tailieunhanh - Báo cáo khoa học: Poly(ADP-ribose) polymerase-1 protects excessive DNA strand breaks from deterioration during repair in human cell extracts

Base excision repair (BER), a major pathway for the removal of simple lesions in DNA, requires the co-ordinated action of several repair and ancillary proteins, the impairment of which can lead to genetic instability. We here address the role of poly(ADP-ribose) polymerase-1 (PARP-1) in BER. Using anin vitro cross-linking assay, we reveal that PARP-1 is always involved in repair of a uracil-containing oligonucleotide and that it binds to the damaged DNA during the early stages of repair. | iFEBS Journal Poly ADP-ribose polymerase-1 protects excessive DNA strand breaks from deterioration during repair in human cell extracts Jason L. Parsons Irina I. Dianova Sarah L. Allinson and Grigory L. Dianov MRC Radiation and Genome Stability Unit Harwell Oxfordshire UK Keywords base excision repair DNA polymerase b DNA repair poly ADP-ribose polymerase-1 PARP-1 XRCC1 Correspondence G. L. Dianov Radiation and Genome Stability Unit MedicalResearch Council Harwell Oxfordshire OX11 0RD UK Fax 44 1235 841 200 Tel 44 1235 841 134 E-mail Present address Department of BiologicalSciences Lancaster University Lancaster LA1 4YQ UK Received 13 December 2004 revised 19 January 2005 accepted 24 February 2005 Base excision repair BER a major pathway for the removal of simple lesions in DNA requires the co-ordinated action of several repair and ancillary proteins the impairment of which can lead to genetic instability. We here address the role of poly ADP-ribose polymerase-1 PARP-1 in BER. Using an in vitro cross-linking assay we reveal that PARP-1 is always involved in repair of a uracil-containing oligonucleotide and that it binds to the damaged DNA during the early stages of repair. Inhibition of PARP-1 poly ADP-ribosyl ation by 3-aminobenzamide blocks dissociation of PARP-1 from damaged DNA and prevents further repair. We find that excessive poly ADP-ribosyl ation occurs when repair intermediates containing single-strand breaks are in excess of the repair capacity of the cell extract suggesting that repeated binding of PARP-1 to the nicked DNA occurs. We also find increased sensitivity of repair intermediates to nuclease cleavage in PARP-deficient mouse fibroblasts and after depletion of PARP-1 from HeLa whole cell extracts. Our data support the model in which PARP-1 binding to DNA single-strand breaks or repair intermediates plays a protective role when repair is limited. doi Spontaneously derived DNA lesions such as