DIVERSE REGULATION OF DNA END-JOINING REPAIR ACCURACY IN HUMAN CELLS

Aneesa Talal Al-Soodani

DNA damage occurs in our cells all the time, therefore, it is extremely important that such lesions are not only processed in a timely manner, but are appropriately repaired as well. DNA double-strand breaks (DSBs) are the most toxic type of DNA damage that occurs in all cell types. Fortunately, repair mechanisms have evolved such as non-homologous end-joining (NHEJ) and homologous recombination (HR). Defects in these repair pathways can be mutagenic and increase the risk of cancer development. However, active production of DSBs is used in clinics to treat specific malignancies. Therefore, delineating how cells respond to DNA damage, and how appropriate DNA repair pathways are chosen, has been a topic of study for many years. Recently, it was discovered that the competition between BRCA1 and 53BP1 plays an important role in this regard. During S/G2 phases of the cell cycle, BRCA1 promotes end-resection, which is the starting point for HR, whereas 53BP1 inhibits end-resection. On the other hand, BRCA1 is evicted by 53BP1 from DNA DSBs during G1 phase, which promotes NHEJ. This dissertation focuses on the role of two DNA repair proteins, the MutS homologue 5 (hMSH5) and the lysine acetyl-transferase (KAT) p300, in NHEJ-mediated DSB repair. Previous studies have shown that hMSH5 plays an important role in HR repair, and this current study shows that hMSH5 regulates NHEJ-mediated repair. 53BP1 chromatin spreading is delayed in hMSH5-depleted cells, and this leads to extensive end-processing at repair joints. The average length of microhomologies (MHs) is also reduced in hMSH5-deficient cells. Second, I show that p300 chromatin association is reduced in response to IR-induced DNA damage, whereas p300 KAT-inactivation impairs its dissociation and decreases IR-induced 53BP1 recruitment. Conversely, p300 silencing increases IR-induced 53BP1 chromatin spread and decreases BRCA1 chromatin association. 53BP1 deficiency promotes a similar increase in DSB mis-joining as KAT-inactive p300. In short, p300 reduces DNA end mis-joining through the action of 53BP1. These observations demonstrate a diverse regulation of DNA end-joining accuracy through proper 53BP1 chromatin spreading. These insights will help us understand how cancer progresses through genomic instability, and aid in the discovery of new chemotherapeutic targets.

DIVERSE REGULATION OF DNA END-JOINING REPAIR ACCURACY IN HUMAN CELLS

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