Eukaryotic Cell doi:10.1128/EC.00069-07
Copyright (c) 2007, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.
THE MUTUAL DEPENDENCE OF CANDIDA EST1P AND EST3P IN TELOMERASE ASSEMBLY AND ACTIVATION
Min Hsu,
Eun Young Yu,
Sunitha M. Singh,
and
Neal F. Lue*
Department of Microbiology & Immunology, W. R. Hearst Microbiology Research Center, Weill Medical College of Cornell University, 1300 York Avenue, New York, NY 10021
* To whom correspondence should be addressed. Email:
nflue{at}med.cornell.edu.
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Abstract |
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Telomerase is an RNA-protein complex responsible for extending one strand of the telomere terminal repeats. Analysis of the telomerase complex in budding yeasts has revealed the presence of one catalytic protein subunit (Est2p/TERT) and at least two non-catalytic components (Est1p and Est3p). The TERT subunit is essential for telomerase catalysis while the function of Est1p and Est3p have not been precisely elucidated. In an earlier study, we showed that telomerase derived from a Candida est1 null mutant is defective in primer utilization in vitro; it exhibits reduced initiation and processivity on primers that terminates in two regions of the telomere repeat. Here we show that telomerase derived from a Candida est3 null mutant has nearly identical defects in primer utilization and processivity. Further analysis revealed an unexpected mutual dependence of Est1p and Est3p in their assembly into the full telomerase complex, which accounts for the similarity between the mutant enzymes. We also developed an affinity isolation and an in vitro re-constitution protocol for the telomerase complex that will facilitate future mechanistic studies.
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