Accumulation of Pax2 Transactivation Domain Interaction Protein (PTIP) at Sites of DNA Breaks via RNF8-dependent Pathway Is Required for Cell Survival after DNA Damage

Abstract

Genomic stability in eukaryotic cells is maintained by the coordination of multiple cellular events including cell cycle checkpoint, DNA repair, transcription, and apoptosis after DNA damage. Pax2 transactivation domain interaction protein (PTIP), a protein that contains six BRCT domains, has been implicated in DNA damage response. In this study we showed that recruitment of PTIP to damaged chromatin depends on DNA damage signaling proteins γH2AX·MDC1·RNF8, which in turn facilitates sustained localization of PA1 (PTIP-associated protein 1) to sites of DNA break. Similar to PTIP, depletion of PA1 increases cellular sensitivity to ionizing radiation. Furthermore, we demonstrated that the N-terminal PA1 binding domain and the C-terminal focus-localization domain of PTIP are critical for PTIP function in DNA damage repair. Interestingly, although PTIP and PA1 associate with MLL (mixed lineage leukemia) complexes and participate in transcriptional regulation, this function of PTIP·PA1 in DNA damage response is likely to be independent of the MLL complexes. Taken together, we propose that a subset of PTIP·PA1 complex is recruited to DNA damage sites via the RNF8-dependent pathway and is required for cell survival in response to DNA damage. Genomic stability in eukaryotic cells is maintained by the coordination of multiple cellular events including cell cycle checkpoint, DNA repair, transcription, and apoptosis after DNA damage. Pax2 transactivation domain interaction protein (PTIP), a protein that contains six BRCT domains, has been implicated in DNA damage response. In this study we showed that recruitment of PTIP to damaged chromatin depends on DNA damage signaling proteins γH2AX·MDC1·RNF8, which in turn facilitates sustained localization of PA1 (PTIP-associated protein 1) to sites of DNA break. Similar to PTIP, depletion of PA1 increases cellular sensitivity to ionizing radiation. Furthermore, we demonstrated that the N-terminal PA1 binding domain and the C-terminal focus-localization domain of PTIP are critical for PTIP function in DNA damage repair. Interestingly, although PTIP and PA1 associate with MLL (mixed lineage leukemia) complexes and participate in transcriptional regulation, this function of PTIP·PA1 in DNA damage response is likely to be independent of the MLL complexes. Taken together, we propose that a subset of PTIP·PA1 complex is recruited to DNA damage sites via the RNF8-dependent pathway and is required for cell survival in response to DNA damage. The genome of all living cells constantly suffers a variety of genomic insults, which if not fixed would lead to genomic instability. Therefore, in response to DNA damage, cells elicit an elaborated signaling network, which is collectively known as the DNA damage response pathway (1Harper J.W. Elledge S.J. Mol. Cell. 2007; 28: 739-745Abstract Full Text Full Text PDF PubMed Scopus (1255) Google Scholar). Through a cascade of sensors, transducers, and effectors, the DNA damage response pathway coordinates a process that include cell cycle checkpoints, DNA repair, cellular senescence, and apoptosis (2Sancar A. Lindsey-Boltz L.A. Unsal-Kacmaz K. Linn S. Annu. Rev. Biochem. 2004; 73: 39-85Crossref PubMed Scopus (2455) Google Scholar, 3Zhou B.B. Elledge S.J. Nature. 2000; 408: 433-439Crossref PubMed Scopus (2596) Google Scholar). The regulation of this pathway is best studied after ionizing radiation (IR). 2The abbreviations used are: IR, ionizing radiation; IRIF, IR-induced foci; PTIP, Pax2 transactivation domain interaction protein; PA1, PTIP-associated protein 1; siRNA, small interfering RNAs; MLL, mixed lineage leukemia; DAPI, 4′,6-diamidino-2-phenylindole; MEF, mouse embryonic fibroblast; HA, hemagglutinin; Gy, gray; UTX, ubiquitously transcribed tetratricopeptide repeat, X chromosome; ATM, ataxia telangiectasia mutated; ATR, ataxia telangiectasia and Rad3related; BRCT, BRCA1 carboxyl termini; FANCN, Fanconi anemia, complementation group N; DSB, double strand break. In response to IR, the histone variant H2AX is phosphorylated by ATM or ATR (4Rogakou E.P. Pilch D.R. Orr A.H. Ivanova V.S. Bonner W.M. J. Biol. Chem. 1998; 273: 5858-5868Abstract Full Text Full Text PDF PubMed Scopus (4046) Google Scholar) and serves as a platform for the recruitment of MDC1, which further facilitates the loading of many checkpoint and repair proteins to sites of DNA damage to form IR-induced foci (IRIF) (5Stucki M. Jackson S.P. DNA Repair (Amst). 2006; 5: 534-543Crossref PubMed Scopus (321) Google Scholar, 6Lou Z. Minter-Dykhouse K. Franco S. Gostissa M. Rivera M.A. Celeste A. Manis J.P. van Deursen J. Nussenzweig A. Paull T.T. Alt F.W. Chen J. Mol. Cell. 2006; 21: 187-200Abstract Full Text Full Text PDF PubMed Scopus (478) Google Scholar). More recent studies revealed that phosphorylation of MDC1 binds directly to and accumulates the E3 ubiquitin ligase complex, RNF8·Ubc13 at DNA damage sites, which ubiquitinates H2AX, H2A, and possibly additional proteins and allows the recruitment of 53BP1 and the ubiquitin-interacting motif domain-containing protein RAP80 at sites of breaks after DNA damage (7Huen M.S. Grant R. Manke I. Minn K. Yu X. Yaffe M.B. Chen J. Cell. 2007; 131: 901-914Abstract Full Text Full Text PDF PubMed Scopus (803) Google Scholar, 8Kim H. Chen J. Yu X. Science. 2007; 316: 1202-1205Crossref PubMed Scopus (434) Google Scholar, 9Wang B. Elledge S.J. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 20759-20763Crossref PubMed Scopus (361) Google Scholar, 10Wang B. Matsuoka S. Ballif B.A. Zhang D. Smogorzewska A. Gygi S.P. Elledge S.J. Science. 2007; 316: 1194-1198Crossref PubMed Scopus (553) Google Scholar, 11Mailand N. Bekker-Jensen S. Faustrup H. Melander F. Bartek J. Lukas C. Lukas J. Cell. 2007; 131: 887-900Abstract Full Text Full Text PDF PubMed Scopus (896) Google Scholar, 12Kolas N.K. Chapman J.R. Nakada S. Ylanko J. Chahwan R. Sweeney F.D. Panier S. Mendez M. Wildenhain J. Thomson T.M. Pelletier L. Jackson S.P. Durocher D. Science. 2007; 318: 1637-1640Crossref PubMed Scopus (694) Google Scholar, 13Sobhian B. Shao G. Lilli D.R. Culhane A.C. Moreau L.A. Xia B. Livingston D.M. Greenberg R.A. Science. 2007; 316: 1198-1202Crossref PubMed Scopus (542) Google Scholar). Although RAP80 specifically recruits BRCA1 to sites of DNA break, it remains to be determined how 53BP1 localizes to DNA damage sites via this ubiquitination-dependent cascade and whether additional DNA damage checkpoint and repair proteins would localize to sites of DNA break through the same or a similar mechanism. PTIP was originally identified as Pax2 transactivation domain interaction protein in a yeast two-hybrid screen (14Lechner M.S. Levitan I. Dressler G.R. Nucleic Acids Res. 2000; 28: 2741-2751Crossref PubMed Scopus (60) Google Scholar). Subsequently, several studies suggest that PTIP is an essential component of histone H3K4 methyltransferase complexes, which may be involved in the regulation of gene expression through modulation of H3K4 methylation (15Cho Y.W. Hong T. Hong S. Guo H. Yu H. Kim D. Guszczynski T. Dressler G.R. Copeland T.D. Kalkum M. Ge K. J. Biol. Chem. 2007; 282: 20395-20406Abstract Full Text Full Text PDF PubMed Scopus (400) Google Scholar, 16Patel S.R. Kim D. Levitan I. Dressler G.R. Dev. Cell. 2007; 13: 580-592Abstract Full Text Full Text PDF PubMed Scopus (168) Google Scholar). The importance of PTIP functions in vivo is revealed by the observations that PTIP null embryos die at E9.5 due to widespread cell death (17Cho E.A. Prindle M.J. Dressler G.R. Mol. Cell Biol. 2003; 23: 1666-1673Crossref PubMed Scopus (68) Google Scholar). In addition, PTIP null cells show a very high number of DNA breaks during S-phase and an inability to progress through mitosis, suggesting a defect in DNA repair (17Cho E.A. Prindle M.J. Dressler G.R. Mol. Cell Biol. 2003; 23: 1666-1673Crossref PubMed Scopus (68) Google Scholar). A possible function of PTIP in DNA damage repair is also suggested by the presence of six BRCT domains in PTIP, as BRCT domains are phosphoprotein binding domains that preferentially bind to Ser(P)/Thr(P) motifs, especially those generated by ATM and ATR (18Manke I.A. Lowery D.M. Nguyen A. Yaffe M.B. Science. 2003; 302: 636-639Crossref PubMed Scopus (542) Google Scholar). Indeed, PTIP has been shown to interact with 53BP1 in response to DNA damage, and this interaction requires ATM-dependent phosphorylation of 53BP1 (18Manke I.A. Lowery D.M. Nguyen A. Yaffe M.B. Science. 2003; 302: 636-639Crossref PubMed Scopus (542) Google Scholar, 19Jowsey P.A. Doherty A.J. Rouse J. J. Biol. Chem. 2004; 279: 55562-55569Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar). The four C-terminal BRCT domains of PTIP are required for its foci formation after IR and its interaction with 53BP1 via 53BP1 Ser-25 phosphorylation (19Jowsey P.A. Doherty A.J. Rouse J. J. Biol. Chem. 2004; 279: 55562-55569Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar). However, the exact function of PTIP in the DNA damage-responsive pathway and where it fits in the well defined DNA damage-signaling cascade remain elusive. Here we report that PTIP acts downstream of γH2AX·MDC1·RNF8 in the DNA damage signal transduction cascade. In addition, we showed that PTIP forms a stable complex with PTIP-associated protein 1 (PA1), and this complex is required for cell survival after IR. Cell Culture and Plasmids-HeLa, 293T, and U2OS cells maintained in with and at in a with embryonic cells in with and at in (15Cho Y.W. Hong T. Hong S. Guo H. Yu H. Kim D. Guszczynski T. Dressler G.R. Copeland T.D. Kalkum M. Ge K. J. Biol. Chem. 2007; 282: 20395-20406Abstract Full Text Full Text PDF PubMed Scopus (400) Google Scholar) with to PTIP and of the PTIP gene was by with H2AX, and cells and its cells with and and and and cells Z. Minter-Dykhouse K. Franco S. Gostissa M. Rivera M.A. Celeste A. Manis J.P. van Deursen J. Nussenzweig A. Paull T.T. Alt F.W. Chen J. Mol. Cell. 2006; 21: 187-200Abstract Full Text Full Text PDF PubMed Scopus (478) Google Scholar, M.S. Grant R. Manke I. Minn K. Yu X. Yaffe M.B. Chen J. Cell. 2007; 131: 901-914Abstract Full Text Full Text PDF PubMed Scopus (803) Google Scholar, R. PubMed Scopus Google Scholar, X. S. M. R. Chen J. Dev. 2006; PubMed Scopus Google Scholar, I. K. T. Chen J. J. Cell Biol. PubMed Scopus Google Scholar). and a of at cells by PTIP was and to N-terminal and binding or or the generated by and by or to the by with PA1 The was and a MDC1, Z. Minter-Dykhouse K. Franco S. Gostissa M. Rivera M.A. Celeste A. Manis J.P. van Deursen J. Nussenzweig A. Paull T.T. Alt F.W. Chen J. Mol. Cell. 2006; 21: 187-200Abstract Full Text Full Text PDF PubMed Scopus (478) Google Scholar, M.S. Grant R. Manke I. Minn K. Yu X. Yaffe M.B. Chen J. Cell. 2007; 131: 901-914Abstract Full Text Full Text PDF PubMed Scopus (803) Google Scholar, Y.W. Hong T. Hong S. Guo H. Yu H. Kim D. Guszczynski T. Dressler G.R. Copeland T.D. Kalkum M. Ge K. J. Biol. Chem. 2007; 282: 20395-20406Abstract Full Text Full Text PDF PubMed Scopus (400) Google Scholar, J. Chen J. 2006; PubMed Scopus Google Scholar). and and PTIP was the was and after of PTIP and cells and used to cells in the presence of cells as or in for the of stable The PTIP identified and by and on or with a and to for fixed in for and in for at with in at for with cells with or for at with and with with a and with 1 and 1 of A and on for of cell by the and with protein with PA1 or for at The with and in loading on and to and was with as Cell cell cycle cells with in of and fixed with the of of at fixed cells and in A for and with for on a and cell cycle was cells used for The stable cells with on for of cell by with for 1 at The proteins with and with for 1 at The further with for 1 at and with The proteins to to and by The of proteins revealed by by the interfering PTIP or PA1 A was used as The of PA1 and are and of and PTIP are and The of and are and The of H2AX is The for MDC1 and (7Huen M.S. Grant R. Manke I. Minn K. Yu X. Yaffe M.B. Chen J. Cell. 2007; 131: 901-914Abstract Full Text Full Text PDF PubMed Scopus (803) Google Scholar). The of is cells at with the to of sensitivity as Z. Minter-Dykhouse K. X. Chen J. Nature. 2003; PubMed Scopus Google Scholar). cells in and with ionizing radiation with for to to fixed and with and a with the of independent PTIP, or PA1 U2OS cells with of ionizing and for 1 checkpoint as Z. Minter-Dykhouse K. X. Chen J. Nature. 2003; PubMed Scopus Google Scholar). PA1 a PTIP-associated an to PTIP functions in DNA damage we a cell a and binding PTIP for the of PTIP binding the of PTIP-associated proteins revealed by In with recent report (15Cho Y.W. Hong T. Hong S. Guo H. Yu H. Kim D. Guszczynski T. Dressler G.R. Copeland T.D. Kalkum M. Ge K. J. Biol. Chem. 2007; 282: 20395-20406Abstract Full Text Full Text PDF PubMed Scopus (400) Google we identified PA1 as a PTIP-associated protein and to an interaction not PTIP and PA1 also proteins suggesting that proteins associate with in the binding of PA1 to PTIP was not after cells to IR how proteins may function together, we whether PTIP and PA1 expression would be in In addition, we also cells in by In we cells to the cell cycle by at shown in the of PTIP and PA1 that PTIP and PA1 are ubiquitously cell the on PTIP required for its interaction with PA1, we a of of PTIP with PA1 in revealed that the interaction PTIP and PA1 was by the of the BRCT domain of PTIP, suggesting that this N-terminal domain of PTIP is involved in its interaction with PA1 we that the of PA1, not its is critical for interaction and we showed that PA1 binds directly to PTIP not to PTIP or PTIP that although PTIP·PA1 interaction requires the domain of PTIP, this interaction in a PTIP PA1 to of DNA studies suggested that PTIP forms foci in response to ionizing radiation (19Jowsey P.A. Doherty A.J. Rouse J. J. Biol. Chem. 2004; 279: 55562-55569Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar). of the sensitivity of we cells Indeed, PTIP foci formation was after ionizing radiation also PTIP foci formation in cells with to DNA damage, including and suggesting that PTIP to a variety of we PA1 foci formation in cells with the same that DNA damage The that DNA PTIP and PA1 foci with those that PTIP and PA1 to sites of DNA IR-induced PA1 foci formation was in U2OS cells with PTIP PTIP the same in cells with or PA1 suggest that PTIP PA1 to sites of DNA damage. further PA1 and PA1 to not form DNA foci However, with PTIP, PA1 foci after IR localization of PA1 also requires the as PTIP of its N-terminal which has PA1 binding to PA1 to foci that a interaction PTIP and PA1 is required for PTIP to PA1 to sites of DNA PTIP of the in to DNA that PTIP is required for PA1 localization after DNA damage, we would be the signaling that PTIP localization to DNA damage in PTIP and PA1 in cells with H2AX, MDC1, or also used a of cell in known to be involved in DNA damage checkpoint response. shown in in to PTIP foci formation was in and that the localization of PTIP depends on a signaling cascade H2AX, MDC1, and the we PTIP localization in cells suggesting that ATM is not essential for PTIP localization after DNA damage. is of PTIP foci formation in cells or cells with the same cells with BRCA1 or that PTIP acts downstream of H2AX, MDC1, and is independent of BRCA1 or in the DNA damage-signaling cascade. PTIP foci formation would on several DNA repair and or shown in PTIP in all that PTIP foci formation is independent of repair proteins after IR. that PTIP requires we determined whether RNF8-dependent events are for PTIP at sites of DNA we not PTIP that for we cells Interestingly, of PTIP to DNA damage sites was in cells cells with not with or domain that loading of and its to protein at the of breaks are events for sustained of was to function with in a DNA damage-signaling pathway to of a number of checkpoint proteins at sites of DNA breaks (7Huen M.S. Grant R. Manke I. Minn K. Yu X. Yaffe M.B. Chen J. Cell. 2007; 131: 901-914Abstract Full Text Full Text PDF PubMed Scopus (803) Google Scholar, 12Kolas N.K. Chapman J.R. Nakada S. Ylanko J. Chahwan R. Sweeney F.D. Panier S. Mendez M. Wildenhain J. Thomson T.M. Pelletier L. Jackson S.P. Durocher D. Science. 2007; 318: 1637-1640Crossref PubMed Scopus (694) Google Scholar). with this we that PTIP foci formation was in and are required for the of PTIP at or the sites of DNA MLL in DNA with shown that PTIP with the mixed lineage and histone complexes (15Cho Y.W. Hong T. Hong S. Guo H. Yu H. Kim D. Guszczynski T. Dressler G.R. Copeland T.D. Kalkum M. Ge K. J. Biol. Chem. 2007; 282: 20395-20406Abstract Full Text Full Text PDF PubMed Scopus (400) Google Scholar, I. T. K. T. A. L. Mol. Cell. Biol. 2007; PubMed Scopus Google Scholar) and the that similar to PTIP, MLL complexes also participate in DNA damage response. a for MLL complexes in DNA damage we whether protein or MLL would be to form shown in UTX, of component of complex, localize to In addition, of has on PTIP or PA1 formation a and a MLL as a we a of PTIP and PA1 that is not with MLL complexes that PTIP with functions are in the Taken together, we that PTIP in DNA damage response in a that is independent of the MLL protein complexes. PTIP to DNA and report that the C-terminal BRCT domains of PTIP are for its localization to breaks (19Jowsey P.A. Doherty A.J. Rouse J. J. Biol. Chem. 2004; 279: 55562-55569Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar). the localization of PTIP, we used a of of of the N-terminal BRCT domains and and the very C-terminal BRCT domains and as well as the of PTIP on PTIP foci of domain or domain PTIP localization after IR observations suggest that this of BRCT domains, BRCT and is essential for PTIP localization to DNA The of PTIP localization on the of an protein in the localization of we showed that PTIP not bind to ubiquitin A study has shown that the BRCT domain of PTIP has binding (18Manke I.A. Lowery D.M. Nguyen A. Yaffe M.B. Science. 2003; 302: 636-639Crossref PubMed Scopus (542) Google suggesting that an phosphorylated protein may be required for PTIP localization after DNA damage. further that the of this BRCT domain is required for PTIP we the on the of BRCT localization of in in and in of PTIP to foci formation of PTIP in after DNA damage is with and that the of PTIP is required for PTIP localization in response to DNA damage. Furthermore, in with that PTIP is required for PA1 foci PTIP with of its or domain to PA1 to suggesting that PTIP localization to sites of DNA break is a for the localization of of PTIP and 53BP1 at DNA of is well that PTIP with 53BP1 after IR (18Manke I.A. Lowery D.M. Nguyen A. Yaffe M.B. Science. 2003; 302: 636-639Crossref PubMed Scopus (542) Google Scholar, 19Jowsey P.A. Doherty A.J. Rouse J. J. Biol. Chem. 2004; 279: 55562-55569Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar, P.A. R. Rouse J. Nucleic Acids Res. 2007; PubMed Scopus Google Scholar). the C-terminal BRCT domains of PTIP been shown to its interaction with 53BP1 as well as its localization after DNA damage (18Manke I.A. Lowery D.M. Nguyen A. Yaffe M.B. Science. 2003; 302: 636-639Crossref PubMed Scopus (542) Google Scholar, 19Jowsey P.A. Doherty A.J. Rouse J. J. Biol. Chem. 2004; 279: 55562-55569Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar, P.A. R. Rouse J. Nucleic Acids Res. 2007; PubMed Scopus Google we whether PTIP localization would on shown in PTIP foci formation was similar in and we also 53BP1 localization after IR in suggesting that although downstream of proteins are recruited to DNA damage sites through independent a of of PTIP, we showed that the interaction PTIP and 53BP1 was by of of the four C-terminal BRCT domains of PTIP suggesting that all four C-terminal BRCT domains of PTIP are for its interaction with the for 53BP1 binding is its which requires and PTIP and PA1 for Cell after PTIP is involved in DNA damage its function in this process remains to be the of PTIP in IR-induced checkpoint a a defect in IR-induced checkpoint However, cells showed checkpoint PTIP in cells also to a defect in checkpoint which is very similar to those in cells with PA1 that PTIP and PA1 may a in this checkpoint after DNA damage. we whether PTIP would be required for cell survival after IR. cells that or of PTIP as Although cells with PTIP was to cell survival after IR similar to cells or cells with the or of PTIP, cells with the or domain to suggest that PTIP is required for cell survival after IR and that the N-terminal domain and C-terminal or domains are critical for this function of Similar to PTIP, depletion of PA1 IR suggesting that the PTIP·PA1 complex is required for cell survival in response to DNA damage. PTIP is known to be involved in DNA damage response (19Jowsey P.A. Doherty A.J. Rouse J. J. Biol. Chem. 2004; 279: 55562-55569Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar, P.A. R. Rouse J. Nucleic Acids Res. 2007; PubMed Scopus Google Scholar). In this study we demonstrated that PTIP acts downstream of the well defined γH2AX·MDC1·RNF8 DNA damage signaling pathway The loading of PTIP at DNA damage foci on the protein at the sites of DNA PTIP further its binding PA1 via a interaction PTIP and we showed that the PTIP·PA1 complex is required for cell survival and checkpoint after IR. studies a of PTIP in the known DNA damage-signaling cascade. study also demonstrated that PTIP is required for cell survival and checkpoint after ionizing its in checkpoint is we show that the focus-localization of PTIP is critical for its function in cell survival IR and a that sustained of DNA damage and repair proteins at sites of DNA breaks are for DNA repair after DNA damage. we not how PTIP accumulates at DNA damage studies suggest that RNF8-dependent ubiquitin at DNA damage sites as sites for the recruitment of downstream proteins or protein complexes (7Huen M.S. Grant R. Manke I. Minn K. Yu X. Yaffe M.B. Chen J. Cell. 2007; 131: 901-914Abstract Full Text Full Text PDF PubMed Scopus (803) Google Scholar, 11Mailand N. Bekker-Jensen S. Faustrup H. Melander F. Bartek J. Lukas C. Lukas J. Cell. 2007; 131: 887-900Abstract Full Text Full Text PDF PubMed Scopus (896) Google Scholar, 12Kolas N.K. Chapman J.R. Nakada S. Ylanko J. Chahwan R. Sweeney F.D. Panier S. Mendez M. Wildenhain J. Thomson T.M. Pelletier L. Jackson S.P. Durocher D. Science. 2007; 318: 1637-1640Crossref PubMed Scopus (694) Google Scholar). ubiquitin-interacting protein RAP80 be recruited to DNA damage sites via its to bind to ubiquitin and serves as an for the further recruitment of BRCA1 H. Chen J. Yu X. Science. 2007; 316: 1202-1205Crossref PubMed Scopus (434) Google Scholar, 9Wang B. Elledge S.J. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 20759-20763Crossref PubMed Scopus (361) Google Scholar, 10Wang B. Matsuoka S. Ballif B.A. Zhang D. Smogorzewska A. Gygi S.P. Elledge S.J. Science. 2007; 316: 1194-1198Crossref PubMed Scopus (553) Google Scholar, 13Sobhian B. Shao G. Lilli D.R. Culhane A.C. Moreau L.A. Xia B. Livingston D.M. Greenberg R.A. Science. 2007; 316: 1198-1202Crossref PubMed Scopus (542) Google Scholar). The localization of PTIP is to a BRCT which is best known for its to bind to phosphorylated proteins (18Manke I.A. Lowery D.M. Nguyen A. Yaffe M.B. Science. 2003; 302: 636-639Crossref PubMed Scopus (542) Google Scholar). is that this BRCT domain of PTIP would interact directly with ubiquitin at DNA damage Indeed, we to ubiquitin binding of PTIP that PTIP recruitment may be very similar to the recruitment of BRCA1 as is identified protein that has ubiquitin binding similar to which allows its localization to DNA damage protein may be phosphorylated PTIP through a interaction via the PTIP The in the is to this and proteins and how proteins DNA damage signal transduction and proteins including PTIP in the DNA damage response. binding to PA1, PTIP also with In this study we showed that although PTIP and 53BP1 associate with (18Manke I.A. Lowery D.M. Nguyen A. Yaffe M.B. Science. 2003; 302: 636-639Crossref PubMed Scopus (542) Google Scholar, 19Jowsey P.A. Doherty A.J. Rouse J. J. Biol. Chem. 2004; 279: 55562-55569Abstract Full Text Full Text PDF PubMed Scopus (62) Google Scholar, P.A. R. Rouse J. Nucleic Acids Res. 2007; PubMed Scopus Google Scholar) and downstream of in the DNA damage-signaling cascade (7Huen M.S. Grant R. Manke I. Minn K. Yu X. Yaffe M.B. Chen J. Cell. 2007; 131: 901-914Abstract Full Text Full Text PDF PubMed Scopus (803) Google Scholar, 11Mailand N. Bekker-Jensen S. Faustrup H. Melander F. Bartek J. Lukas C. Lukas J. Cell. 2007; 131: 887-900Abstract Full Text Full Text PDF PubMed Scopus (896) Google Scholar, 12Kolas N.K. Chapman J.R. Nakada S. Ylanko J. Chahwan R. Sweeney F.D. Panier S. Mendez M. Wildenhain J. Thomson T.M. Pelletier L. Jackson S.P. Durocher D. Science. 2007; 318: 1637-1640Crossref PubMed Scopus (694) Google localize to sites of DNA damage that interaction at DNA damage sites may be for DNA repair However, we not an defect in cell survival after ionizing radiation in cells PTIP specifically in PTIP or suggest that an interaction PTIP and 53BP1 may be essential for a repair process and its be revealed by IR sensitivity used which of multiple DNA repair Indeed, we whether PTIP or PA1 would be involved in a DNA repair for repair. shown in we protein A foci foci and gene in or suggesting that this complex may not a in repair. that similar to PTIP·PA1 complex may participate in of studies PTIP whether similar to PTIP the would be involved in DNA repair all in the Chen especially for and also S. for the and the for