Polymerization defects within human telomerase are distinct from telomerase RNA and TEP1 binding

The minimal, active core of human telomerase is postulated to contain two c omponents, the telomerase RNA hTER and the telomerase reverse transcriptase hTERT. The reconstitution of human telomerase activity in vitro has facili tated the identification of sequences within the telomerase RNA and the RT motifs of hTERT that are essential for telomerase activity. However, the pr ecise role of residues outside the RT domain of hTERT is unknown. Here we h ave delineated several regions within hTERT that are important for telomera se catalysis, primer use, and interaction with the telomerase RNA and the t elomerase-associated protein TEP1. In particular, certain deletions of the amino and carboxy terminus of hTERT that retained an interaction with telom erase RNA and TEP1 were nonetheless completely inactive in vitro and in viv o. Furthermore, hTERT truncations lacking the amino terminus that were comp etent to bind the telomerase RNA were severely compromised for the ability to elongate telomeric and nontelomeric primers. These results suggest that the interaction of telomerase RNA with hTERT can be functionally uncoupled from polymerization, and that there are regions outside the RT domain of hT ERT that are critical for telomerase activity and primer use. These results establish that the human telomerase RT possesses unique polymerization det erminants that distinguish it from other RTs.