Poster abstracts

Poster number 60 submitted by Yiheng Hu

53BP1 dynamics regulated in a RNF8/ RNF168- Dependent manner in Double- stranded DNA Break Repair

Yiheng Hu (The Ohio State University), Chao Wang (The Ohio State University), Kun Huang (The Ohio State University), Neelima Mondal (Jawaharlal Nehru University), Jeffrey D. Parvin (The Ohio State University)

Abstract:
53BP1 regulates DNA double- stranded break (DSB) repair pathways. In functional assays for DSB repair, we found that 53BP1 was important in the non-homologous end-joining pathway, and this activity was dependent upon RNF8 and RNF168. We observed that upon irradiation of cells, 53BP1 is degraded everywhere except at DNA damage sites, and this proteasome- mediated degradation of 53BP1 was dependent on RNF8 and RNF168. The puzzle is that 53BP1 has been characterized to form intense irradiation induced foci (IRIF), and this is in apparent contradiction to our observation of its degradation following DNA damage. Image analysis of 53BP1 IRIF confirmed our results. 53BP1 protein was diffusely abundant in nuclei, and upon ionizing radiation, 53BP1 was everywhere degraded except at DNA damage sites where the 53BP1 protein is stabilized. Depletion of RNF8 or RNF168 blocked the degradation of the diffusely nuclear 53BP1, and IRIF did not form. Our results suggest that 53BP1 functions specifically in the NHEJ pathway, and its function is dependent on degradation of most nuclear 53BP1 protein by RNF8/RNF168. We infer from these results that 53BP1 is required to recruit a factor to sites of NHEJ repair, and if it remains in high concentration throughout the nucleus, then the critical repair factor fails to bind to the DNA lesion.

References:
1. Kuniyoshi Iwabuchi et al., ‘Potential Role for 53BP1 in DNA End-joining Repair Through Direct Interaction with DNA.’, The Journal of biological chemistry, 278 (2003), 36487–95
2. Yaron Galanty et al., ‘Mammalian SUMO E3-ligases PIAS1 and PIAS4 Promote Responses to DNA Double-strand Breaks.’, Nature, 462 (2009), 935–9
3. Niels Mailand et al., ‘RNF8 Ubiquitylates Histones at DNA Double-strand Breaks and Promotes Assembly of Repair Proteins.’, Cell, 131 (2007), 887–900
4. Jiangbo Tang et al., ‘Acetylation Limits 53BP1 Association with Damaged Chromatin to Promote Homologous Recombination.’, Nature structural & molecular biology, 2013

Keywords: 53BP1, Double- stranded break repair, RNF8 RNF168