Feedback control of telomere length D Shore and A Bianchi In ﬁssion yeast, the telomere duplex DNA-binding protein is called Taz1, a single-Myb domain containing protein similar to mammalian TRF1 and TRF2. Taz1 recruits both Rap1 and Rif1 orthologues to ﬁssion yeast telomeres, in which these proteins negatively regulate telomerase activity (reviewed in Rog and Cooper, 2008). In this system, Rif1 interacts with the telomere independently of Rap1, which instead recruits a novel telomeric protein called Poz1 (Miyoshi et al, 2008). Poz1 in turn interacts with Tpz1, the ﬁssion yeast orthologue of mammalian TPP1 (see below), which binds to both Pot1 (protection of telomeres 1; Baumann and Cech, 2001), a single-stranded DNA-binding OB-fold protein, and Ccq1, a protein involved in telomerase recruitment (Miyoshi et al, 2008; Tomita and Cooper, 2008). One current model for telomere length regulation in ﬁssion yeast (Miyoshi et al, 2008) posits a switch between telomer- ase inactive and active forms actuated by a transfer of the Poz1–Tpz1–Pot1–Ccq1 complex from Rap1 to the 30 single- stranded overhang. Precisely how this switch would be regulated, presumably as a function of TG repeat tract length, is still unclear. In mammals, telomeric duplex DNA is bound by two Myb domain protein homodimers, TRF1 and TRF2. In these sys- tems, the Rap1 orthologue is recruited by a direct interaction with TRF2, whereas a novel protein, TIN2, interacts with both TRF1 and TRF2 and provides the key bridge to the TPP1–POT1 telomere end-binding complex. Together, this complex of six proteins is referred to as the ‘shelterin’ complex, to denote its key role in both telomere protection and telomerase-based telomere maintenance (de Lange, 2005). It is unclear whether a telomere-length dependent switch in telomerase regulation similar to the one postulated above for ﬁssion yeast also operates in mammals, but this is of course a possibility given the similarities between the telomeric complexes in the two systems. Indeed, a mechan- ism to relay information regarding repeat-array length to the overhang that is the substrate for telomerase has been proposed in human cells, based on the dual ability of POT1 to indirectly interact with double-stranded repeat binding factors and also to directly bind the single-stranded overhang (Loayza and De Lange, 2003; Wang et al, 2007; Xin et al, 2007; see Figure 4). Consistent with these ideas, TPP1 inter- acts with telomerase and has been proposed to recruit the enzyme to chromosome ends (Xin et al, 2007). Again, the details of how this process might be regulated are still unknown, and the involvement of additional proteins is likely, perhaps including human Est1. Interestingly, the human MRX complex (MRN, where NBS1 is the human orthologue of Xrs2) and Ku86 both have roles in the telomerase pathway (O’Connor et al, 2004), just as in the yeast systems, but their mechanism of action might differ considerably in the mammalian system. MRN, which binds telomeres in late S/G2 phase of the cell cycle (Verdun et al, 2005; Verdun and Karlseder, 2006), recruits ATM (the mammalian Tel1 orthologue) to telomeres. One of ATM functions is phosphorylation of TRF1, which leads to its dissociation from telomeres (Wu et al, 2007), a curious twist on the situation in budding yeast in which the Rap1–Rif complex instead acts to inhibit telomere binding of Tel1 (ATM) (Hirano et al, 2009). An attractive notion, still untested, is that MRN–ATM association with telomeres is modulated by telomere repeat tract length, with ATM then acting on (several?) telomeric proteins to positively inﬂuence telomerase action. Regarding the apparent special role of TPP1 in ﬁssion yeast and mammalian telomere replication, we note that both S. cerevisiae and C. albicans Est3 proteins have recently been proposed to be structural homologues of TPP1 (Lee et al, 2008; Yu et al, 2008). Despite this structural similarity and a possible shared role in telomerase stimulation, the yeast Est3 proteins might have evolved at least partly distinct telomeric functions (e.g. with regard to telomere capping), or at least use different interacting partners. These ﬁndings point to how evolution has used a basic toolbox of telomeric proteins to derive different functional modules, and emphasize the importance of studying telomere biology in different organisms. Telomeric single-stranded DNA can form alternative DNA structures that may have important functions in both telo- mere capping and length regulation. In many different organ- isms, telomeres can adopt a so-called ‘t-loop’ conformation (Grifﬁth et al, 1999), in which the 30 single-stranded overhang is base paired to internal repeats in the T2AG3 array, pre- sumably hiding the chromosome end from the DNA damage checkpoint machinery, but perhaps also blocking telomerase access. TRF2, by promoting positive supercoiling, can induce t-loop formation in vitro (Amiard et al, 2007). Dramatic telomere shortening events can occur through homologous recombination at t-loop junctions (Wang et al, 2004), but it still remains to be seen whether t-loops have a function in homeostatic regulation of the telomerase pathway of length regulation. In addition to the t-loop, G-rich telomere over- hangs can form so-called G-quadruplex structures, which have been extensively studied in vitro. The best evidence Telomerase TERT TERC RAP1 TRF2 TIN2 TRF1 POT1 TPP1 TERT TERC Figure 4 Possible mechanisms of telomerase regulation by the shelterin complex in mammalian cells. The formation of a t-loop by the shelterin complex (not indicated here) is thought to sequester the telomere 30 terminus from telomerase. Opening of the t-loop will generate a structure (left) that may still provide only limited access to telomerase, perhaps due to the TPP1–TIN2 interaction. In this case, a transition to a more open structure (right) might be required to allow efﬁcient recruitment of telomerase by POT1–TPP1. See main text for more details and a description of related events thought to occur at telomeres in the ﬁssion yeast S. pombe. 2318 The EMBO Journal VOL 28 | NO 16 | 2009 & 2009 European Molecular Biology Organization