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Published online 26 March 2004

Nucleic Acids Research, 2004, Vol. 32, No. 6 1917-1927
© 2004 Oxford University Press

Humanized telomeres and an attempt to express a functional human telomerase in yeast

Amadou Bah, François Bachand1, Éveline Clair, Chantal Autexier1 and Raymund J. Wellinger*

Department of Microbiology and Infectiology, ARN/RNA Group, Faculty of Medicine, Université de Sherbrooke, Sherbrooke, Canada and 1 Department of Anatomy and Cell Biology, Department of Medicine, McGill University, and Bloomfield Center for Research in Aging, Lady Davis Institute for Medical Research, The Sir Mortimer B. Davis-Jewish General Hospital, Montreal, Canada

*To whom correspondence should be addressed. Tel: +1 819 564 5214; Fax: +1 819 564 5392; Email: Raymund.Wellinger{at}Usherbrooke.ca
Present addresses:
F. Bachand, Department of Cancer Biology, Dana Farber Cancer Institute, Boston, MA, USA
É. Clair, Department of Biochemistry, McGill University, Montreal, Canada

Received December 23, 2003; Revised February 18, 2004; Accepted March 8, 2004

The maintenance of telomeric repeat DNA depends on an evolutionarily conserved reverse trans criptase called telomerase. In vitro, only the catalytic subunit and a telomerase-associated RNA are required for the synthesis of species-specific repeat DNA. In an attempt to establish a heterologous system for the study of the human telomerase enzyme, we expressed the two core components and predicted regulatory subunits in the yeast Saccharomyces cerevisiae. We show that adequate substrates for human telomerase can be generated; the expressed enzyme was localized in the nucleus and it had the capacity to synthesize human-specific repeats in vitro. However, there was no evidence for human telomerase activity at yeast telomeres in vivo. Therefore functional replacement of the yeast telomerase by the human enzyme may require additional human-specific components. We also replaced the template region of the yeast telomerase RNA with one that dictates the synthesis of vertebrate repeats and performed a detailed molecular analysis of the composition of the telomeres upon outgrowth of such strains. The results suggest that vertebrate repeats on yeast telomeres are subject to a very high degree of repeat turnover and show that an innermost tract of 50 bp of yeast repeats are resistant to replacement.


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