Altered levels of Polo-kinase Cdc5 impact on DNA double strand break response and genome stability in budding yeast

DNA double strand breaks are one of the most deleterious lesions affecting genome stability. Their accurate repair is governed by several post-translational modifications of repair factors tightly linked with cell cycle. Polo-kinase 1 (PLK1) is key regulator of mitotic transition, which is also associated with several cancers. Even though it is linked with high genome instability the underlying mechanisms are still not fully uncovered. Here, using the budding yeast as a models system, we investigate the role of Polo-kinase Cdc5 in DSB processing and repair pathway choice. Using Tetra-OFF system for Cdc5 in JKM139 cells1, we observe faster 5\u2019-3\u2019 nucleolytic degradation (resection) after inducing one irreparable DNA double strand break (DSB). On the contrary, Cdc5 overexpression slows down the DSB processing2. . We also observed increase in repair through non-homologous end joining (NHEJ) upon Cdc5 overexpression; whereas absence of Cdc5 (Tet-off) supresses the NHEJ pathway. Consistent with the biochemical analysis, the cells with hypomorphic mutant Cdc5-T238A3 exhibit 50% reduction in survival by end joining pathway after one irreparable DSB in JKM139 cells. Moreover, we further characterized the requirement of Cdc5 activity in Rad51 and Rad52 dependent DSB repair pathway choice. Our data suggests that absence of Cdc5 (Tet-off) does not inhibit the DSB repair through single strand annealing pathway, but it largely abrogates the repair through the pathway of break induced replication (BIR). Indeed the Cdc5-T238A mutant cells have reduced survival through BIR using a specific genetic background4. Taken all the data together, we suggest that Cdc5 levels and activity impact on DSB processing and repair pathway choice, affecting genome stability. References: 1. Moore, J. K. & Haber, J. E. Cell cycle and genetic requirements of two pathways of nonhomologous end-joining repair of double-strand breaks in Saccharomyces cerevisiae. Mol. Cell. Biol. 16, 2164\u20132173 (1996). 2. Donnianni, R. A. et al. Elevated levels of the polo kinase Cdc5 override the Mec1/ATR checkpoint in budding yeast by acting at different steps of the signaling pathway. PLoS Genet. 6, 1\u201314 (2010). 3. Rawal, C. C. et al. Reduced kinase activity of polo kinase Cdc5 affects chromosome stability and DNA damage response in S. cerevisiae. Cell Cycle 15, (2016). 4. Lydeard, J. R., Jain, S., Yamaguchi, M. & Haber, J. E. Break-induced replication and telomerase-independent telomere maintenance require Pol32. Nature 448, 820\u2013823 (2007)