Caenorhabditis elegans POT-2 telomere protein represses a mode of alternative lengthening of telomeres with normal telomere lengths

Abstract

Canonical telomere repeats at chromosome termini can be maintained by a telomerase-independent pathway termed alternative lengthening of telomeres (ALT). Human cancers that survive via ALT can exhibit long and heterogeneous telomeres, although many telomerase-negative tumors possess telomeres of normal length. Here, we report that Caenorhabditis elegans telomerase mutants that survived via ALT possessed either long or normal telomere lengths. Most ALT strains displayed end-to-end chromosome fusions, suggesting that critical telomere shortening occurred before or concomitant with ALT. ALT required the 9-1-1 DNA damage response complex and its clamp loader, HPR-17. Deficiency for the POT-2 telomere binding protein promoted ALT in telomerase mutants, overcame the requirement for the 9-1-1 complex in ALT, and promoted ALT with normal telomere lengths. We propose that telomerase-deficient human tumors with normal telomere lengths could represent a mode of ALT that is facilitated by telomere capping protein dysfunction. PROTECTION OF TELOMERES-2 | telomere maintenanceT elomeres are tandem repeat tracts that cap the ends of linear chromosomes and erode with each cell cycle because of the inability of canonical DNA polymerases to completely replicate chromosome termini (1). However, the ribonucleoprotein telomerase combats this erosion by adding de novo telomeric repeats via reverse transcription (2). Progressive telomere shortening during proliferation of human somatic cells can trigger senescence, which serves as a major tumor suppression mechanism (3). Bypass of senescence and further cell proliferation leads to critical telomere shortening and crisis, resulting in high levels of cell death due to chromosome instability (4). Many tumors overcome this proliferation barrier by activating expression of the telomerase reverse transcriptase. The remaining 10-15% of human tumors do not possess telomerase activity (5, 6).S. cerevisiae strains deficient for telomerase can survive with long and heterogeneous telomeres composed of amplified telomere repeats (type II survivors) (7,8). Analogous to type II survivors, long and heterogeneous telomeres occur in almost all in vitro immortalized cell lines and tumor-derived cells lines that are telomerase-negative (9-11). Furthermore, extensive analysis of telomerase activity and telomere length from primary human tumor samples has revealed that certain telomerase-negative tumors possess long and heterogeneous telomeres, defined as alternative lengthening of telomeres (ALT) tumors (9). However, the majority (81/108) of primary tumors lacking telomerase activity have telomeres of normal lengths (9,(12)(13)(14)(15). This subset of telomerase-negative tumors may have become neoplastic before exhausting their telomere repeat reserves (9, 16). Alternatively, these tumors could have evolved via the same telomeredriven crisis events that give rise to ALT tumors, followed by activation of a distinct form of ALT that maintains telomeres of apparently normal lengths (9).Homologous recombinatio...

Caenorhabditis elegans POT-1 and POT-2 Repress Telomere Maintenance Pathways

“…Thus, although POT-1 mildly represses telomere shortening in the absence of telomerase, deficiency for pot-1 or both pot-1 and pot-2 failed to enhance the onset of senescence in the absence of telomerase. This increased trans -generational lifespan is consistent with recent reports that either POT-1 or POT-2 can suppress the telomerase-independent telomere maintenance mechanism termed ALT (Cheng et al 2012; Lackner et al 2012). …”

“…We observed wild-type levels of telomeric C-circles in early-generation pot-2 strains with short telomeres, and large 5- to 7-fold increases in C-circles in late-generation pot-2 strains with long telomeres (Figure 4). Previously established trt-1 ; pot-2 ALT strains with telomeres of normal lengths (Cheng et al 2012) displayed little or no increases in C-circle formation, indicating that the high levels of C-circles in late-generation pot-2 strains could require the presence of extremely long telomeres and possibly the activity of the telomerase reverse transcriptase. Consistent with the notion that telomerase could contribute to C-circle formation, trt-1 ; pot-1 ALT strains with either short or very long telomeres displayed modestly elevated levels of C-circles (Figure 4C).…”

“…Recent independent reports have indicated that deficiency for pot-1 or pot-2 can promote the telomerase-independent telomere maintenance pathway ALT in trt-1 mutants (Cheng et al 2012; Lackner et al 2012). POT-1 has been previously shown to repress C-circle formation (Raices et al 2008), a bona fide marker of ALT, and here we show the same function for POT-2 (Figure 4), suggesting that DNA replication intermediates relevant to ALT may occur in animals lacking either of these proteins.…”

“…POT-1 has been previously shown to repress C-circle formation (Raices et al 2008), a bona fide marker of ALT, and here we show the same function for POT-2 (Figure 4), suggesting that DNA replication intermediates relevant to ALT may occur in animals lacking either of these proteins. We previously observed an ALT phenotype that allows telomerase mutants to escape senescence indefinitely, but only when hundreds of animals were transferred weekly (Cheng et al 2012). In the present study, we transferred only 6 animals once a week, which we expected might preclude the onset of a full-blown ALT phenotype.…”

Chromosome Evolution in Dendropsophini (Amphibia, Anura, Hylinae)

“…According to Jones and Houben [2003 and references therein], spontaneous amplification of coding and noncoding tandem repeat sequences derived from A chromosomes seems to be strongly associated with the origin and evolution of B chromosomes. A telomeraseindependent lengthening of the telomeres [Cheng et al, 2012;Lovejoy et al, 2012] may explain this particular phenomenon in S. lacteus , although the repetitive nature of the telomeric tandem array, which is a target of molecular alterations in the number of repetitions, is an appropriate explanation [Jones and Houben, 2003]. However, a more exhaustive analysis is necessary to prove its consistency as a good cytological landmark.…”