AN IN-VITRO ASSAY FOR SACCHAROMYCES TELOMERASE REQUIRES EST1

Telomerase activity was demonstrated in cell-free extracts from S. cer evisiae through the use of a PCR-based assay. As expected, this activi ty was eliminated by RNase or phenol treatment of the extract and was dependent on dGTP and dTTP. Telomerase was not detected in extracts pr epared from cells grown far similar to 30 or more cell divisions in th e absence of the EST1 product, Est1p. TLC1 RNA, which determines the s equence of telomeric DNA in vivo, was present in normal amounts in est 1 Delta cells. Moreover, TLC1 RNA specifically precipitated with epito pe-tagged Est1p. These data indicate that Est1p is either a subunit of yeast telomerase or an accessory protein associated with telomerase t hat is essential in vitro for its activity.