Fig. 6.

Proposed mechanism for de novo telomere addition illustrated by addition of ADD1 onto a TGG breakpoint by two cycles of annealing and synthesis. In vitro studies indicate telomerase requires a 3′ single-stranded substrate (16) that could be revealed by resection of the broken chromosome V (44). Initial annealing of TGG at the preferred location allows up to five bases in the first annealing-synthesis-dissociation cycle; synthesis of more than five bases would not generate an ADD1 addition sequence. Dissociation after either ¹⁰G¹⁰ or ¹⁰GT¹¹ are added will generate new ends that will preferentially reanneal to the initial annealing registration (Fig. 5) and therefore give the appearance of high processivity through this region when analyzing only bulk telomeric sequences. On the other hand, dissociation of longer fragments generated by addition of ¹⁰GTG¹², ¹⁰GTGT¹³, ¹⁰GTGTG¹⁴, or ¹⁰GTGTGG¹⁵ will generate new ends that will preferentially allow reannealing to a second, common registration on the TLC1 template (only the specific case of ¹⁰GTGT¹³ addition in the first cycle is illustrated). Synthesis in the second registration is sufficient to add final nucleotides of the 11-nt ADD1 sequence. The robustness of reannealing of potential intermediate sequences to the first and second registrations on the TLC1 template explains the high frequency of ADD1 additions (Fig. 4a).