Human Telomeres Maintain Their Overhang Length at Senescence

Abstract

Normal human cells in culture enter replicative senescence after a finite number of population doublings. The exact molecular mechanisms triggering the growth arrest are poorly understood. A recent report on the disappearance of the G-rich 3′ telomeric overhang in senescent cells led to the hypothesis that loss of the 3′ G-rich overhang is the molecular signal that triggers senescence. Here, we describe a quantitative assay to measure the length of the G-rich 3′ telomeric overhangs from cultured cells. Using both this assay and the conventional nondenaturing hybridization assay for measuring G-rich overhangs, we show that normal human fibroblasts can maintain their overhangs at senescence. Furthermore, cells do not lose their overhangs when they bypass senescence after the inactivation of p53 and Rb. We thus conclude that a global reduction in overhang length is not the molecular signal that triggers replicative senescence.

SUPPLEMENTAL MATERIAL

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

ACKNOWLEDGMENTS

We are grateful to B. Chabot (Université de Sherbrooke, Québec, Canada) for the GST-UP1 construct. We also thank anonymous reviewers for their helpful comments on the manuscript. We thank O. Bechter and N. Forsyth for discussion and critical comments on the manuscript and W. Walker and J. Sherrell for technical assistance.

This work was supported by a Ruth L. Kirschstein National Research Service Award Individual Fellowship (W.C.) and NIH AG01228. W.E.W. is an Ellison Foundation Senior Scholar.