UC Berkeley

Telomeres are specialized protein-DNA structures that protect the ends of linear chromosomes, and they are maintained by the telomerase ribonucleoprotein (RNP) enzyme complex. Recombinant telomerase RNP with catalytic activity contains, at a minimum, the catalytic reverse transcriptase subunit (TERT) and the telomerase RNA (TER). However, endogenous telomerase is a much larger holoenzyme complex, with telomerase-associated subunits that contribute to RNP assembly and regulation. Telomerase-associated subunits may also directly affect the biochemical features of telomerase catalytic activity. In vitro reconstitution of Tetrahymena thermophila minimal RNP results in telomerase activity with low repeat addition processivity (RAP), while the endogenously assembled complexes can have both low and high RAP.

Knowledge of a comprehensive list of telomerase-associated proteins expands our understanding of how telomerase-associated proteins regulate telomerase. Therefore, I optimized an affinity purification of TERT-associated proteins and identified telomerase-associated proteins to add to the list of known subunits. I have also begun characterizing the architecture of the telomerase holoenzyme: three subunits form a subcomplex that bridges the minimal RNP and a processivity factor, which contains telomeric single-stranded DNA-binding activity. With these telomerase subunits, a telomerase complex with high RAP can be reconstituted in vitro. The work described here expands the known functions of telomerase-associated proteins and the molecular mechanisms of telomere length regulation.