53BP1 and double-strand break repair pathway choice in cancer

The cellular response to DNA double-strand breaks (DSB) is principally the result of two competing repair pathways. The Tumour Protein P53 Binding Protein 1 (TP53BP1) gene product plays a central role in DSB repair pathway choice, promoting non-homologous end-joining (NHEJ) and counteracting homologous recombination (HR). As with all DNA damage response (DDR) proteins, cell-cycle regulation of 53BP1 is critical to its function. The inappropriate activity of 53BP1 in mitosis, during which most of the DDR is inactivated to preserve genome stability, has been shown to have adverse effects on cell division that are characteristic of cancers. We explored the consequences of replication stress, associated with oncogene activity, and the impact of aberrantly active 53BP1 in mitosis. This work reveals interference with normal mitotic mechanisms for detecting and resolving stalled replication intermediates; most evidently manifest as reduced mitotic DNA synthesis (MiDAS), and increased accumulation of acentric micronuclei and G1 nuclear bodies. We propose that the functional sequestration of TOPBP1 may be responsible for these chromosomal mis-segregation defects. The mechanism by which 53BP1 (together with RIF1 and MAD2L2) promote NHEJ remains elusive. Genome instability in HR-defective cancers, principally those harbouring a mutation in BRCA1, can be rescued by the concomitant loss of either 53BP1, RIF1 or MAD2L2. This phenomenon has relevance for the development of PARP (Poly(ADP-ribose) polymerase) inhibitor resistance, now being used in BRCA1-deficient breast, ovarian and prostate cancers based on the principle of ‘synthetic lethality’. As part of a search for other resistance mechanisms, we identified the shieldin complex (SHLD1/C20orf196 and SHLD2/FAM35A) that when lost suppressed PARP inhibitor sensitivity in BRCA1-deficient cancer cells. We demonstrate that SHLD1/2 represent the terminal end-effectors of the 53BP1-RIF1-MAD2L2 axis, supporting NHEJ activity. We identified three OB-fold domains in SHLD2/FAM35A, that bind single-stranded DNA and are necessary for NHEJ. Loss of SHLD1 or SHLD2 (SHLD1/2) caused a hyper-resection phenotype, and which, in the absence of BRCA1 was associated with rescuing RAD51 loading. Downregulation in SHLD1/2 expression was also associated with intrinsic and acquired resistance across a panel of BRCA1-deficient patient-derived xenograft models. Promisingly, this mechanism may impart an acquired vulnerability to cisplatin or radiotherapy. Thus, we have characterised a critical component of the 53BP1 axis which is deterministic in repair pathway choice and carries implications for the treatment of cancers.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2019

Author dc:creator

• Dev, Harveer Singh

Advisor dc:contributor.advisor

• Jackson, Stephen Phillip

• DNA repair
• BRCA1
• 53BP1
• Shieldin
• Cancer
• PARP inhibitor
• Olaparib
• Homologous recombination
• Non-homologous end-joining
• Double-strand break repair
• Repair pathway choice

Statement dc:rights

• Page 47, Figure from Pederson et al 2015 Journal of Cell Biology. Reproduced with permission of ROCKEFELLER UNIVERSITY PRESS in the format Thesis/Dissertation via Copyright Clearance Center. (Order Detail ID: 71982557) Page 209-229, Article reproduced in full from Dev et al 2018 Nature Cell Biology. "Ownership of copyright in original research articles remains with the Author, and provided that, when reproducing the contribution or extracts from it or from the Supplementary Information, the Author acknowledges first and reference publication in the Journal, the Author retains the following non-exclusive rights: To reproduce the contribution in whole or in part in any printed volume (book or thesis) of which they are the author(s)."

Licence

• https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/7aea3c6e-0ce1-4f4b-946a-4e39e1ce0e9b/download
• https://www.rioxx.net/licenses/all-rights-reserved/

Language dc:language

en

Author Identifier

0000-0003-2874-6894
0000-0001-9317-7937

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/297644