DNA-Directed Polymerase Subunits Play a Vital Role in Human Telomeric Overhang Processing.

UNLABELLED: Telomeres consist of TTAGGG repeats bound by the shelterin complex and end with a 3' overhang. In humans, telomeres shorten at each cell division, unless telomerase (TERT) is expressed and able to add telomeric repeats. For effective telomere maintenance, the DNA strand complementary to that made by telomerase must be synthesized. Recent studies have discovered a link between different activities necessary to process telomeres in the S phase of the cell cycle to reform a proper overhang. Notably, the human CST complex (CTC1/STN1/TEN1), known to interact functionally with the polymerase complex (POLA/primase), was shown to be important for telomere processing. Here, focus was paid to the catalytic (POLA1/p180) and accessory (POLA2/p68) subunits of the polymerase, and their mechanistic roles at telomeres. We were able to detect p68 and p180 at telomeres in S-phase using chromatin immunoprecipitation. We could also show that the CST, shelterin, and polymerase complexes interact, revealing contacts occurring at telomeres. We found that the polymerase complex could associate with telomerase activity. Finally, depletion of p180 by siRNA led to increased overhang amounts at telomeres. These data support a model in which the polymerase complex is important for proper telomeric overhang processing through fill-in synthesis, during S phase. These results shed light on important events necessary for efficient telomere maintenance and protection. IMPLICATIONS: This study describes the interplay between DNA replication components with proteins that associate with chromosome ends, and telomerase. These interactions are proposed to be important for the processing and protection of chromosome ends. ©2014 American Association for Cancer Research.