DNA double strand break repair in human bladder cancer is error prone and involves microhomology-associated end-joining.
In human cells DNA double strand breaks (DSBs) can be repaired by the non-homologous end-joining (NHEJ) pathway. In a background of NHEJ deficiency, DSBs with mismatched ends can be joined by an error-prone mechanism involving joining between regions of nucleotide microhomology. The majority of joins formed from a DSB with partially incompatible 3' overhangs by cell-free extracts from human glioblastoma (MO59K) and urothelial (NHU) cell lines were accurate and produced by the overlap/fill-in of mismatched termini by NHEJ. However, repair of DSBs by extracts using tissue from four high-grade bladder carcinomas resulted in no accurate join formation. Junctions were formed by the non-random deletion of terminal nucleotides and showed a preference for annealing at a microhomology of 8 nt buried within the DNA substrate; this process was not dependent on functional Ku70, DNA-PK or XRCC4. Junctions were repaired in the same manner in MO59K extracts in which accurate NHEJ was inactivated by inhibition of Ku70 or DNA-PK(cs). These data indicate that bladder tumour extracts are unable to perform accurate NHEJ such that error-prone joining predominates. Therefore, in high-grade tumours mismatched DSBs are repaired by a highly mutagenic, microhomology-mediated, alternative end-joining pathway, a process that may contribute to genomic instability observed in bladder cancer.
Bentley
Johanne
J
Cancer Research UK Clinical Centre, St James's University Hospital, Leeds, LS9 7TF, UK. J.Bentley@cancermed.leeds.ac.uk
Diggle
Christine P
CP
Harnden
Patricia
P
Knowles
Margaret A
MA
Kiltie
Anne E
AE
D034961:Antigens, Nuclear / Q000502:physiology; D001483:Base Sequence; D045744:Cell Line, Tumor; D004249:DNA Damage; D004260:DNA Repair; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins / Q000502:physiology; D006801:Humans; D000072200:Ku Autoantigen; D009687:Nuclear Proteins; D017346:Protein Serine-Threonine Kinases / Q000502:physiology; D011995:Recombination, Genetic; D017385:Sequence Homology; D014516:Ureteral Neoplasms / Q000235:genetics / Q000378:metabolism; D001749:Urinary Bladder Neoplasms / Q000235:genetics* / Q000378:metabolism
D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; C101467:XRCC4 protein, human; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; D017346:Protein Serine-Threonine Kinases; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen