Susan L Smith, Ph.D.

TIN2 Stability Is Regulated by the E3 Ligase Siah2
Bhanot, Monica; Smith, Susan
2012 JAN ;32(2):376-384, Molecular & cellular biology
Telomeres are coated by shelterin, a six-subunit complex that is required for protection and replication of chromosome ends. The central subunit TIN2, with binding sites to three subunits (TRF1, TRF2, and TPP1), is essential for stability and function of the complex. Here we show that TIN2 stability is regulated by the E3 ligase Siah2. We demonstrate that TIN2 binds to Siah2 and is ubiquitylated in vivo. We show using purified proteins that Siah2 acts as an E3 ligase to directly ubiquitylate TIN2 in vitro. Depletion of Siah2 led to stabilization of TIN2 protein, indicating that Siah2 regulates TIN2 protein levels in vivo. Overexpression of Siah2 in human cells led to loss of TIN2 at telomeres that was dependent on the presence of the catalytic RING domain of Siah2. In contrast to RNAi-mediated depletion of TIN2 that led to loss of TRF1 and TRF2 at telomeres, Siah2-mediated depletion of TIN2 allowed TRF1 and TRF2 to remain on telomeres, indicating a different fate for shelterin subunits when TIN2 is depleted posttranslationally. TPP1 was lost from telomeres, although its protein level was not reduced. We speculate that Siah2-mediated removal of TIN2 may allow dynamic remodeling of the shelterin complex and its associated factors during the cell cycle
— id: 150790, year: 2012, vol: 32, page: 376, stat: Journal Article,

A role for sister telomere cohesion in telomere elongation by telomerase
Houghtaling, Benjamin R; Canudas, Silvia; Smith, Susan
2012 Jan 1;11(1):19-25, Cell cycle
Telomere length homeostasis is achieved by a balance of telomere shortening caused by DNA replication and nucleolytic attack and telomere lengthening by telomerase. The importance of telomere length maintenance to human health is best illustrated by dyskeratosis congenita (DC) a disease of telomere shortening caused by mutations in telomerase subunits. DC patients suffer stem cell depletion and die of bone marrow stem cell failure. Recently a new class of particularly severe DC patients was found to harbor mutations in the shelterin subunit TIN2. The DC-TIN2 mutations were clustered in small domain of unknown function. In a recently published study we showed that the DC mutation cluster in TIN2 harbored a binding site for heterochromatin protein 1 (HP1) and further, that HP1 binding to TIN2 was required for sister telomere cohesion in S phase and for telomere length maintenance by telomerase. We briefly review and discuss the implications of our findings in this Extra View, and present some new data that may shed light on how sister telomere cohesion could influence telomere elongation by telomerase
— id: 149956, year: 2012, vol: 11, page: 19, stat: Journal Article,

Poly(ADP-ribose) polymerase 3 (PARP3), a newcomer in cellular response to DNA damage and mitotic progression
Boehler, Christian; Gauthier, Laurent R; Mortusewicz, Oliver; Biard, Denis S; Saliou, Jean-Michel; Bresson, Anne; Sanglier-Cianferani, Sarah; Smith, Susan; Schreiber, Valerie; Boussin, Francois; Dantzer, Francoise
2011 Feb 15;108(7):2783-2788, Proceedings of the National Academy of Sciences of the United States of America
The ADP ribosyl transferase [poly(ADP-ribose) polymerase] ARTD3(PARP3) is a newly characterized member of the ARTD(PARP) family that catalyzes the reaction of ADP ribosylation, a key posttranslational modification of proteins involved in different signaling pathways from DNA damage to energy metabolism and organismal memory. This enzyme shares high structural similarities with the DNA repair enzymes PARP1 and PARP2 and accordingly has been found to catalyse poly(ADP ribose) synthesis. However, relatively little is known about its in vivo cellular properties. By combining biochemical studies with the generation and characterization of loss-of-function human and mouse models, we describe PARP3 as a newcomer in genome integrity and mitotic progression. We report a particular role of PARP3 in cellular response to double-strand breaks, most likely in concert with PARP1. We identify PARP3 as a critical player in the stabilization of the mitotic spindle and in telomere integrity notably by associating and regulating the mitotic components NuMA and tankyrase 1. Both functions open stimulating prospects for specifically targeting PARP3 in cancer therapy
— id: 134153, year: 2011, vol: 108, page: 2783, stat: Journal Article,

A role for heterochromatin protein 1{gamma} at human telomeres
Canudas, Silvia; Houghtaling, Benjamin R; Bhanot, Monica; Sasa, Ghadir; Savage, Sharon A; Bertuch, Alison A; Smith, Susan
2011 Sep 1;25(17):1807-1819, Genes & development
Human telomere function is mediated by shelterin, a six-subunit complex that is required for telomere replication, protection, and cohesion. TIN2, the central component of shelterin, has binding sites to three subunits: TRF1, TRF2, and TPP1. Here we identify a fourth partner, heterochromatin protein 1gamma (HP1gamma), that binds to a conserved canonical HP1-binding motif, PXVXL, in the C-terminal domain of TIN2. We show that HP1gamma localizes to telomeres in S phase, where it is required to establish/maintain cohesion. We further demonstrate that the HP1-binding site in TIN2 is required for sister telomere cohesion and can impact telomere length maintenance by telomerase. Remarkably, the PTVML HP1-binding site is embedded in the recently identified cluster of mutations in TIN2 that gives rise to dyskeratosis congenita (DC), an inherited bone marrow failure syndrome caused by defects in telomere maintenance. We show that DC-associated mutations in TIN2 abrogate binding to HP1gamma and that DC patient cells are defective in sister telomere cohesion. Our data indicate a novel requirement for HP1gamma in the establishment/maintenance of cohesion at human telomeres and, furthermore, may provide insight into the mechanism of pathogenesis in TIN2-mediated DC
— id: 137073, year: 2011, vol: 25, page: 1807, stat: Journal Article,

Differential regulation of telomere and centromere cohesion by the Scc3 homologues SA1 and SA2, respectively, in human cells
Canudas, Silvia; Smith, Susan
2009 Oct 19;187(2):165-173, Journal of cell biology
Replicated sister chromatids are held together until mitosis by cohesin, a conserved multisubunit complex comprised of Smc1, Smc3, Scc1, and Scc3, which in vertebrate cells exists as two closely related homologues (SA1 and SA2). Here, we show that cohesin(SA1) and cohesin(SA2) are differentially required for telomere and centromere cohesion, respectively. Cells deficient in SA1 are unable to establish or maintain cohesion between sister telomeres after DNA replication in S phase. The same phenotype is observed upon depletion of the telomeric protein TIN2. In contrast, in SA2-depleted cells telomere cohesion is normal, but centromere cohesion is prematurely lost. We demonstrate that loss of telomere cohesion has dramatic consequences on chromosome morphology and function. In the absence of sister telomere cohesion, cells are unable to repair chromatid breaks and suffer sister telomere loss. Our studies elucidate the functional distinction between the Scc3 homologues in human cells and further reveal an essential role for sister telomere cohesion in genomic integrity
— id: 104723, year: 2009, vol: 187, page: 165, stat: Journal Article,

Sister telomeres rendered dysfunctional by persistent cohesion are fused by NHEJ
Hsiao, Susan J; Smith, Susan
2009 Feb 23;184(4):515-526, Journal of cell biology
Telomeres protect chromosome ends from being viewed as double-strand breaks and from eliciting a DNA damage response. Deprotection of chromosome ends occurs when telomeres become critically short because of replicative attrition or inhibition of TRF2. In this study, we report a novel form of deprotection that occurs exclusively after DNA replication in S/G2 phase of the cell cycle. In cells deficient in the telomeric poly(adenosine diphosphate ribose) polymerase tankyrase 1, sister telomere resolution is blocked. Unexpectedly, cohered sister telomeres become deprotected and are inappropriately fused. In contrast to telomeres rendered dysfunctional by TRF2, which engage in chromatid fusions predominantly between chromatids from different chromosomes (Bailey, S.M., M.N. Cornforth, A. Kurimasa, D.J. Chen, and E.H. Goodwin. 2001. Science. 293:2462-2465; Smogorzewska, A., J. Karlseder, H. Holtgreve-Grez, A. Jauch, and T. de Lange. 2002. Curr. Biol. 12:1635-1644), telomeres rendered dysfunctional by tankyrase 1 engage in chromatid fusions almost exclusively between sister chromatids. We show that cohered sister telomeres are fused by DNA ligase IV-mediated nonhomologous end joining. These results demonstrate that the timely removal of sister telomere cohesion is essential for the formation of a protective structure at chromosome ends after DNA replication in S/G2 phase of the cell cycle
— id: 93576, year: 2009, vol: 184, page: 515, stat: Journal Article,

The long and short of it: a new isoform of TIN2 in the nuclear matrix
Smith, Susan
2009 Mar 15;8(6):797-798, Cell cycle
— id: 137817, year: 2009, vol: 8, page: 797, stat: Journal Article,

The SAGA continues...to the end
Smith, Susan
2009 Aug 14;35(3):256-258, Molecular cell
In this issue of Molecular Cell, Atanassov et al. (2009) show that the GCN5-containing SAGA complex regulates telomere function via deubiquitination and stabilization of the telomere repeat binding factor TRF1
— id: 101646, year: 2009, vol: 35, page: 256, stat: Journal Article,

Tankyrase 1 and tankyrase 2 are essential but redundant for mouse embryonic development
Chiang, Y Jeffrey; Hsiao, Susan J; Yver, Dena; Cushman, Samuel W; Tessarollo, Lino; Smith, Susan; Hodes, Richard J
2008 ;3(7):e2639-e2639, PLoS ONE
Tankyrases are proteins with poly(ADP-ribose) polymerase activity. Human tankyrases post-translationally modify multiple proteins involved in processes including maintenance of telomere length, sister telomere association, and trafficking of glut4-containing vesicles. To date, however, little is known about in vivo functions for tankyrases. We recently reported that body size was significantly reduced in mice deficient for tankyrase 2, but that these mice otherwise appeared developmentally normal. In the present study, we report generation of tankyrase 1-deficient and tankyrase 1 and 2 double-deficient mice, and use of these mutant strains to systematically assess candidate functions of tankyrase 1 and tankyrase 2 in vivo. No defects were observed in development, telomere length maintenance, or cell cycle regulation in tankyrase 1 or tankyrase 2 knockout mice. In contrast to viability and normal development of mice singly deficient in either tankyrase, deficiency in both tankyrase 1 and tankyrase 2 results in embryonic lethality by day 10, indicating that there is substantial redundancy between tankyrase 1 and tankyrase 2, but that tankyrase function is essential for embryonic development
— id: 88580, year: 2008, vol: 3, page: e2639, stat: Journal Article,

Tankyrase function at telomeres, spindle poles, and beyond
Hsiao, Susan J; Smith, Susan
2008 Jan;90(1):83-92, Biochimie
Telomeres have special needs; they require distinct mechanisms for their protection, replication, and separation at mitosis. A dedicated six-subunit protein complex termed shelterin attends to these needs. But shelterin cannot do it alone and often relies on recruits from other cellular locales. One such recruit is tankyrase 1, a poly(ADP-ribose) polymerase that is brought to telomeres by the shelterin DNA binding subunit TRF1, where it functions in telomere length regulation and sister chromatid separation. An understanding of how tankyrase 1 functions at telomeres has been confounded by its complexity; it localizes to multiple subcellular sites, it has many diverse binding partners, and it has a closely related homolog (tankyrase 2) with which it may functionally overlap. This review summarizes our current knowledge of tankyrases focusing on their localization, binding partners, and function
— id: 76329, year: 2008, vol: 90, page: 83, stat: Journal Article,

Protein requirements for sister telomere association in human cells
Canudas, Silvia; Houghtaling, Benjamin R; Kim, Ju Youn; Dynek, Jasmin N; Chang, William G; Smith, Susan
2007 Nov 28;26(23):4867-4878, EMBO journal
Previous studies in human cells indicate that sister telomeres have distinct requirements for their separation at mitosis. In cells depleted for tankyrase 1, a telomeric poly(ADP-ribose) polymerase, sister chromatid arms and centromeres separate normally, but telomeres remain associated and cells arrest in mitosis. Here, we use biochemical and genetic approaches to identify proteins that might mediate the persistent association at sister telomeres. We use immunoprecipitation analysis to show that the telomeric proteins, TRF1 (an acceptor of PARsylation by tankyrase 1) and TIN2 (a TRF1 binding partner) each bind to the SA1 ortholog of the cohesin Scc3 subunit. Sucrose gradient sedimentation shows that TRF1 cosediments with the SA1-cohesin complex. Depletion of the SA1 cohesin subunit or the telomeric proteins (TRF1 and TIN2) restores the normal resolution of sister telomeres in mitosis in tankyrase 1-depleted cells. Moreover, depletion of TRF1 and TIN2 or SA1 abrogates the requirement for tankyrase 1 in mitotic progression. Our studies indicate that sister telomere association in human cells is mediated by a novel association between a cohesin subunit and components of telomeric chromatin
— id: 75398, year: 2007, vol: 26, page: 4867, stat: Journal Article,

Tankyrase 2 poly(ADP-ribose) polymerase domain-deleted mice exhibit growth defects but have normal telomere length and capping
Hsiao, Susan J; Poitras, Marc F; Cook, Brandoch D; Liu, Yie; Smith, Susan
2006 Mar;26(6):2044-2054, Molecular & cellular biology
Regulation of telomere length maintenance and capping are a critical cell functions in both normal and tumor cells. Tankyrase 2 (Tnks2) is a poly(ADP-ribose) polymerase (PARP) that has been shown to modify itself and TRF1, a telomere-binding protein. We show here by overexpression studies that tankyrase 2, like its closely related homolog tankyrase 1, can function as a positive regulator of telomere length in human cells, dependent on its catalytic PARP activity. To study the role of Tnks2 in vivo, we generated mice with the Tnks2 PARP domain deleted. These mice are viable and fertile but display a growth retardation phenotype. Telomere analysis by quantitative fluorescence in situ hybridization (FISH), flow-FISH, and restriction fragment analysis showed no change in telomere length or telomere capping in these mice. To determine the requirement for Tnks2 in long-term maintenance of telomeres, we generated embryonic stem cells with the Tnks2 PARP domain deleted and observed no change, even upon prolonged growth, in telomere length or telomere capping. Together, these results suggest that Tnks2 has a role in normal growth and development but is not essential for telomere length maintenance or telomere capping in mice
— id: 64582, year: 2006, vol: 26, page: 2044, stat: Journal Article,

NuMA is a major acceptor of poly(ADP-ribosyl)ation by tankyrase 1 in mitosis
Chang, William; Dynek, Jasmin N; Smith, Susan
2005 Oct 15;391(2):177-184, Biochemical journal
Tankyrase 1 is a PARP [poly(ADP-ribose) polymerase] that localizes to multiple subcellular sites, including telomeres and mitotic centrosomes. Previous studies demonstrated that cells deficient in tankyrase 1 suffered a block in resolution of sister telomeres and arrested in early anaphase [Dynek and Smith (2004) Science 304, 97-100]. This phenotype was dependent on the catalytic PARP activity of tankyrase 1. To identify critical acceptors of PARsylation [poly(ADP-ribosyl)ation] by tankyrase 1 in mitosis, tankyrase 1 immunoprecipitates were analysed for associated PARsylated proteins. We identified NuMA (nuclear mitotic apparatus protein) as a major acceptor of poly(ADP-ribose) from tankyrase 1 in mitosis. We showed by immunofluorescence and immunoprecipitation that association between tankyrase 1 and NuMA increases dramatically at the onset of mitosis, concomitant with PARsylation of NuMA. Knockdown of tankyrase 1 by siRNA (small interfering RNA) eliminates PARsylation of NuMA in mitosis, confirming tankyrase 1 as the PARP responsible for this modification. However, even in the absence of tankyrase 1 and PARsylation, NuMA localizes to spindle poles. By contrast, siRNA knockdown of NuMA results in complete loss of tankyrase 1 from spindle poles. We discuss our result in terms of a model where PARsylation of NuMA by tankyrase 1 in mitosis could play a role in sister telomere separation and/or mitotic progression
— id: 58149, year: 2005, vol: 391, page: 177, stat: Journal Article,

Resolution of sister telomere association is required for progression through mitosis
Dynek, Jasmin N; Smith, Susan
2004 Apr 2;304(5667):97-100, Science
Cohesins keep sister chromatids associated from the time of their replication in S phase until the onset of anaphase. In vertebrate cells, two distinct pathways dissociate cohesins, one acts on chromosome arms and the other on centromeres. Here, we describe a third pathway that acts on telomeres. Knockdown of tankyrase 1, a telomeric poly(ADP-ribose) polymerase caused mitotic arrest. Chromosomes aligned normally on the metaphase plate but were unable to segregate. Sister chromatids separated at centromeres and arms but remained associated at telomeres, apparently through proteinaceous bridges. Thus, telomeres may require a unique tankyrase 1-dependent mechanism for sister chromatid resolution before anaphase
— id: 42576, year: 2004, vol: 304, page: 97, stat: Journal Article,

A dynamic molecular link between the telomere length regulator TRF1 and the chromosome end protector TRF2
Houghtaling, Benjamin R; Cuttonaro, Leanora; Chang, William; Smith, Susan
2004 Oct 21;14(18):1621-1631, Current biology. CB
BACKGROUND: Human telomeres are coated by the telomere repeat binding proteins TRF1 and TRF2, which are believed to function independently to regulate telomere length and protect chromosome ends, respectively. RESULTS: Here, we show that TRF1 and TRF2 are linked via TIN2, a previously identified TRF1-interacting protein, and its novel binding partner TINT1. TINT1 localized to telomeres via TIN2, where it functioned as a negative regulator of telomerase-mediated telomere elongation. TIN2 associated with TINT1, and TRF1 or TRF2 throughout the cell cycle, revealing a partially redundant unit in telomeric chromatin that may provide flexibility in telomere length control. Indeed, when TRF1 was removed from telomeres by overexpression of the positive telomere length regulator tankyrase 1, the TIN2/TINT1 complex remained on telomeres via an increased association with TRF2. CONCLUSIONS: Our findings suggest a dynamic cross talk between TRF1 and TRF2 and provide a molecular mechanism for telomere length homeostasis by TRF2 in the absence of TRF1
— id: 46295, year: 2004, vol: 14, page: 1621, stat: Journal Article,

Functional subdomain in the ankyrin domain of tankyrase 1 required for poly(ADP-ribosyl)ation of TRF1 and telomere elongation
Seimiya, Hiroyuki; Muramatsu, Yukiko; Smith, Susan; Tsuruo, Takashi
2004 Mar;24(5):1944-1955, Molecular & cellular biology
In human cells, telomere elongation by telomerase is repressed in cis by the telomeric protein TRF1. Tankyrase 1 binds TRF1 via its ankyrin domain and poly(ADP-ribosyl)ates it. Overexpression of tankyrase 1 in telomerase-positive cells releases TRF1 from telomeres, resulting in telomere elongation. The tankyrase 1 ankyrin domain is classified into five conserved subdomains, ARCs (ankyrin repeat clusters) I to V. Here, we investigated the biological significance of the ARCs. First, each ARC worked as an independent binding site for TRF1. Second, ARCs II to V recognized the N-terminal acidic domain of TRF1 whereas ARC I bound a discrete site between the homodimerization and the Myb-like domains of TRF1. Inactivation of TRF1 binding in the C-terminal ARC, ARC V, either by deletion or point mutation, significantly reduced the ability of tankyrase 1 to poly(ADP-ribosyl)ate TRF1, release TRF1 from telomeres, and elongate telomeres. In contrast, other ARCs, ARC II and/or IV, inactivated by point mutations still retained the biological function of tankyrase 1. On the other hand, ARC V per se was not sufficient for telomere elongation, suggesting a structural role for multiple ARCs. This work provides evidence that specific ARC-TRF1 interactions play roles in the essential catalytic function of tankyrase 1
— id: 137818, year: 2004, vol: 24, page: 1944, stat: Journal Article,

TRF1 is degraded by ubiquitin-mediated proteolysis after release from telomeres
Chang, William; Dynek, Jasmin N; Smith, Susan
2003 Jun 1;17(11):1328-1333, Genes & development
Mammalian telomeres are coated by the sequence-specific, DNA-binding protein, TRF1, a negative regulator of telomere length. Previous results showed that ADP-ribosylation of TRF1 by tankyrase 1 released TRF1 from telomeres and promoted telomere elongation. We now show that loss of TRF1 from telomeres results in ubiquitination and degradation of TRF1 by the proteasome and that degradation is required to keep TRF1 off telomeres. Ubiquitination of TRF1 is regulated by its telomere-binding status; only the telomere-unbound form of TRF1 is ubiquitinated. Our findings suggest a novel mechanism of sequential post translational modification of TRF1 (ADP-ribosylation and ubiquitination) for regulating access of telomerase to telomeres
— id: 36833, year: 2003, vol: 17, page: 1328, stat: Journal Article,

Role for the Related Poly(ADP-Ribose) Polymerases Tankyrase 1 and 2 at Human Telomeres
Cook, Brandoch D; Dynek, Jasmin N; Chang, William; Shostak, Grigoriy; Smith, Susan
2002 Jan 1;22(1):332-342, Molecular & cellular biology
Telomere maintenance is essential for the continuous growth of tumor cells. In most human tumors telomeres are maintained by telomerase, a specialized reverse transcriptase. Tankyrase 1, a human telomeric poly(ADP-ribose) polymerase (PARP), positively regulates telomere length through its interaction with TRF1, a telomeric DNA-binding protein. Tankyrase 1 ADP-ribosylates TRF1, inhibiting its binding to telomeric DNA. Overexpression of tankyrase 1 in the nucleus promotes telomere elongation, suggesting that tankyrase 1 regulates access of telomerase to the telomeric complex. The recent identification of a closely related homolog of tankyrase 1, tankyrase 2, opens the possibility for a second PARP at telomeres. We therefore sought to establish the role of tankyrase 1 at telomeres and to determine if tankyrase 2 might have a telomeric function. We show that endogenous tankyrase 1 is a component of the human telomeric complex. We demonstrate that telomere elongation by tankyrase 1 requires the catalytic activity of the PARP domain and does not occur in telomerase-negative primary human cells. To investigate a potential role for tankyrase 2 at telomeres, recombinant tankyrase 2 was subjected to an in vitro PARP assay. Tankyrase 2 poly(ADP-ribosyl)ated itself and TRF1. Overexpression of tankyrase 2 in the nucleus released endogenous TRF1 from telomeres. These findings establish tankyrase 2 as a bona fide PARP, with itself and TRF1 as acceptors of ADP-ribosylation, and suggest the possibility of a role for tankyrase 2 at telomeres
— id: 24853, year: 2002, vol: 22, page: 332, stat: Journal Article,

The telomeric poly(ADP-ribose) polymerase, tankyrase 1, contains multiple binding sites for telomeric repeat binding factor 1 (TRF1) and a novel acceptor, 182-kDa tankyrase-binding protein (TAB182)
Seimiya, Hiroyuki; Smith, Susan
2002 Apr 19;277(16):14116-14126, Journal of biological chemistry
Tankyrase 1, a human telomeric poly(ADP-ribose) polymerase, was originally identified through its interaction with TRF1, a negative regulator of telomere length. Tankyrase 1 ADP-ribosylates TRF1 in vitro, and its overexpression induces telomere elongation in human cancer cells. In addition to its telomeric localization, tankyrase 1 resides at multiple subcellular sites, suggesting additional functions for this protein. Here we identify TAB182, a novel tankyrase 1-binding protein of 182 kDa. TAB182 displays a complex pattern of subcellular localization. TAB182 localizes to the nucleus in a heterochromatic staining pattern and to the cytoplasm, where it co-stains with the cortical actin network. TAB182 coimmunoprecipitates with tankyrase 1 from human cells and serves as an acceptor of poly(ADP-ribosyl)ation by tankyrase 1 in vitro. Like TRF1, TAB182 binds to the ankyrin domain (comprising 24 ankyrin repeats) of tankyrase 1. Surprisingly, dissection of this domain reveals multiple discrete and overlapping binding sites for TRF1 and TAB182. Thus, we demonstrate five well conserved ankyrin repeat clusters in tankyrase 1. Although each of the five ankyrin repeat clusters independently binds to TRF1, only three of the five bind toTAB182. These findings suggest that tankyrase 1 may act as a scaffold for large molecular mass complexes made up of multiple binding proteins. We discuss potential roles for tankyrase 1-mediated higher order complexes at telomeres and at other subcellular sites
— id: 39708, year: 2002, vol: 277, page: 14116, stat: Journal Article,

The world according to PARP
Smith S
2001 Mar;26(3):174-179, Trends in biochemical sciences
An immediate cellular response to DNA damage is the synthesis of poly(ADP-ribose) by the enzyme poly(ADP-ribose) polymerase (PARP). This nuclear enzyme and the unique post-translational modification it catalyzes have long been considered to function exclusively in cellular surveillance of genotoxic stress. The recent identification of multiple members of a PARP family might force a revision of this concept. The novel primary structures and subcellular localizations for some of these PARPs suggests new and unexpected roles for poly(ADP-ribosyl)ation in telomere replication and cellular transport
— id: 20290, year: 2001, vol: 26, page: 174, stat: Journal Article,

Mammalian meiotic telomeres: protein composition and redistribution in relation to nuclear pores
Scherthan H; Jerratsch M; Li B; Smith S; Hulten M; Lock T; de Lange T
2000 Dec;11(12):4189-4203, Molecular biology of the cell
Mammalian telomeres consist of TTAGGG repeats, telomeric repeat binding factor (TRF), and other proteins, resulting in a protective structure at chromosome ends. Although structure and function of the somatic telomeric complex has been elucidated in some detail, the protein composition of mammalian meiotic telomeres is undetermined. Here we show, by indirect immunofluorescence (IF), that the meiotic telomere complex is similar to its somatic counterpart and contains significant amounts of TRF1, TRF2, and hRap1, while tankyrase, a poly-(ADP-ribose)polymerase at somatic telomeres and nuclear pores, forms small signals at ends of human meiotic chromosome cores. Analysis of rodent spermatocytes reveals Trf1 at mouse, TRF2 at rat, and mammalian Rap1 at meiotic telomeres of both rodents. Moreover, we demonstrate that telomere repositioning during meiotic prophase occurs in sectors of the nuclear envelope that are distinct from nuclear pore-dense areas. The latter form during preleptotene/leptotene and are present during entire prophase I
— id: 20289, year: 2000, vol: 11, page: 4189, stat: Journal Article,

Tankyrase promotes telomere elongation in human cells
Smith S; de Lange T
2000 Oct 19;10(20):1299-1302, Current biology. CB
Human telomeres are maintained by telomerase, a reverse transcriptase that adds telomeric repeats to chromosome ends [1,2]. In human tumors and immortalized cells, telomeres are often maintained at a constant length setting [3,4], indicating that telomerase-mediated telomere elongation is tightly regulated. Tankyrase, a telomeric poly(ADP-ribose) polymerase (PARP) [5], was identified through its interaction with TRF1 [6], a negative regulator of telomere extension by telomerase [7]. Tankyrase-mediated ADP-ribosylation inhibits binding of TRF1 to telomeric repeats in vitro [5], suggesting that tankyrase might regulate TRF1 and therefore control telomere dynamics in vivo. Here, we present evidence that tankyrase acts as a positive regulator of telomere elongation in vivo, apparently by inhibiting TRF1. Overexpression of tankyrase in the nucleus diminished the level of unmodified TRF1 in immunoblots and led to reduced immunofluorescence of TRF1 at interphase telomeres. Long-term overexpression of tankyrase in telomerase-positive human cells resulted in a gradual and progressive elongation of telomeres. A PARP-deficient form of tankyrase failed to affect TRF1 and did not alter telomere length dynamics, consistent with ADP-ribosylation of TRF1 as the main cause of altered telomere homeostasis. Our results indicate that tankyrase can induce telomere elongation in human cells. We propose that tankyrase-mediated ADP-ribosylation of TRF1 opens the telomeric complex, allowing access to telomerase
— id: 20288, year: 2000, vol: 10, page: 1299, stat: Journal Article,

Recombination: a means to an end in human cells
Smith, S
2000 Dec;26(4):388-389, Nature genetics
— id: 137819, year: 2000, vol: 26, page: 388, stat: Journal Article,

Cell cycle dependent localization of the telomeric PARP, tankyrase, to nuclear pore complexes and centrosomes
Smith S; de Lange T
1999 Nov;112(Pt 21):3649-3656, Journal of cell science
Tankyrase is a human poly(ADP-ribose) polymerase that was initially identified through its interaction with the telomeric protein TRF1, a negative regulator of telomere length. In vitro poly(ADP-ribosyl)ation by tankyrase inhibits TRF1 binding to telomeric DNA suggesting a role for tankyrase in telomere function. We previously demonstrated that tankyrase co-localizes with TRF1 at the ends of human chromosomes in metaphase. Here we show that tankyrase localizes to additional subcellular sites in a cell cycle dependent manner. In interphase, tankyrase co-localized with TRF1 to telomeres, but in addition was found to reside at nuclear pore complexes, as evidenced by indirect immunofluorescence, subcellular fractionation and immunoelectron microscopy. At mitosis, concomitant with nuclear envelope breakdown and nuclear pore complex disassembly, tankyrase was found to relocate around the pericentriolar matrix of mitotic centrosomes. This complex staining pattern along with the observation that tankyrase did not contain a nuclear localization signal suggested that its telomeric localization might be regulated, perhaps by TRF1. Indeed, localization of exogenously-expressed tankyrase to telomeres was dependent upon co-transfection with TRF1. These data indicate that the subcellular localization of tankyrase can be regulated by both the cell cycle and TRF1
— id: 8360, year: 1999, vol: 112, page: 3649, stat: Journal Article,

Chromosomal mapping of the tankyrase gene in human and mouse
Zhu L; Smith S; de Lange T; Seldin MF
1999 Apr 15;57(2):320-321, Genomics
— id: 8359, year: 1999, vol: 57, page: 320, stat: Journal Article,

Tankyrase, a poly(ADP-ribose) polymerase at human telomeres
Smith S; Giriat I; Schmitt A; de Lange T
1998 Nov 20;282(5393):1484-1487, Science
Tankyrase, a protein with homology to ankyrins and to the catalytic domain of poly(adenosine diphosphate-ribose) polymerase (PARP), was identified and localized to human telomeres. Tankyrase binds to the telomeric protein TRF1 (telomeric repeat binding factor-1), a negative regulator of telomere length maintenance. Like ankyrins, tankyrase contains 24 ankyrin repeats in a domain responsible for its interaction with TRF1. Recombinant tankyrase was found to have PARP activity in vitro, with both TRF1 and tankyrase functioning as acceptors for adenosine diphosphate (ADP)-ribosylation. ADP-ribosylation of TRF1 diminished its ability to bind to telomeric DNA in vitro, suggesting that telomere function in human cells is regulated by poly(ADP-ribosyl)ation
— id: 8358, year: 1998, vol: 282, page: 1484, stat: Journal Article,

TRF1 is a dimer and bends telomeric DNA
Bianchi A; Smith S; Chong L; Elias P; de Lange T
1997 Apr 1;16(7):1785-1794, EMBO journal
TRF1 is a mammalian telomeric protein that binds to the duplex array of TTAGGG repeats at chromosome ends. TRF1 has homology to the DNA-binding domain of the Myb family of transcription factors but, unlike most Myb-related proteins, TRF1 carries one rather than multiple Myb-type DNA-binding motifs. Here we show that TRF1 binds DNA as a dimer using a large conserved domain near the N-terminus of the protein for TRF1-TRF1 interactions. Dimerization was observed both in a complex with DNA and in the yeast two-hybrid assay. TRF1 dimers were found to require both Myb repeats for the formation of a stable complex with DNA, indicating a parallel between the DNA-binding mode of TRF1 and other Myb-related proteins. TRF1 was found to have a number of biochemical similarities to Rap1p, a distantly related DNA-binding protein that functions at telomeres in yeast. Rap1p and TRF1 both require two Myb motifs for DNA binding and both factors bind along their cognate telomeric sequences without showing strong cooperative interactions between adjacent proteins. Furthermore, TRF1 was found to bend its telomeric site to an angle of -120 degrees. Since Rap1p similarly distorts telomeric DNA, we propose that DNA bending is important for the function of telomeres in yeast and mammals
— id: 20287, year: 1997, vol: 16, page: 1785, stat: Journal Article,

TRF1, a mammalian telomeric protein
Smith S; de Lange T
1997 Jan;13(1):21-26, Trends in genetics
Telomerase adds TTAGGG repeats onto mammalian chromosome ends, replenishing the terminal sequence loss incurred during DNA replication. This maintenance of telomeric DNA preserves binding sites for telomeric proteins, which form a protective nucleoprotein complex at chromosome ends. The recent isolation of TRF1, the mammalian telomeric-repeat binding factor, should now allow the structure and function of the telomeric complex to be examined in detail
— id: 20286, year: 1997, vol: 13, page: 21, stat: Journal Article,