Functional Competition between Poly(ADP-ribose) Polymerase and Its 24-kDa Apoptotic Fragment in DNA Repair and Transcription

Abstract

Poly(ADP-ribose) polymerase is a 113-kDa nuclear enzyme that binds to both damaged DNA and to RNA associated with actively transcribed regions of chromatin. Binding of poly(ADP-ribose) polymerase to DNA lesions activates it, catalyzing the covalent addition of multiple ADP-ribose polymers to the enzyme (automodification). During apoptosis, poly(ADP-ribose) polymerase is cleaved by caspase-3, resulting in the formation of an N-terminal 24-kDa fragment, containing the DNA binding domain, and a C-terminal 89-kDa catalytic fragment. The functional relevance of this cleavage is not well understood. We therefore prepared a recombinant 24-kDa poly(ADP-ribose) polymerase fragment and investigated the role of this fragment in DNA repair and transcription. The 24-kDa fragment retained its binding affinity for both DNA breaks and RNA. In an in vitro cell-free DNA repair assay, this fragment inhibited rejoining of DNA breaks and suppressed ADP-ribose polymer formation by competing with poly(ADP-ribose) polymerase in binding to DNA breaks. With regard to transcription, it has recently been demonstrated that binding of poly(ADP-ribose) polymerase to transcribed RNA reduces the rate of transcript elongation and that automodification of poly(ADP-ribose) polymerase bound to DNA breaks results in up-regulation of transcription. We tested the 24-kDa fragment for its ability to suppress transcript elongation, and we found that it competed against the up-regulation of transcription mediated by full-length poly(ADP-ribose) polymerase. The ability of the 24-kDa fragment to inhibit DNA repair, ADP-ribose polymer formation, and damage-dependent up-regulation of transcription may contribute to the apoptotic shift from cell survival to cell death mode. Poly(ADP-ribose) polymerase is a 113-kDa nuclear enzyme that binds to both damaged DNA and to RNA associated with actively transcribed regions of chromatin. Binding of poly(ADP-ribose) polymerase to DNA lesions activates it, catalyzing the covalent addition of multiple ADP-ribose polymers to the enzyme (automodification). During apoptosis, poly(ADP-ribose) polymerase is cleaved by caspase-3, resulting in the formation of an N-terminal 24-kDa fragment, containing the DNA binding domain, and a C-terminal 89-kDa catalytic fragment. The functional relevance of this cleavage is not well understood. We therefore prepared a recombinant 24-kDa poly(ADP-ribose) polymerase fragment and investigated the role of this fragment in DNA repair and transcription. The 24-kDa fragment retained its binding affinity for both DNA breaks and RNA. In an in vitro cell-free DNA repair assay, this fragment inhibited rejoining of DNA breaks and suppressed ADP-ribose polymer formation by competing with poly(ADP-ribose) polymerase in binding to DNA breaks. With regard to transcription, it has recently been demonstrated that binding of poly(ADP-ribose) polymerase to transcribed RNA reduces the rate of transcript elongation and that automodification of poly(ADP-ribose) polymerase bound to DNA breaks results in up-regulation of transcription. We tested the 24-kDa fragment for its ability to suppress transcript elongation, and we found that it competed against the up-regulation of transcription mediated by full-length poly(ADP-ribose) polymerase. The ability of the 24-kDa fragment to inhibit DNA repair, ADP-ribose polymer formation, and damage-dependent up-regulation of transcription may contribute to the apoptotic shift from cell survival to cell death mode. poly(ADP-ribose) polymerase N-methyl-N′-nitro-N-nitrosoguanidine enzyme-linked immunosorbent assay terminal dUTP nick-end labeling Dulbecco's modified Eagle's medium. Poly(ADP-ribose) polymerase (PARP)1 is a highly abundant nuclear enzyme present at about 2 × 105 molecules per nucleus (1Ludwig A. Behnke B. Holtlund J. Hilz H. J. Biol. Chem. 1988; 263: 6993-6999Abstract Full Text PDF PubMed Google Scholar). This enzyme is composed of an N-terminal DNA binding domain, containing two zinc finger motifs, a C-terminal NAD+ binding domain, catalyzing the synthesis of ADP-ribose polymers from its substrate, NAD+, and an automodification site, which unites the N-terminal and C-terminal domains (2de Murcia G. Ménissier-de Murcia J. Schreiber V. BioEssays. 1991; 13: 455-462Crossref PubMed Scopus (93) Google Scholar). Poly(ADP-ribosyl)ation by PARP at the automodification site of the protein is initiated by the binding of the zinc fingers to DNA breaks (3Althaus F.R. Richter C. ADP-ribosylation of Proteins: Enzymology and Biological Significance. Springer-Verlag, Berlin1987: 3-113Google Scholar, 4Lindahl T. Satoh M.S. Poirier G.G. Klungland A. Trends Biochem. Sci. 1995; 20: 405-411Abstract Full Text PDF PubMed Scopus (578) Google Scholar). As a consequence of this automodification, the binding affinity of PARP for DNA is reduced, resulting in dissociation of PARP from DNA breaks (5Zahradka P. Ebisuzaki K. Eur. J. Biochem. 1982; 127: 579-585Crossref PubMed Scopus (146) Google Scholar) and thereby allowing the DNA repair machinery to access the sites of DNA damage (6Satoh M.S. Lindahl T. Nature. 1992; 356: 356-358Crossref PubMed Scopus (975) Google Scholar). In cells where DNA breaks are generated by DNA-damaging agents, PARP is activated and automodified (3Althaus F.R. Richter C. ADP-ribosylation of Proteins: Enzymology and Biological Significance. Springer-Verlag, Berlin1987: 3-113Google Scholar, 4Lindahl T. Satoh M.S. Poirier G.G. Klungland A. Trends Biochem. Sci. 1995; 20: 405-411Abstract Full Text PDF PubMed Scopus (578) Google Scholar), leading to the conclusion that PARP is involved in the cellular response to genetic damage, particularly in the repair of damaged DNA (3Althaus F.R. Richter C. ADP-ribosylation of Proteins: Enzymology and Biological Significance. Springer-Verlag, Berlin1987: 3-113Google Scholar). However, PARP has been shown to lack DNA repair activity in itself (6Satoh M.S. Lindahl T. Nature. 1992; 356: 356-358Crossref PubMed Scopus (975) Google Scholar, 7Vodenicharov M.D. Sallmann F.R. Wang Z.-Q. Satoh M.S. Poirier G.G. Nucleic Acids Res. 2000; 28: 3887-3896Crossref PubMed Scopus (117) Google Scholar). Alternatively, it has been suggested that PARP is involved in chromatin stabilization (8Simbulan-Rosenthal C.M. Haddad B.R. Rosenthal D.S. Weaver Z. Coleman A. Luo R. Young H.M. Wang Z.Q. Ried T. Smulson M.E. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: 13191-13196Crossref PubMed Scopus (110) Google Scholar), in DNA replication (9Dantzer F. Nasheuer H.P. Vonesch J.L. de Murcia G. Menissier-de Murcia J. Nucleic Acids Res. 1998; 26: 1891-1898Crossref PubMed Scopus (138) Google Scholar, 10Simbulan-Rosenthal C.M. Rosenthal D.S. Boulares A.H. Hickey R.J. Malkas L.H. Coll J.M. Smulson M.E. Biochemistry. 1998; 37: 9363-9370Crossref PubMed Scopus (77) Google Scholar), and in transcription (11Kannan P., Yu, Y. Wankhade S. Tainsky M.A. Nucleic Acids Res. 1999; 27: 866-874Crossref PubMed Scopus (124) Google Scholar, 12Meisterernst M. Stelzer G. Roeder R.G. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 2261-2265Crossref PubMed Scopus (145) Google Scholar, 13Anderson M.G. Scoggin K.E. Simbulan-Rosenthal C.M. Steadman J.A. J. Virol. 2000; 74: 2169-2177Crossref PubMed Scopus (64) Google Scholar), although the roles played by PARP in these processes are not yet understood. In nuclear localization experiments, PARP is observed in clear foci, associated both with regions of chromatin actively transcribed by RNA polymerase II as well as with nucleoli where rRNA is synthesized by RNA polymerase I (14Fakan S. Leduc Y. Lamarre D. Brunet G. Poirier G.G. Exp. Cell Res. 1988; 179: 517-526Crossref PubMed Scopus (31) Google Scholar). Dispersal of the foci upon treatment of cells with the transcription inhibitors actinomycin D or 5,6-dichloro-1-β-ribofuranosylbenzimidazole suggests an involvement of PARP in transcription (15Desnoyers S. Kaufmann S.H. Poirier G.G. Exp. Cell Res. 1996; 227: 146-153Crossref PubMed Scopus (58) Google Scholar). In addition, such foci are also dispersed by treatment of isolated nuclei with RNase (16Kaufmann S.H. Brunet G. Talbot B. Lamarr D. Dumas C. Shaper J.H. Poirier G. Exp. Cell Res. 1991; 192: 524-535Crossref PubMed Scopus (52) Google Scholar). These observations thus suggest an interaction between PARP and transcribed RNA. Recently, we demonstrated that RNA-bound PARP reduces the rate of RNA elongation by RNA polymerase II and that automodification of PARP in response to DNA damage up-regulates transcription (17Vispé S. Yung T.M.C. Ritchot J. Serizawa H. Satoh M.S. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 9886-9891Crossref PubMed Scopus (75) Google Scholar). Since DNA-damaging agents induce RNA damage as well, we proposed that this up-regulation allows cells to compensate for the loss of damaged RNA that occurs collaterally with DNA damage and that this pathway is required for cell survival following exposure to DNA-damaging agents (17Vispé S. Yung T.M.C. Ritchot J. Serizawa H. Satoh M.S. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 9886-9891Crossref PubMed Scopus (75) Google Scholar). When cells are exposed to sufficiently high levels of DNA-damaging agents, they commit to cell death by inducing either apoptosis or necrosis. During apoptosis PARP is cleaved by the apoptosis-specific protease, caspase-3, resulting in the formation of an N-terminal 24-kDa fragment, containing the DNA binding domain, and a C-terminal 89-kDa catalytic domain, containing the automodification site (18Kaufmann S.H. Cancer Res. 1989; 49: 5870-5878PubMed Google Scholar, 19Duriez P.J. Shah G.M. Biochem. Cell Biol. 1997; 75: 337-349Crossref PubMed Scopus (417) Google Scholar, 20Lazebnik Y.A. Kaufmann S.H. Desnoyers S. Poirier G.G. Earnshaw W.C. Nature. 1994; 371: 346-347Crossref PubMed Scopus (2351) Google Scholar). Recently, Halappanavar et al. (21Halappanavar S. J. Earnshaw W.C. Shah G.M. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar) and et al. de G. V. de Murcia G. Murcia J.M. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar) in PARP of the of DNA This suggests that cleavage of PARP has a role in In addition, and Wang Z. Wang Z.Q. Biol. 1999; PubMed Scopus Google Scholar) and Boulares et al. A.H. V. Wang G. S. Smulson M. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar) suggested that cleavage of PARP is also required for cell and Wang Z. Wang Z.Q. Biol. 1999; PubMed Scopus Google Scholar) found cell death by Boulares et A.H. V. Wang G. S. Smulson M. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar) demonstrated a of cell death by PARP in PARP Since the 24-kDa fragment the DNA binding domain, which is of binding to DNA breaks M.E. D. M. A. S. A. Simbulan-Rosenthal C. Rosenthal D. A. A. Cancer Res. 1998; Google Scholar), it has been that the 24-kDa fragment of PARP and the of apoptosis P.J. Shah G.M. Biochem. Cell Biol. 1997; 75: 337-349Crossref PubMed Scopus (417) Google Scholar). However, of the 24-kDa fragment to we prepared the 24-kDa apoptotic fragment of PARP and the 24-kDa fragment against the of PARP in DNA repair, ADP-ribose polymer formation, and transcription. cells from Cell The against the automodification of PARP and the against the DNA binding of PARP finger P.J. Desnoyers S. Shah G.M. B. S. Poirier G.G. Talbot B. 1997; PubMed Scopus Google Scholar) by G. G. PARP or the to the 24-kDa DNA binding of PARP which is found in apoptotic cells (18Kaufmann S.H. Cancer Res. 1989; 49: 5870-5878PubMed Google Scholar, 19Duriez P.J. Shah G.M. Biochem. Cell Biol. 1997; 75: 337-349Crossref PubMed Scopus (417) Google Scholar, 20Lazebnik Y.A. Kaufmann S.H. Desnoyers S. Poirier G.G. Earnshaw W.C. Nature. 1994; 371: 346-347Crossref PubMed Scopus (2351) Google Scholar), and to cells with in 2 of in the of and for and of PARP or the 24-kDa fragment in the of for at The at × for and the in and The resulting in of containing 2 and and PARP or the 24-kDa fragment by a at × the for of PARP or the 24-kDa fragment. to a and with PARP a of containing The of by of The of with to the and to a DNA and with the with of by of containing PARP with of containing The with to the to a with PARP a of containing and the against to as for PARP that the DNA The against to The recombinant PARP and the 24-kDa fragment by PARP and the 24-kDa fragment a with the P.J. Desnoyers S. Shah G.M. B. S. Poirier G.G. Talbot B. 1997; PubMed Scopus Google Scholar) which the zinc finger 2 present in both PARP and the 24-kDa fragment, and with to addition of to the PARP and the 24-kDa fragment at the DNA for the assay, a at for of with of in a the by the The with its in a containing and by for at by at and at The with and and the resulting in and The binding of the with of PARP or the 24-kDa fragment in a containing and for at in a by and the and exposed to for or for by an RNA as (17Vispé S. Yung T.M.C. Ritchot J. Serizawa H. Satoh M.S. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 9886-9891Crossref PubMed Scopus (75) Google Scholar). RNA synthesized RNA polymerase to the I The RNA with PARP or the 24-kDa fragment and by the either exposed to for or for by prepared from cells following the of al. J.L. A. M. PubMed Scopus Google Scholar). The cell-free DNA repair assay of of II containing an of DNA per (6Satoh M.S. Lindahl T. Nature. 1992; 356: 356-358Crossref PubMed Scopus (975) Google Scholar), and of the 24-kDa fragment in the or of 2 M.S. Poirier G.G. Lindahl T. J. Biol. Chem. Full Text PDF PubMed Google Scholar). of the and by the of ADP-ribose polymers in the cell-free DNA repair assay, of and to the by addition of and activity retained a as M.S. Poirier G.G. Lindahl T. J. Biol. Chem. Full Text PDF PubMed Google Scholar). elongation as (17Vispé S. Yung T.M.C. Ritchot J. Serizawa H. Satoh M.S. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 9886-9891Crossref PubMed Scopus (75) Google Scholar). a containing a with a at of the to of RNA polymerase II from DNA The DNA with RNA polymerase II by H. H. Yung Nature. 1995; PubMed Scopus Google Scholar) in the of and at for as by et al. A. 1996; PubMed Scopus Google Scholar). initiated by addition of and in the or of PARP or the 24-kDa fragment. and RNA a the exposed to for containing the by and in a transcription assay with nuclear The of the for at in a containing transcription of nuclear and of either PARP or the 24-kDa fragment. The by addition of of 2 and with by and by The to the to the 24-kDa DNA binding of PARP with in Dulbecco's modified Eagle's for at and to cells to for at cell containing and cells for which the with and the cells exposed of treatment at the with and cells for 2 in cell death the cells in and with and to the cells by the of the 24-kDa PARP fragment DNA repair and transcription, we prepared the recombinant 24-kDa fragment and full-length As shown in the 24-kDa fragment, which to an of about to as and The of full-length recombinant PARP about with observed of the 24-kDa fragment and PARP by the P.J. Desnoyers S. Shah G.M. B. S. Poirier G.G. Talbot B. 1997; PubMed Scopus Google Scholar), which the zinc finger 2 of PARP and the 24-kDa fragment. with either the 24-kDa fragment or the zinc finger found in PARP and the 24-kDa fragment to DNA (2de Murcia G. Ménissier-de Murcia J. Schreiber V. BioEssays. 1991; 13: 455-462Crossref PubMed Scopus (93) Google Scholar, J. D. M.A. M. Poirier G.G. 1999; PubMed Scopus Google Scholar), the of DNA a As shown in in observed the with the 24-kDa fragment. of the PARP of the 24-kDa fragment, with the in the of the 24-kDa fragment. In addition, the found at the of the of inhibited of the that formation of the to binding of the 24-kDa fragment or PARP to the DNA not The in 2 a between activity associated with the and the of the 24-kDa fragment or PARP and to that the 24-kDa fragment has about of the binding activity of full-length cleavage of PARP by caspase-3, the resulting 24-kDa fragment DNA binding In the of substrate, NAD+, PARP binds to and DNA thereby DNA repair (6Satoh M.S. Lindahl T. Nature. 1992; 356: 356-358Crossref PubMed Scopus (975) Google Scholar). dissociation of PARP from DNA breaks by automodification is a for DNA repair (6Satoh M.S. Lindahl T. Nature. 1992; 356: 356-358Crossref PubMed Scopus (975) Google Scholar). Since the 24-kDa fragment is of binding to DNA breaks the automodification site, these DNA breaks and inhibit DNA repair in the of this a cell-free DNA repair assay containing DNA cell-free and of the 24-kDa fragment in the or of As shown in of DNA breaks in the of NAD+ to of DNA repair by bound and dissociation of PARP from DNA breaks initiated by addition of NAD+, about of DNA breaks This DNA repair inhibited by addition of the 24-kDa fragment and of the 24-kDa fragment to PARP to inhibit DNA repair by and of PARP from of the 24-kDa with the that the 24-kDa fragment, full-length binds to and DNA breaks in the of We the of ADP-ribose polymers generated in the cell-free assay in the or of the recombinant 24-kDa fragment. As observed M.S. Poirier G.G. Lindahl T. J. Biol. Chem. Full Text PDF PubMed Google Scholar), of cell-free with DNA breaks formation of ADP-ribose polymers However, addition of the 24-kDa fragment inhibited ADP-ribose polymer formation, that the 24-kDa fragment competed with PARP in binding to DNA breaks and thereby the of in the of Since ADP-ribose polymers are synthesized from NAD+, of poly(ADP-ribose) formation the of the of NAD+ present in the following the cell-free DNA repair assay, to the assay and the to a M.S. Nucleic Acids Res. 1996; PubMed Scopus Google Scholar) for of of PARP of the NAD+, addition of the 24-kDa fragment in a in not these results suggest that the 24-kDa fragment DNA repair and ADP-ribose formation by binding to and DNA breaks in with full-length In PARP has been observed to with particularly with actively transcribed this has been shown to by binding of PARP to transcribed RNA (14Fakan S. Leduc Y. Lamarre D. Brunet G. Poirier G.G. Exp. Cell Res. 1988; 179: 517-526Crossref PubMed Scopus (31) Google Scholar, S. Kaufmann S.H. Poirier G.G. Exp. Cell Res. 1996; 227: 146-153Crossref PubMed Scopus (58) Google Scholar, S.H. Brunet G. Talbot B. Lamarr D. Dumas C. Shaper J.H. Poirier G. Exp. Cell Res. 1991; 192: 524-535Crossref PubMed Scopus (52) Google Scholar). We recently demonstrated that binding of PARP to RNA reduces the rate of RNA elongation by RNA polymerase II and that formation of DNA breaks and automodification of PARP the PARP thus RNA synthesis (17Vispé S. Yung T.M.C. Ritchot J. Serizawa H. Satoh M.S. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 9886-9891Crossref PubMed Scopus (75) Google Scholar). the 24-kDa fragment binds to RNA RNA prepared by the from I and and with either the 24-kDa fragment or As shown in PARP the of RNA a a The 24-kDa fragment also the of RNA that the fragment, full-length is of binding to RNA The in a between activity associated with the and the of 24-kDa fragment or to that the 24-kDa fragment has about of the binding activity of full-length We tested the 24-kDa fragment of elongation of RNA a elongation with a at prepared (17Vispé S. Yung T.M.C. Ritchot J. Serizawa H. Satoh M.S. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 9886-9891Crossref PubMed Scopus (75) Google Scholar) and as a During the RNA polymerase II from the where the and to the in the of in the of and resulting in the formation The initiated by and either the 24-kDa fragment or As shown in the formation of generated by of RNA polymerase II at sites to transcript with that PARP transcript elongation (17Vispé S. Yung T.M.C. Ritchot J. Serizawa H. Satoh M.S. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 9886-9891Crossref PubMed Scopus (75) Google Scholar), addition of PARP inhibited the of The addition of the 24-kDa fragment in of the that the 24-kDa fragment also to suppress RNA synthesis by RNA polymerase RNA been demonstrated to in of RNA polymerase II transcription C.M. Biochem. 1997; PubMed Scopus Google Scholar). In (17Vispé S. Yung T.M.C. Ritchot J. Serizawa H. Satoh M.S. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 9886-9891Crossref PubMed Scopus (75) Google Scholar), we suggested that the rate of transcript elongation in the of PARP occurs as the of PARP binding to and these DNA damage, automodification of PARP the of thereby transcription (17Vispé S. Yung T.M.C. Ritchot J. Serizawa H. Satoh M.S. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 9886-9891Crossref PubMed Scopus (75) Google Scholar). the the 24-kDa fragment the automodification site and may therefore with full-length PARP and inhibit its ability to transcription of DNA this we transcription nuclear in the or of the 24-kDa fragment. transcription of nuclear that about 2 of As shown in from addition of NAD+, PARP that bound to the of DNA and an in with (17Vispé S. Yung T.M.C. Ritchot J. Serizawa H. Satoh M.S. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 9886-9891Crossref PubMed Scopus (75) Google Scholar). However, the addition of the 24-kDa fragment to the in addition to PARP and in the of of the 24-kDa fragment to PARP to the transcription by and 2 of PARP from nuclear of the 24-kDa that these in inhibit up-regulation of RNA of the 24-kDa fragment cells apoptosis, cells with for The cells exposed to for and in for an 2 which apoptosis by cells with for of As shown in and a in the we found following treatment of cells with with and with a and in such of cells either the which the automodification of or the against zinc finger 2 of both PARP and the 24-kDa fragment P.J. Desnoyers S. Shah G.M. B. S. Poirier G.G. Talbot B. 1997; PubMed Scopus Google a PARP of per × cells of the 24-kDa fragment per × Since the to by the of cells following with we that the of 24-kDa fragment to PARP in cells the 24-kDa fragment This to cells to the apoptotic of During apoptosis, the abundant nuclear enzyme PARP is cleaved by caspase-3, a 24-kDa fragment containing the DNA binding (18Kaufmann S.H. Cancer Res. 1989; 49: 5870-5878PubMed Google Scholar, 19Duriez P.J. Shah G.M. Biochem. Cell Biol. 1997; 75: 337-349Crossref PubMed Scopus (417) Google Scholar, 20Lazebnik Y.A. Kaufmann S.H. Desnoyers S. Poirier G.G. Earnshaw W.C. Nature. 1994; 371: 346-347Crossref PubMed Scopus (2351) Google Scholar). In this we demonstrated that the 24-kDa fragment, full-length is of binding to both DNA breaks and although the affinity of the 24-kDa fragment for DNA breaks and RNA to and to full-length However, the high affinity of PARP for DNA breaks J. D. M.A. M. Poirier G.G. 1999; PubMed Scopus Google Scholar) and RNA H. K. T. J. Biol. Chem. Full Text PDF PubMed Google Scholar), the affinity is and with a functional role for the 24-kDa fragment in Since apoptosis in PARP cells Z.Q. C. M. M. K. 1997; PubMed Scopus Google Scholar, Y. Shah G.M. Biochem. Res. 1998; PubMed Scopus Google Scholar, Murcia J.M. C. C. M. B. M. M. A. M. C. P. de Murcia G. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: PubMed Scopus Google Scholar), cleavage of PARP may not required for apoptosis per However, it has been suggested that PARP to cells survival (21Halappanavar S. J. Earnshaw W.C. Shah G.M. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar, Murcia J.M. C. C. M. B. M. M. A. M. C. P. de Murcia G. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: PubMed Scopus Google Scholar, P. D. S. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). a in the of full-length PARP as a consequence of cleavage by may this In addition, the that the resulting 24-kDa fragment the DNA binding of PARP the automodification site may it to as a in to full-length automodification both the DNA binding activity and its activity of PARP (5Zahradka P. Ebisuzaki K. Eur. J. Biochem. 1982; 127: 579-585Crossref PubMed Scopus (146) Google Scholar). In we demonstrated that the 24-kDa fragment inhibited DNA repair, ADP-ribose polymer formation, and damage-dependent up-regulation of transcription mediated by of which shift the cell from survival to death of DNA repair by the 24-kDa fragment for by the that the 24-kDa fragment, full-length PARP DNA thus DNA repair from access to sites of DNA repair inhibited where PARP automodification is suppressed (6Satoh M.S. Lindahl T. Nature. 1992; 356: 356-358Crossref PubMed Scopus (975) Google Scholar), automodification is for the dissociation of PARP from DNA breaks. Alternatively, in cells of a PARP fragment containing the DNA binding and automodification site the catalytic also DNA repair to of the DNA breaks M. A. Ménissier-de Murcia J. J.H. de Murcia G. J. PubMed Scopus Google Scholar). As such the of cells to DNA-damaging agents (3Althaus F.R. Richter C. ADP-ribosylation of Proteins: Enzymology and Biological Significance. Springer-Verlag, Berlin1987: 3-113Google Scholar, M. A. Ménissier-de Murcia J. J.H. de Murcia G. J. PubMed Scopus Google Scholar). of DNA repair by binding of the 24-kDa fragment to DNA breaks may also cells to DNA-damaging Binding of the 24-kDa fragment to DNA breaks the formation of ADP-ribose polymers and the of NAD+ not has been suggested that of NAD+ by of PARP results in the rate of synthesis and cell death by Z. Wang Z.Q. Biol. 1999; PubMed Scopus Google Scholar, C. Trends Sci. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar, S.H. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: PubMed Scopus Google Scholar, K. K. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar, J. S. J. Cell Sci. 2000; Google Scholar). In of NAD+ and apoptosis and of Z. Wang Z.Q. Biol. 1999; PubMed Scopus Google Scholar, S.H. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: PubMed Scopus Google Scholar, K. K. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar, J. S. J. Cell Sci. 2000; Google Scholar). with these results also the that the 24-kDa fragment allows cells to an pathway by of The 24-kDa fragment of up-regulation of transcription We recently that PARP reduces the rate of transcript elongation by RNA polymerase II and that and automodification of as occurs in response to DNA damage, this thereby resulting in up-regulation of transcription (17Vispé S. Yung T.M.C. Ritchot J. Serizawa H. Satoh M.S. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 9886-9891Crossref PubMed Scopus (75) Google Scholar). Since DNA-damaging agents induce RNA damage as well, we proposed that such up-regulation allows cells to compensate for the loss of RNA following exposure to DNA-damaging agents, and that this pathway is required for cell The of such a pathway by the 24-kDa PARP fragment may therefore cells to DNA-damaging of the 24-kDa fragment in et al. J. S. J. Cell Sci. 2000; Google Scholar) found a in ADP-ribose polymer synthesis exposure of the cells to as well as of In the a in ADP-ribose polymer formation in the of about of the 24-kDa fragment in or about an of 24-kDa fragment to full-length PARP per In addition, with the from et al. J. S. J. Cell Sci. 2000; Google Scholar), we also found of apoptosis by in cells a of 24-kDa fragment to full-length PARP that addition of a of 24-kDa fragment to full-length PARP inhibited both DNA repair of full-length PARP from cell-free of the 24-kDa and up-regulation of transcription 2 of full-length PARP from nuclear of the 24-kDa by and the 24-kDa fragment, by competing with and in to may contribute to the by which cells commit to apoptosis A. J. Biochem. 1997; PubMed Scopus Google Scholar). We G. G. Poirier for the and the C. for and and for