Murine Pif1 Interacts with Telomerase and Is Dispensable for Telomere Function In Vivo

Abstract

Pif1 is a 5′-to-3′ DNA helicase critical to DNA replication and telomere length maintenance in the budding yeast Saccharomyces cerevisiae. ScPif1 is a negative regulator of telomeric repeat synthesis by telomerase, and recombinant ScPif1 promotes the dissociation of the telomerase RNA template from telomeric DNA in vitro. In order to dissect the role of mPif1 in mammals, we cloned and disrupted the mPif1 gene. In wild-type animals, mPif1 expression was detected only in embryonic and hematopoietic lineages. mPif1^−/− mice were viable at expected frequencies, displayed no visible abnormalities, and showed no reproducible alteration in telomere length in two different null backgrounds, even after several generations. Spectral karyotyping of mPif1^−/− fibroblasts and splenocytes revealed no significant change in chromosomal rearrangements. Furthermore, induction of apoptosis or DNA damage revealed no differences in cell viability compared to what was found for wild-type fibroblasts and splenocytes. Despite a novel association of mPif1 with telomerase, mPif1 did not affect the elongation activity of telomerase in vitro. Thus, in contrast to what occurs with ScPif1, murine telomere homeostasis or genetic stability does not depend on mPif1, perhaps due to fundamental differences in the regulation of telomerase and/or telomere length between mice and yeast or due to genetic redundancy with other DNA helicases.

SUPPLEMENTAL MATERIAL

We thank Maria Mateyak, Virginia Zakian, John Murnane, and Maria Blasco for sharing unpublished results. We thank David Sealey for the chromatin immunoprecipitation analysis of mPif1 (unpublished) and for comments on the manuscript. We thank Mike Tyers, Daniel Durocher, and Carol Greider for advice and comments.

We thank Tak W. Mak for funding Andrew Wakeham's contribution. This study was funded by a grant from the NIH/NIA Longevity Assurance Genes Program (AG024398) and National Cancer Institute of Canada Research grant no. 15072 (using funds from the Canadian Cancer Society) to L.H.