biology
The functions of human Dual-specificity tyrosine (Y)-Regulated Kinase 1A, or DYRK1A, include cell cycle control and differentiation. DYRK1A is required for assembly of the DREAM complex and repression of the cell cycle-dependent genes, such as BRCA1 and RAD51, in quiescence. Our lab previously reported that overexpression of DYRK1A inhibits the accumulation of a DNA repair protein 53BP1, at the DNA double-stranded breaks (DSB). Accumulation of 53BP1 is attributed to repair by non-homologous end joining (NHEJ) over homologous recombination (HRR). The function of 53BP1 is opposed by RNF169, a ubiquitin-binding protein that also accumulates at the DSB sites and promotes HRR. It was found that DYRK1A interacts with RNF169 to regulate the displacement of 53BP1 from the DSB sites. This study focuses on RNF169 in order to understand the role of DYRK1A in DNA damage response. We used the Multi-Dimensional Protein Identification Technology (MudPIT) proteomic analysis to identify RNF169-interacting proteins. Human cancer U-2 OS cells stably expressing HA-tagged RNF169, as well as control cells were used for immunoprecipitation. The samples were sent to Stowers Institute for Medical Research for MudPIT proteomic analysis. In order to understand the regulation of DNA repair by DYRK1A, the RNA sequencing dataset was analyzed as part of other studies in the lab. The expression of the mRNA for repair factors RAD51 and BRCA1 was found to be regulated by DYRK1A. To determine the significance of this finding, an experiment was designed to assess BRCA1 and RAD51 protein levels in the normal U-2 OS cells and in the cells lacking DYRK1A (U-2 OS DYRK1A knockout cells) after inducing DNA damage by gamma irradiation. It was found that the levels of RAD51, BRCA1 and 53BP1 levels were increased with DYRK1A KO. These results were consistent with the finding that DNA repair efficiency is increased with DYRK1A KO. Further studies can help to understand if these effects are mediated by DYRK1A-regulated DREAM complex.
Larisa Litovchick, M.D., Ph.D.
Sarah Golding, Ph.D.
Virginia Commonwealth University. Undergraduate Research Opportunities Program
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