TY - JOUR
T1 - CRISPR screens identify genomic ribonucleotides as a source of PARP-trapping lesions
AU - Zimmermann, Michal
AU - Murina, Olga
AU - Reijns, Martin A. M.
AU - Agathanggelou, Angelo
AU - Challis, Rachel
AU - Tarnauskaite, Zygimante
AU - Muir, Morwenna
AU - Fluteau, Adeline
AU - Aregger, Michael
AU - McEwan, Andrea
AU - Yuan, Wei
AU - Clarke, Matthew
AU - Lambros, Maryou B.
AU - Paneesha, Shankara
AU - Moss, Paul
AU - Chandrashekhar, Megha
AU - Angers, Stephane
AU - Moffat, Jason
AU - Brunton, Valerie G.
AU - Hart, Traver
AU - de Bono, Johann
AU - Stankovic, Tatjana
AU - Jackson, Andrew P.
AU - Durocher, Daniel
PY - 2019/1/12
Y1 - 2019/1/12
N2 - The observation that BRCA1- and BRCA2-deficient cells are sensitive to inhibitors of poly(ADP–ribose) polymerase (PARP) has spurred the development of cancer therapies that use these inhibitors to target deficiencies in homologous recombination. The cytotoxicity of PARP inhibitors depends on PARP trapping, the formation of non-covalent protein–DNA adducts composed of inhibited PARP1 bound to DNA lesions of unclear origins. To address the nature of such lesions and the cellular consequences of PARP trapping, we undertook three CRISPR (clustered regularly interspersed palindromic repeats) screens to identify genes and pathways that mediate cellular resistance to olaparib, a clinically approved PARP inhibitor. Here we present a high-confidence set of 73 genes, which when mutated cause increased sensitivity to PARP inhibitors. In addition to an expected enrichment for genes related to homologous recombination, we discovered that mutations in all three genes encoding ribonuclease H2 sensitized cells to PARP inhibition. We establish that the underlying cause of the PARP-inhibitor hypersensitivity of cells deficient in ribonuclease H2 is impaired ribonucleotide excision repair. Embedded ribonucleotides, which are abundant in the genome of cells deficient in ribonucleotide excision repair, are substrates for cleavage by topoisomerase 1, resulting in PARP-trapping lesions that impede DNA replication and endanger genome integrity. We conclude that genomic ribonucleotides are a hitherto unappreciated source of PARP-trapping DNA lesions, and that the frequent deletion of RNASEH2B in metastatic prostate cancer and chronic lymphocytic leukaemia could provide an opportunity to exploit these findings therapeutically.
AB - The observation that BRCA1- and BRCA2-deficient cells are sensitive to inhibitors of poly(ADP–ribose) polymerase (PARP) has spurred the development of cancer therapies that use these inhibitors to target deficiencies in homologous recombination. The cytotoxicity of PARP inhibitors depends on PARP trapping, the formation of non-covalent protein–DNA adducts composed of inhibited PARP1 bound to DNA lesions of unclear origins. To address the nature of such lesions and the cellular consequences of PARP trapping, we undertook three CRISPR (clustered regularly interspersed palindromic repeats) screens to identify genes and pathways that mediate cellular resistance to olaparib, a clinically approved PARP inhibitor. Here we present a high-confidence set of 73 genes, which when mutated cause increased sensitivity to PARP inhibitors. In addition to an expected enrichment for genes related to homologous recombination, we discovered that mutations in all three genes encoding ribonuclease H2 sensitized cells to PARP inhibition. We establish that the underlying cause of the PARP-inhibitor hypersensitivity of cells deficient in ribonuclease H2 is impaired ribonucleotide excision repair. Embedded ribonucleotides, which are abundant in the genome of cells deficient in ribonucleotide excision repair, are substrates for cleavage by topoisomerase 1, resulting in PARP-trapping lesions that impede DNA replication and endanger genome integrity. We conclude that genomic ribonucleotides are a hitherto unappreciated source of PARP-trapping DNA lesions, and that the frequent deletion of RNASEH2B in metastatic prostate cancer and chronic lymphocytic leukaemia could provide an opportunity to exploit these findings therapeutically.
U2 - 10.1038/s41586-018-0291-z
DO - 10.1038/s41586-018-0291-z
M3 - Article
SN - 0028-0836
VL - 559
SP - 285
EP - 289
JO - Nature
JF - Nature
IS - 7713
ER -