{"doi":"10.1074/jbc.m409293200","title":"Telosome, a Mammalian Telomere-associated Complex Formed by Multiple Telomeric Proteins","abstract":null,"journal":"Journal of Biological Chemistry","year":2004,"id":590141,"datarank":11.593735519686714,"base_score":5.993961427306569,"endowment":5.993961427306569,"self_citation_contribution":0.8990942140959856,"citation_network_contribution":10.694641305590729,"self_endowment_contribution":0.8990942140959856,"citer_contribution":10.694641305590729,"corpus_percentile":null,"corpus_rank":null,"citation_count":400,"citer_count":200,"citers_with_citation_signal":200,"citers_with_endowment":200,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":null,"is_data_producer":false,"deposit_databanks":null,"is_oa":false,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":null,"fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1509949,"name":"Matthew S. O'Connor","orcid":null,"position":1,"is_corresponding":false},{"id":473560,"name":"Jun Qin","orcid":"0000-0002-2263-128X","position":2,"is_corresponding":false},{"id":1182260,"name":"Zhou Songyang","orcid":"0000-0002-7824-3761","position":3,"is_corresponding":false},{"id":1122676,"name":"Dan Liu","orcid":"0000-0001-8789-0218","position":0,"is_corresponding":false}],"reference_count":0,"raw_metadata":{"has_enrichment":true,"resolved":true,"title":"Telosome, a Mammalian Telomere-associated Complex Formed by Multiple Telomeric Proteins","abstract":"In mammalian cells, telomere-binding proteins TRF1 and TRF2 play crucial roles in telomere biology. They interact with several other telomere regulators including TIN2, PTOP, POT1, and RAP1 to ensure proper maintenance of telomeres. TRF1 and TRF2 are believed to exert distinct functions. TRF1 forms a complex with TIN2, PTOP, and POT1 and regulates telomere length, whereas TRF2 mediates t-loop formation and end protection. However, whether cross-talk occurs between the TRF1 and TRF2 complexes and how the signals from these complexes are integrated for telomere maintenance remain to be elucidated. Through gel filtration and co-immunoprecipitation experiments, we found that TRF1 and TRF2 are in fact subunits of a telomere-associated high molecular weight complex (telosome) that also contains POT1, PTOP, RAP1, and TIN2. We demonstrated that the TRF1-interacting protein TIN2 binds TRF2 directly and in vivo, thereby bridging TRF2 to TRF1. Consistent with this multi-protein telosome model, stripping TRF1 off the telomeres by expressing tankyrase reduced telomere recruitment of not only TIN2 but also TRF2. These results help to unify previous observations and suggest that telomere maintenance depends on the multi-subunit telosome. In mammalian cells, telomere-binding proteins TRF1 and TRF2 play crucial roles in telomere biology. They interact with several other telomere regulators including TIN2, PTOP, POT1, and RAP1 to ensure proper maintenance of telomeres. TRF1 and TRF2 are believed to exert distinct functions. TRF1 forms a complex with TIN2, PTOP, and POT1 and regulates telomere length, whereas TRF2 mediates t-loop formation and end protection. However, whether cross-talk occurs between the TRF1 and TRF2 complexes and how the signals from these complexes are integrated for telomere maintenance remain to be elucidated. Through gel filtration and co-immunoprecipitation experiments, we found that TRF1 and TRF2 are in fact subunits of a telomere-associated high molecular weight complex (telosome) that also contains POT1, PTOP, RAP1, and TIN2. We demonstrated that the TRF1-interacting protein TIN2 binds TRF2 directly and in vivo, thereby bridging TRF2 to TRF1. Consistent with this multi-protein telosome model, stripping TRF1 off the telomeres by expressing tankyrase reduced telomere recruitment of not only TIN2 but also TRF2. These results help to unify previous observations and suggest that telomere maintenance depends on the multi-subunit telosome. The homeostasis of mammalian telomeres is regulated by a number of telomere-associated proteins. Among these proteins, TRF1 and TRF2 directly bind double-stranded telomere DNA and interact with a number of proteins to maintain telomere length and structure (1de Lange T. Oncogene. 2002; 21: 532-540Crossref PubMed Google Scholar, 2Kim Sh S.H. Kaminker P. Campisi J. Oncogene. 2002; 21: 503-511Crossref PubMed Google Scholar). It has been shown that the amount of telomere-bound TRF1 correlates with telomere length. Overexpression of TRF1 shortened telomeres in human cells, whereas dominant negative TRF1 led to elongated telomeres (3van Steensel B. de Lange T. Nature. 1997; 385: 740-743Crossref PubMed Scopus (1055) Google Scholar, 4Smith S. de Lange T. Curr. Biol. 2000; 10: 1299-1302Abstract Full Text Full Text PDF PubMed Scopus (347) Google Scholar, 5Smogorzewska A. van Steensel B. Bianchi A. Oelmann S. Schaefer M.R. Schnapp G. de Lange T. Mol. Cell. Biol. 2000; 20: 1659-1668Crossref PubMed Scopus (630) Google Scholar). TRF1 may control the length of telomere repeats through multiple mechanisms. For example, TRF1 can control telomerase access through its interaction with TIN2, PTOP/PIP1, and the single-stranded telomere DNA-binding protein POT1 (6Loayza D. De Lange T. Nature. 2003; 424: 1013-1018Crossref PubMed Scopus (544) Google Scholar, 7Liu D. Safari A. O'Connor M.S. Chan D.W. Laegeler A. Qin J. Songyang Z. Nat. Cell Biol. 2004; 6: 673-680Crossref PubMed Scopus (335) Google Scholar, 8Ye J.Z. Hockemeyer D. Krutchinsky A.N. Loayza D. Hooper S.M. Chait B.T. de Lange T. Genes Dev. 2004; 18: 1649-1654Crossref PubMed Scopus (351) Google Scholar). TRF1 may also regulate telomerase activity through its interaction with PINX1 (9Zhou X.Z. Lu K.P. Cell. 2001; 107: 347-359Abstract Full Text Full Text PDF PubMed Scopus (264) Google Scholar). In comparison, TRF2 has an essential role in telomere end protection and t-loop formation (1de Lange T. Oncogene. 2002; 21: 532-540Crossref PubMed Google Scholar, 10Griffith J.D. Comeau L. Rosenfield S. Stansel R.M. Bianchi A. Moss H. de Lange T. Cell. 1999; 97: 503-514Abstract Full Text Full Text PDF PubMed Scopus (1946) Google Scholar, 11Wei C. Price M. Cell Mol. Life Sci. 2003; 60: 2283-2294Crossref PubMed Scopus (44) Google Scholar). Interference of endogenous TRF2 activity by expressing dominant negative forms of TRF2 markedly increased the rate of telomere end-to-end fusions (12van Steensel B. Smogorzewska A. de Lange T. Cell. 1998; 92: 401-413Abstract Full Text Full Text PDF PubMed Scopus (1456) Google Scholar). Consistent with this role of TRF2, TRF2 forms a complex with RAP1 and associates with several proteins involved in DNA damage and repair responses, notably RAD50/MER11/NBS1, Ku86, and ERCC1/XPF (13Zhu X.D. Kuster B. Mann M. Petrini J.H. de Lange T. Nat. Genet. 2000; 25: 347-352Crossref PubMed Scopus (512) Google Scholar, 14Zhu X.D. Niedernhofer L. Kuster B. Mann M. Hoeijmakers J.H. de Lange T. Mol. Cell. 2003; 12: 1489-1498Abstract Full Text Full Text PDF PubMed Scopus (330) Google Scholar, 15O'Connor M.S. Safari A. Liu D. Qin J. Songyang Z. J. Biol. Chem. 2004; 279: 28585-28591Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar). These findings have pointed to distinct biological functions of TRF1 and TRF2. Some recent findings, however, suggest a more complex picture. For instance, overexpression of TRF2 caused telomere shortening in primary cells (16Karlseder J. Smogorzewska A. de Lange T. Science. 2002; 295: 2446-2449Crossref PubMed Scopus (668) Google Scholar). In mouse embryonic stem cells, the conditional knockout of TRF1 led to significantly reduced levels of TRF2 at the telomeres, suggesting that TRF2 telomere localization may be partially regulated by TRF1 (17Iwano T. Tachibana M. Reth M. Shinkai Y. J. Biol. Chem. 2003; 279: 1442-1448Abstract Full Text Full Text PDF PubMed Scopus (92) Google Scholar). In addition, chromosome end-to-end fusion was detected in TRF1 knock-out cells, indicating that telomere end protection was compromised. Despite the wealth of information, the functional relationship between TRF1 and TRF2 in telomere maintenance remains unclear. Notably, a recent report demonstrated a direct interaction between TRF2 and the TRF1-interacting protein, TIN2 (18Kim S.H. Beausejour C. Davalos A.R. Kaminker P. Heo S.J. Campisi J. J. Biol. Chem. 2004; 279: 43799-43804Abstract Full Text Full Text PDF PubMed Scopus (153) Google Scholar). Such findings further suggest that cross-talk probably occurs between the TRF1 and TRF2 complexes. However, whether TIN2 can simultaneously associate with both TRF1 and TRF2 in the same complex remains to be demonstrated. In addition to TRF1, several other telomeric proteins have been shown to be regulators of telomere length (1de Lange T. Oncogene. 2002; 21: 532-540Crossref PubMed Google Scholar, 2Kim Sh S.H. Kaminker P. Campisi J. Oncogene. 2002; 21: 503-511Crossref PubMed Google Scholar, 7Liu D. Safari A. O'Connor M.S. Chan D.W. Laegeler A. Qin J. Songyang Z. Nat. Cell Biol. 2004; 6: 673-680Crossref PubMed Scopus (335) Google Scholar, 8Ye J.Z. Hockemeyer D. Krutchinsky A.N. Loayza D. Hooper S.M. Chait B.T. de Lange T. Genes Dev. 2004; 18: 1649-1654Crossref PubMed Scopus (351) Google Scholar, 19Kim S.H. Kaminker P. Campisi J. Nat. Genet. 1999; 23: 405-412Crossref PubMed Scopus (429) Google Scholar, 20Li B. Oestreich S. de Lange T. Cell. 2000; 101: 471-483Abstract Full Text Full Text PDF PubMed Scopus (2) Google Scholar, 21Baumann P. Cech T.R. Science. 2001; 292: 1171-1175Crossref PubMed Scopus (810) Google Scholar, 22Colgin L.M. Baran K. Baumann P. Cech T.R. Reddel R.R. Curr. Biol. 2003; 13: 942-946Abstract Full Text Full Text PDF PubMed Scopus (182) Google Scholar). Both inhibition of endogenous RAP1, TIN2, POT1, or PTOP expression through RNA interference (RNAi) 1The abbreviations used are: RNAi, RNA interference; TANK, tankyrase; GST, glutathione S-transferase; PD, poly(ADP-ribose) polymerase-dead. and expression of dominant negative forms of these four proteins resulted in elongated telomeres in cultured cells (3van Steensel B. de Lange T. Nature. 1997; 385: 740-743Crossref PubMed Scopus (1055) Google Scholar, 6Loayza D. De Lange T. Nature. 2003; 424: 1013-1018Crossref PubMed Scopus (544) Google Scholar, 7Liu D. Safari A. O'Connor M.S. Chan D.W. Laegeler A. Qin J. Songyang Z. Nat. Cell Biol. 2004; 6: 673-680Crossref PubMed Scopus (335) Google Scholar, 8Ye J.Z. Hockemeyer D. Krutchinsky A.N. Loayza D. Hooper S.M. Chait B.T. de Lange T. Genes Dev. 2004; 18: 1649-1654Crossref PubMed Scopus (351) Google Scholar, 15O'Connor M.S. Safari A. Liu D. Qin J. Songyang Z. J. Biol. Chem. 2004; 279: 28585-28591Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar, 23Ye J.Z. de Lange T. Nat. Genet. 2004; 36: 618-623Crossref PubMed Scopus (155) Google Scholar). These observations suggest that RAP1, TIN2, POT1, and PTOP may function in the same pathway. All four proteins, RAP1, TIN2, POT1, and PTOP, directly or indirectly associate with TRF1 or TRF2 (7Liu D. Safari A. O'Connor M.S. Chan D.W. Laegeler A. Qin J. Songyang Z. Nat. Cell Biol. 2004; 6: 673-680Crossref PubMed Scopus (335) Google Scholar, 8Ye J.Z. Hockemeyer D. Krutchinsky A.N. Loayza D. Hooper S.M. Chait B.T. de Lange T. Genes Dev. 2004; 18: 1649-1654Crossref PubMed Scopus (351) Google Scholar, 20Li B. Oestreich S. de Lange T. Cell. 2000; 101: 471-483Abstract Full Text Full Text PDF PubMed Scopus (2) Google Scholar), pointing to a possible functional connection among these six telomeric proteins. In this report, we present evidence demonstrating that the TRF1 and TRF2 complexes do indeed interact with each other, as TRF1, TRF2, RAP1, TIN2, POT1, and PTOP can form a protein complex in vivo to regulate telomeres. Preparation of Nuclear Extracts—HeLa S3 cells grown in suspension to 1 × 106 cells/ml were collected and washed in cold phosphate-buffered saline and hypotonic buffer (10 mm Tris, pH 7.3, 10 mm KCl, 1.5 mm MgCl2, 0.2 mm phenylmethylsulfonyl fluoride, and 10 mm 2-mercaptoethanol). The cells were then allowed to swell for 15 min in hypotonic buffer, homogenized until cell membrane lysis was ∼80%. The lysates were resuspended in low salt buffer (20 mm KCl, 20 mm Tris, pH 7.3, 25% glycerol, 1.5 mm MgCl2, and 0.2 mm EDTA) and homogenized briefly to break the nuclear membrane. An equal volume of high salt buffer (1.2 m KCl, 20 mm Tris, pH 7.3, 25% glycerol, 1.5 mm MgCl2, and 0.2 mm EDTA) was added followed by agitation for 30 min at 4 °C and centrifuged at 20,000 × g for 30 min. The supernatant was dialyzed in BC0 buffer (20 mm Tris, pH 7.3, 20% glycerol, 0.2 mm EDTA, 0.2 mm phenylmethylsulfonyl fluoride, and 10 mm 2-mercaptoethanol) for 3 h and centrifuged again. The cleared supernatant was then aliquoted and stored at -80 °C. Salt extraction and fractionation of HT1080 cells were performed as previously described previously (23Ye J.Z. de Lange T. Nat. Genet. 2004; 36: 618-623Crossref PubMed Scopus (155) Google Scholar). The cells were extracted in a low salt buffer (20 mm Hepes pH 7.9, 5 mm MgCl2, 1 mm DTT, 25% glycerol, protease inhibitors, 0.2% Nonidet P-40, and 150 mm KCl). The resulting supernatant was the 150 mm fraction. The pellet was further extracted with a similar buffer but containing 420 mm KCl. Chromatin-bound proteins were in the 420 mm KCl fraction. Immunoprecipitation and Mass Spectrometry—For large-scale affinity purification, ∼70 mg of nuclear protein extracts were incubated with 100 μl of anti-FLAG M2-agarose beads (Sigma) for 3 h at 4 °C. The beads were then washed 4 times with NETN (20 mm Tris, pH 8.0, 100 mm NaCl, 0.5% Nonidet P-40, and 1 mm EDTA), and the bound protein was eluted twice with 100 μlof200 μg/ml FLAG peptide-(DYKDDDDK) (Sigma) in NETN. The eluent was resolved on a 8–12% SDS-PAGE gradient gel (Bio-Rad) and visualized by Coomassie Blue staining. Specific bands were then excised, digested with trypsin, and subjected to ion-trap mass spectrometry as previously described (24Ogryzko V.V. Kotani T. Zhang X. Schiltz R.L. Howard T. Yang X.J. Howard B.H. Qin J. Nakatani Y. Cell. 1998; 94: 35-44Abstract Full Text Full Text PDF PubMed Scopus (467) Google Scholar). Peptides were identified using PROWL (prowl.rockefeller.edu/). For small-scale immunoprecipitation experiments, 1 mg of nuclear extracts was incubated for 2 h at 4 °C with 5 μg of anti-FLAG M2 (Sigma), anti-hRap1 (Bethyl Laboratories), anti-TRF2 (Oncogene), anti-POT1N, anti-TIN2C, or anti-PTOP 466 antibodies (7Liu D. Safari A. O'Connor M.S. Chan D.W. Laegeler A. Qin J. Songyang Z. Nat. Cell Biol. 2004; 6: 673-680Crossref PubMed Scopus (335) Google Scholar) and 15 μl of protein A or protein G-agarose beads (Santa Cruz Biotechnology). The beads were then washed four times with 0.5 ml of NETN, boiled in 2× SDS loading buffer, and resolved on 8 or 10% SDS-PAGE   of the     were performed as described previously (7Liu D. Safari A. O'Connor M.S. Chan D.W. Laegeler A. Qin J. Songyang Z. Nat. Cell Biol. 2004; 6: 673-680Crossref PubMed Scopus (335) Google Scholar).  cell nuclear extracts were  on      gel filtration    The resulting  were resolved by SDS-PAGE and  with    of  and Cell  of  and      and    were  in the    as previously described  M.S. Safari A. Liu D. Qin J. Songyang Z. J. Biol. Chem. 2004; 279: 28585-28591Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar). The    and     were a   from   de Lange (23Ye J.Z. de Lange T. Nat. Genet. 2004; 36: 618-623Crossref PubMed Scopus (155) Google Scholar). The   were used to   cells to   for the   of  or HT1080  These cells were  with 2 μg/ml  for 3    to  cells  expressing  and its  or   and       and nuclear   were  by SDS-PAGE and  to    The primary antibodies  anti-FLAG M2 (Sigma) and anti-TRF2  The     was    human POT1 protein     anti-TIN2C, and anti-PTOP 466 antibodies were previously described (7Liu D. Safari A. O'Connor M.S. Chan D.W. Laegeler A. Qin J. Songyang Z. Nat. Cell Biol. 2004; 6: 673-680Crossref PubMed Scopus (335) Google Scholar). These antibodies were  by the     was a   from the de Lange  (3van Steensel B. de Lange T. Nature. 1997; 385: 740-743Crossref PubMed Scopus (1055) Google Scholar). The  antibodies     and     In       POT1, RAP1, and TIN2 were  using  beads    1 μg of  fusion proteins on beads was used for each   In   and   of human TRF1 and TRF2 were   using the In     The  were washed  times with NETN, eluted with 2× SDS buffer, resolved by  and  to    followed by  using a      localization of telomere-associated proteins were visualized through   as previously described  B. Oestreich S. de Lange T. Cell. 2000; 101: 471-483Abstract Full Text Full Text PDF PubMed Scopus (2) Google Scholar).  were grown  on    in the       in phosphate-buffered   with   in phosphate-buffered  and   in the    containing  mm  The cells were   for 1 h at  °C in     with  primary and   antibodies for 1 h each at   and then visualized  a     The primary antibodies used      and  anti-TRF2    antibodies were        and         of a     proteins can be    for   and  of   proteins by immunoprecipitation and mass  We   a   to  the molecular  that regulate human telomeres. In   of the  RAP1 protein  we identified several proteins that are  to interact with RAP1, including    and TRF2   (13Zhu X.D. Kuster B. Mann M. Petrini J.H. de Lange T. Nat. Genet. 2000; 25: 347-352Crossref PubMed Scopus (512) Google Scholar, 15O'Connor M.S. Safari A. Liu D. Qin J. Songyang Z. J. Biol. Chem. 2004; 279: 28585-28591Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar).  the  of the    TIN2 as a  of the RAP1 complex as  The same RAP1 complex was  to form  in the  of   suggesting that the  between the   were not  through DNA  not  The  of TIN2 in the  complex was   TIN2 is a TRF1-interacting protein  S.H. Kaminker P. Campisi J. Nat. Genet. 1999; 23: 405-412Crossref PubMed Scopus (429) Google Scholar).   and  of  proteins further  the  of a  protein  as we found TRF2 and RAP1 to  with TRF1, TIN2, PTOP, and POT1 (7Liu D. Safari A. O'Connor M.S. Chan D.W. Laegeler A. Qin J. Songyang Z. Nat. Cell Biol. 2004; 6: 673-680Crossref PubMed Scopus (335) Google Scholar).  mass spectrometry   the six proteins to be the   of the  complex (7Liu D. Safari A. O'Connor M.S. Chan D.W. Laegeler A. Qin J. Songyang Z. Nat. Cell Biol. 2004; 6: 673-680Crossref PubMed Scopus (335) Google Scholar).  TIN2, PTOP, and POT1 have been shown to complex with TRF1 (6Loayza D. De Lange T. Nature. 2003; 424: 1013-1018Crossref PubMed Scopus (544) Google Scholar, 7Liu D. Safari A. O'Connor M.S. Chan D.W. Laegeler A. Qin J. Songyang Z. Nat. Cell Biol. 2004; 6: 673-680Crossref PubMed Scopus (335) Google Scholar, 8Ye J.Z. Hockemeyer D. Krutchinsky A.N. Loayza D. Hooper S.M. Chait B.T. de Lange T. Genes Dev. 2004; 18: 1649-1654Crossref PubMed Scopus (351) Google Scholar),   that TRF2 and RAP1 may interact with the TRF1  resulting in the formation of a  complex at the telomeres.   the interaction between the six telomeric proteins, we   co-immunoprecipitation  using nuclear extracts from  cells and antibodies  endogenous POT1, PTOP, TIN2, RAP1, or TRF2. Consistent with  previous observations (7Liu D. Safari A. O'Connor M.S. Chan D.W. Laegeler A. Qin J. Songyang Z. Nat. Cell Biol. 2004; 6: 673-680Crossref PubMed Scopus (335) Google Scholar), endogenous POT1 and PTOP  with TIN2, whereas immunoprecipitation with anti-PTOP and  antibodies   POT1   Notably, TRF2 was also  to  with POT1, PTOP, and TIN2. In the   antibodies  TRF2 or RAP1   POT1 and TIN2 as  These     findings that TRF2 and RAP1 associate with the TRF1 complex and suggest a cross-talk between the TRF1 and TRF2 complexes. We performed gel filtration   using  nuclear   shown in   endogenous TRF1, TRF2, TIN2, RAP1, PTOP, and POT1  in a  molecular complex   indicating that the  proteins  indeed form a  complex that contains the  telomeric proteins identified to  in mammalian   on the   we  this   containing the six telomeric proteins, the telosome. It  be  that  of the telomeric proteins  RAP1 and  were also  at  molecular weight    may be other telomere complexes in addition to the telosome. TIN2   TRF2 Both in  and in  and  TRF2 to the TRF1    how the   was  to the TRF1  in the telosome. RAP1 contains an     a   and a   RAP1     B. Oestreich S. de Lange T. Cell. 2000; 101: 471-483Abstract Full Text Full Text PDF PubMed Scopus (2) Google Scholar). We    to   of the  of RAP1 were  for its  with TIN2.  shown in   anti-FLAG immunoprecipitation of  RAP1   endogenous TRF2 and TIN2. An  of a  of RAP1    in the   M.S. Safari A. Liu D. Qin J. Songyang Z. J. Biol. Chem. 2004; 279: 28585-28591Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar)  that the   and   were  in  RAP1 interaction with TRF2 and TIN2   However, the RAP1       to  with not only endogenous TRF2 but also TIN2, indicating that RAP1 may associate with TIN2 through TRF2.  these  suggest a direct interaction between TRF2 and TIN2  other  of the TRF1    this  TRF2 was in   and incubated with  fusion telomeric proteins.  shown in   in   TRF2  bound   a   to  but not   but not     in   TRF1      both TRF1 and TRF2 can directly interact with TIN2.    between  TRF2 and  TIN2 were detected in  cells  not  Consistent with a recent report on TIN2 interaction with TRF2 (18Kim S.H. Beausejour C. Davalos A.R. Kaminker P. Heo S.J. Campisi J. J. Biol. Chem. 2004; 279: 43799-43804Abstract Full Text Full Text PDF PubMed Scopus (153) Google Scholar),  results  that TIN2  the  between the TRF1 and TRF2 complexes.   of     by  further  the functional  of telosome as  as the interaction between TRF1 and TRF2, we  whether telomere localization of telosome  in  TRF2, was regulated by TRF1 using HT1080 cells expressing     or      is a  poly(ADP-ribose)   S.    A. de Lange T. Science. 1998;   PubMed Scopus  Google Scholar).  can  TRF1, resulting in TRF1  and  by the    stripping TRF1 off the telomeres  S.    A. de Lange T. Science. 1998;   PubMed Scopus  Google Scholar,      S. Genes Dev. 2003;   PubMed Scopus  Google Scholar). In these cells,  and  were  at   whereas  TIN2 and TRF2 levels were not reduced  not  We then  the levels of  TRF1, TRF2, and TIN2 in these cells using    TRF1, TRF2, and TIN2      of telomeric proteins    previously  (17Iwano T. Tachibana M. Reth M. Shinkai Y. J. Biol. Chem. 2003; 279: 1442-1448Abstract Full Text Full Text PDF PubMed Scopus (92) Google Scholar), telomere-bound TRF1 was  reduced in cells expressing     with  cells    the number and  of TIN2   significantly in    cells   The direct interaction between TIN2 and TRF2  that TRF2 telomere localization may be  in  cells as   in cells in  TRF1 levels were reduced  of   anti-TRF2  also    Consistent with this  the  of  TIN2 and TRF2 in  cells were also  as  by     These results are not only  with the  that TRF2 telomere localization was  in TRF1 knock-out cells (17Iwano T. Tachibana M. Reth M. Shinkai Y. J. Biol. Chem. 2003; 279: 1442-1448Abstract Full Text Full Text PDF PubMed Scopus (92) Google Scholar) but also  further  for the   telosome  In this   TRF1   telosome  thereby  telomere localization of TIN2 and TRF2.  telomere localization of TRF2 may   the formation of telosome. All six proteins, TRF1, TRF2, TIN2, RAP1, POT1, and PTOP, have been shown to   to the telomeres in mammalian   interference with  of the six proteins by  or dominant negative expression has been  to  telomere length or end  (3van Steensel B. de Lange T. Nature. 1997; 385: 740-743Crossref PubMed Scopus (1055) Google Scholar, 6Loayza D. De Lange T. Nature. 2003; 424: 1013-1018Crossref PubMed Scopus (544) Google Scholar, 7Liu D. Safari A. O'Connor M.S. Chan D.W. Laegeler A. Qin J. Songyang Z. Nat. Cell Biol. 2004; 6: 673-680Crossref PubMed Scopus (335) Google Scholar, 8Ye J.Z. Hockemeyer D. Krutchinsky A.N. Loayza D. Hooper S.M. Chait B.T. de Lange T. Genes Dev. 2004; 18: 1649-1654Crossref PubMed Scopus (351) Google Scholar,  Steensel B. Smogorzewska A. de Lange T. Cell. 1998; 92: 401-413Abstract Full Text Full Text PDF PubMed Scopus (1456) Google Scholar, 15O'Connor M.S. Safari A. Liu D. Qin J. Songyang Z. J. Biol. Chem. 2004; 279: 28585-28591Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar, 23Ye J.Z. de Lange T. Nat. Genet. 2004; 36: 618-623Crossref PubMed Scopus (155) Google Scholar).  these proteins probably are the   of telomere  In a recent  by    (18Kim S.H. Beausejour C. Davalos A.R. Kaminker P. Heo S.J. Campisi J. J. Biol. Chem. 2004; 279: 43799-43804Abstract Full Text Full Text PDF PubMed Scopus (153) Google Scholar), the   a direct interaction between TIN2 and TRF2. In this report, we demonstrated that  interaction  only   of the    six telomeric proteins are  to   a high molecular weight  the telosome.  this  was in      J.Z.     M. Loayza D.  Y. Krutchinsky A.N. Chait B.T. De Lange T. J. Biol. Chem. 2004; 279:  Full Text Full Text PDF PubMed Scopus  Google Scholar) also  the  of the  telomeric   is  with   The telosome   to   similar telomere   in human cells were   RAP1, POT1, PTOP, or TIN2 was  through  or dominant negative expression (3van Steensel B. de Lange T. Nature. 1997; 385: 740-743Crossref PubMed Scopus (1055) Google Scholar, 6Loayza D. De Lange T. Nature. 2003; 424: 1013-1018Crossref PubMed Scopus (544) Google Scholar, 7Liu D. Safari A. O'Connor M.S. Chan D.W. Laegeler A. Qin J. Songyang Z. Nat. Cell Biol. 2004; 6: 673-680Crossref PubMed Scopus (335) Google Scholar, 8Ye J.Z. Hockemeyer D. Krutchinsky A.N. Loayza D. Hooper S.M. Chait B.T. de Lange T. Genes Dev. 2004; 18: 1649-1654Crossref PubMed Scopus (351) Google Scholar, 15O'Connor M.S. Safari A. Liu D. Qin J. Songyang Z. J. Biol. Chem. 2004; 279: 28585-28591Abstract Full Text Full Text PDF PubMed Scopus (144) Google Scholar, 23Ye J.Z. de Lange T. Nat. Genet. 2004; 36: 618-623Crossref PubMed Scopus (155) Google Scholar).  of the dominant negative forms of a   the telosome may  its   and  of  of the six proteins by   probably  telosome   the  of telosome subunits may be crucial to its proper  For example, the  of TIN2 through  led to reduced TRF1 localization at the telomeres (23Ye J.Z. de Lange T. Nat. Genet. 2004; 36: 618-623Crossref PubMed Scopus (155) Google Scholar). In further  of a  role of the telosome in  telomere   of TIN2 or TRF1 in  resulted in embryonic   X.D. Niedernhofer L. Kuster B. Mann M. Hoeijmakers J.H. de Lange T. Mol. Cell. 2003; 12: 1489-1498Abstract Full Text Full Text PDF PubMed Scopus (330) Google Scholar,    S.  L. Campisi J.   Mol. Cell. Biol. 2004;   PubMed Scopus  Google Scholar). In the TRF1 knock-out  telomeric localization of TRF2 and TIN2 was also  (17Iwano T. Tachibana M. Reth M. Shinkai Y. J. Biol. Chem. 2003; 279: 1442-1448Abstract Full Text Full Text PDF PubMed Scopus (92) Google Scholar). Both TRF1 and TRF2 can bind telomeric double-stranded  The functional  between these  proteins is probably  to   to    complexes. TRF1  a primary role in telomere length control and cell  whereas TRF2  telomere  from   as DNA  It was  whether    between the TRF1 and TRF2 complexes.  results  that TRF1 and TRF2 interact with each other through TIN2 and  the functional connection between TRF1 and TRF2. The  of the telosome   essential  in telomere  telomere length, and end protection and    and functional cross-talk between its   similar to the   telosome  J.H.     Genes Dev.  6:  PubMed Scopus  Google Scholar,  K.  S.  S.M. Curr.  Cell Biol. 2001; 13:  PubMed Scopus  Google Scholar), the mammalian telosome may  the  telomere-associated complex  telomere maintenance in mammalian   of the six telomeric proteins may interact with  other  proteins to form    for the   and  of signals from   We   Chan and  Safari for","is_dataset_classified":null,"base_score":5.993961427306569,"endowment":5.993961427306569,"datacite_reuse_total":0,"file_count":0,"downloads":0,"views":0,"has_version_chain":false,"is_dataset":false,"is_oa":false,"pmid":"15383534","pmcid":null,"openalex_id":"https://openalex.org/W2051990601","authors":[],"funders":[],"total_grants":0,"fwci":8.1114,"citation_percentile":0.98329103,"influential_citations":0,"citation_trend":[{"year":2012,"count":22},{"year":2013,"count":16},{"year":2014,"count":25},{"year":2015,"count":19},{"year":2016,"count":19},{"year":2017,"count":15},{"year":2018,"count":19},{"year":2019,"count":16},{"year":2020,"count":11},{"year":2021,"count":15},{"year":2022,"count":13},{"year":2023,"count":10},{"year":2024,"count":8},{"year":2025,"count":9},{"year":2026,"count":3}],"oa_status":"hybrid","license":"cc-by","oa_locations":[{"url":"http://www.jbc.org/article/S0021925820694261/pdf","host_type":"journal"},{"url":"http://www.jbc.org/article/S0021925820694261/pdf","host_type":"publisher"},{"url":"https://api.elsevier.com/content/article/PII:S0021925820694261?httpAccept=text/xml","host_type":"publisher"},{"url":"https://api.elsevier.com/content/article/PII:S0021925820694261?httpAccept=text/plain","host_type":"publisher"},{"url":"https://syndication.highwire.org/content/doi/10.1074/jbc.M409293200","host_type":"publisher"},{"url":"https://doi.org/10.1074/jbc.m409293200","host_type":"journal"},{"url":"https://pubmed.ncbi.nlm.nih.gov/15383534","host_type":"repository"}],"fields_of_study":["Telomeres, Telomerase, and Senescence","Genetics, Aging, and Longevity in Model Organisms","Microplastics and Plastic Pollution","Blotting, Western","Cell Line","Cell Nucleus","Electrophoresis, Polyacrylamide Gel","Fluorescent Antibody Technique, Indirect","Gene Deletion","HeLa Cells","Humans","Immunoprecipitation","Mass Spectrometry","Microscopy, Fluorescence","Multiprotein Complexes","Mutation","Protein Binding","Protein Structure, Tertiary","Shelterin Complex","Tankyrases","Telomere","Telomere-Binding Proteins","Telomeric Repeat Binding Protein 1","Telomeric Repeat Binding Protein 2","rap1 GTP-Binding Proteins"],"mesh_terms":["Shelterin Complex","Cell Line","Cell Nucleus","Electrophoresis, Polyacrylamide Gel","HeLa Cells","Humans","Microscopy, Fluorescence","Mutation","Protein Binding","Mass Spectrometry","Blotting, Western","Telomere","Gene Deletion","Protein Structure, Tertiary","Fluorescent Antibody Technique, Indirect","rap1 GTP-Binding Proteins","Telomere-Binding Proteins","Telomeric Repeat Binding Protein 1","Telomeric Repeat Binding Protein 2","Tankyrases","Multiprotein Complexes","Immunoprecipitation","Hela Cells"],"keywords":["Telomere","Rap1","Shelterin","Telomere-binding protein","Telomerase","Biology","Cell biology","Protein subunit","Molecular biology","DNA-binding protein","Genetics","DNA","Gene","Signal transduction","Transcription factor"],"sdg_mappings":[],"linked_datasets":[],"clinical_trials":[],"software_tools":[],"database_accessions":[],"source":"live","citation_network_status":"fetched"},"created_at":"2026-07-24T13:44:37.581560Z","pmid":null,"pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}