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Molecular and Cellular Biology, March 2007, p. 2074-2083, Vol. 27, No. 6
0270-7306/07/$08.00+0     doi:10.1128/MCB.02105-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Positive and Negative Regulation of Tetrahymena Telomerase Holoenzyme{triangledown} ,{dagger}

Keren L. Witkin,{ddagger} Ramadevi Prathapam, and Kathleen Collins*

Department of Molecular and Cell Biology, University of California at Berkeley, Berkeley, California 94720-3204

Received 10 November 2006/ Returned for modification 6 December 2006/ Accepted 31 December 2006

Telomerase replenishes the telomeric repeats that cap eukaryotic chromosome ends. To perform DNA synthesis, the active site of telomerase reverse transcriptase (TERT) copies a template within the integral telomerase RNA (TER). In vivo, TERT and TER and additional subunits form a telomerase holoenzyme capable of telomere elongation. We previously purified epitope-tagged Tetrahymena thermophila TERT and characterized two of the associated proteins. Here we characterize the remaining two proteins that were enriched by TERT purification. The primary sequence of the p75 polypeptide lacks evident homology with other proteins, whereas the p20 polypeptide is the Tetrahymena ortholog of a conserved multifunctional protein, Skp1. Genetic depletion of p75 induced telomere shortening without affecting the accumulation of TER or TERT, suggesting that p75 promotes telomerase function at the telomere. Affinity purification of p75 coenriched telomerase activity and each other known telomerase holoenzyme protein. On the other hand, genetic depletion of Skp1p induced telomere elongation, suggesting that this protein plays a negative regulatory role in the maintenance of telomere length homeostasis. Affinity purification of Skp1p did not detectably enrich active telomerase but did copurify ubiquitin ligase machinery. These studies reveal additional complexity in the positive and negative regulation of Tetrahymena telomerase function.

* Corresponding author. Mailing address: Department of Molecular and Cell Biology, 16 Barker Hall #3204, University of California at Berkeley, Berkeley, CA 94720-3204. Phone: (510) 643-1598. Fax: (510) 643-6334. E-mail: kcollins{at}berkeley.edu.

{triangledown} Published ahead of print on 12 January 2007.

{dagger} Supplemental material for this article may be found at http://mcb.asm.org/.

{ddagger} Current address: Laboratory of Molecular and Cellular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892.

Molecular and Cellular Biology, March 2007, p. 2074-2083, Vol. 27, No. 6
0270-7306/07/$08.00+0     doi:10.1128/MCB.02105-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Couvillion, M. T., Lee, S. R., Hogstad, B., Malone, C. D., Tonkin, L. A., Sachidanandam, R., Hannon, G. J., Collins, K. (2009). Sequence, biogenesis, and function of diverse small RNA classes bound to the Piwi family proteins of Tetrahymena thermophila. Genes Dev. 23: 2016-2032 [Abstract] [Full Text]  
  • Lee, S. R., Talsky, K. B., Collins, K. (2009). A single RNA-dependent RNA polymerase assembles with mutually exclusive nucleotidyl transferase subunits to direct different pathways of small RNA biogenesis. RNA 15: 1363-1374 [Abstract] [Full Text]  


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