Finding the end: recruitment of telomerase to telomeres

Abstract

Telomeres, the ends of linear eukaryotic chromosomes, are characterized by the presence of multiple repeats of a short DNA sequence. This telomeric DNA is protected from illicit repair by telomere-associated proteins, which in mammals form the shelterin complex. Replicative polymerases are unable to synthesize DNA at the extreme ends of chromosomes, but in unicellular eukaryotes such as yeast and in mammalian germ cells and stem cells, telomere length is maintained by a ribonucleoprotein enzyme known as telomerase. Recent work has provided insights into the mechanisms of telomerase recruitment to telomeres, highlighting the contribution of telomere-associated proteins, including TPP1 in humans, Ccq1 in Schizosaccharomyces pombe and Cdc13 and Ku70-Ku80 in Saccharomyces cerevisiae.

Figure 1

Chromosome end-protection versus telomerase recruitment and action. Deprotected, protein-free telomeric DNA (G/C strand duplex at top) could be a substrate for illicit DNA end-joining and cell cycle checkpoint activation events at the telomere. By specifically binding telomeric DNA, telomeric proteins protect chromosome ends from such deleterious processes. DNA sequences are lost at the ends of chromosomes due to incomplete replication by DNA polymerases. Telomerase is recruited to telomeres via interaction with specific telomeric proteins to extend chromosome ends and thereby counter DNA lost from incomplete replication. In the absence of telomerase recruitment or action, telomeric DNA shortens, ultimately leading to cell senescence. Furthermore, shorter telomeric DNA results in a loss of bound telomeric proteins, which could result in deprotected chromosome ends.

Figure 2

Structures of telomeric proteins and telomerase components. a, Crystal structure of TRFH domain (PDB: 1H6O) and dsDNA-bound Myb domain of human TRF1 (PDB: 1W0T). b, Crystal structure of TRFH domain (1H6P) and dsDNA-bound Myb domain of human TRF2 (PDB: 1W0U). c, Crystal structure of ssDNA-bound DNA-binding domain (DBD) of human POT1, comprised of two OB folds (PDB: 1XJV). d, Crystal structure of the N-terminal OB domain of human TPP1 (PDB: 2I46). e, NMR structure of ssDNA-bound DBD of S. cerevisiae Cdc13 (PDB: 1S40). f, Crystal structure of ssDNA-bound structure of the first OB domain (OB1) of S. pomble Pot1 (PDB: 1QZG). g, NMR structures of template-pseudoknot (PK) fragments of human TR containing the indicated secondary structural elements; top structure (PDB: 2L3E); bottom structure (PDB: 2K96). The ‘P’ and ‘J’ elements are RNA double-helical paired regions and joining segments, respectively. h, Crystal structure of T. castaneum TERT with a hybrid RNADNA hairpin representing a putative telomerase-primer-template ternary complex (PDB: 3KYL).

Figure 3

Models for telomerase recruitment in humans and fission yeast. a, Model for telomerase recruitment in humans. The first step of human telomerase recruitment involves the redistribution of telomerase from Cajal bodies (site for accumulation of recruitment-competent telomerase RNP) to telomeres driven by the telomerase-TPP1 interaction^{, ,} . This recruitment is TIN2-dependent because TIN2 is required for recruiting TPP1 to telomeres. It is possible that the TPP1-telomerase interaction is mediated by a yet-to-be-identified Ccq1 homologue in humans. Once at telomeres, telomerase action during S-phase is coordinated with sister telomere cohesion, which is facilitated by the recruitment of HP1, mediated by the PTVML binding-motif of TIN2^, . Telomerase extension then proceeds in a processive manner facilitated by bound POT1-TPP1, which reduces the primer dissociation rate and increases the translocation efficiency of telomerase^{, ,} . Putative interactions are indicated by dashed arrows. b, Model for telomerase recruitment in fission yeast. The Pot1-Tpz1-Ccq1 complex (telomerase stimulatory) associates with the Taz1-Rap1-Poz1 complex (telomerase inhibitory) at long telomeres. Such an association is facilitated by the presumably high local concentration of Taz1-Rap1 proteins at such long telomeres and results in the inhibition of Ccq1 phosphorylation at T93 by Tel1 and Rad3^{, ,} . At short telomeres however, the Pot1-Tpz1-Ccq1 connection to Taz1-Rap1 is lost owing to lower protein concentration of Taz1-Rap1, facilitating phosphorylation of Ccq1, which results in Est1-Ccq1 binding via the interaction between the 14-3-3-like domain of Est1 and the T93-phosphorylated face of Ccq1^{, ,} . Although telomerase is recruited to telomeres via interaction of Trt1 to Ccq1-bound Est1, this state would be in a ‘closed’ configuration with regards to telomerase action, possibly because telomerase tethered to shelterin is somehow unable to extend telomeres. A conformational switch that replaces phosphorylated Ccq1 with TER1 at the phosphoSer/RNA binding surface of the 14-3-3-like domain of Est1 would relieve the telomerase-shelterin tether and lead to an ‘open’ configuration of telomerase that allows it to carry out telomere elongation.

Figure 4

Model for telomerase recruitment in budding yeast. The Ku heterodimer binds TLC1 (the telomerase RNA) and promotes nuclear import and retention of the RNP. The interaction between ssDNA-bound Cdc13 and Est1 is responsible for recruiting telomerase to budding yeast telomeres. It is not known whether Cdc13 is part of the CST complex when it recruits telomerase to telomeres. The Est1-ssDNA interaction may further strengthen the telomerase-telomere tether. A putative Est2-dependent Est3-ssDNA interaction proposed for certain Candida species could serve as yet another mechanism for telomerase recruitment.