Pif1- and Exo1-dependent nucleases coordinate resection at uncapped telomeres

• Main Author: Dewar, James Michael
• Published: University of Newcastle Upon Tyne 2011
• Subjects: 572.8
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.544202

Telomeres are present at the ends of most eukaryotic chromosomes and are bound by specialized telomere ‘capping’ proteins, preventing them from initiating a DNA Damage Response and cell cycle arrest analogous to that triggered by DNA Double Strand Breaks (DSBs). Resection of ‘uncapped’ telomeres and DSBs to generate extensive single-stranded DNA (ssDNA) is one of the most upstream events in this DDR, but while the nuclease activities that resect DSBs are well-defined, only a single nuclease, Exo1, is known to function at uncapped telomeres. This work establishes that the helicase Pif1, is required for a nuclease activity that resects uncapped telomeres in a parallel pathway to one defined by Exo1, Rad27 (Flap Endonuclease 1) and Rad24 (the 9-1-1 clamp loader). Following inactivation of the essential telomere capping protein Cdc13, Pif1 is shown to resect telomeres independently of Exo1 close to the chromosome end, but to play a less-crucial role in extensive resection. Furthermore, elimination of both Pif1 and Exo1 prevents the accumulation of ssDNA, thus eliminating the DDR at uncapped telomeres. Although Pif1 has no role in the resection of DSBs and has primarily been studied as a negative regulator of telomerase, it is shown to contribute to the DDR at uncapped telomeres in cells lacking telomerase and also to be crucial for telomere maintenance in telomerasedeficient cells. Cdc13 inhibits the DDR at uncapped telomeres and is believed to be required for telomerase recruitment. Astonishingly, elimination of Pif1 and Exo1 permits the viability of cells lacking Cdc13 and cdc13Δ exo1Δ pif1Δ mutants maintain and lengthen their telomeres over time, in a manner dependent on Ku, Rad52 and telomerase. Thus, Cdc13 is not a requirement for telomerase recruitment and elimination of the nuclease activities that function at uncapped telomeres can by pass the requirement for otherwise-essential telomere capping proteins.