Targeting Assay To Study the cis Functions of Human Telomeric Proteins: Evidence for Inhibition of Telomerase by TRF1 and for Activation of Telomere Degradation by TRF2

Abstract

We investigated the control of telomere length by the human telomeric proteins TRF1 and TRF2. To this end, we established telomerase-positive cell lines in which the targeting of these telomeric proteins to specific telomeres could be induced. We demonstrate that their targeting leads to telomere shortening. This indicates that these proteins act in cis to repress telomere elongation. Inhibition of telomerase activity by a modified oligonucleotide did not further increase the pace of telomere erosion caused by TRF1 targeting, suggesting that telomerase itself is the target of TRF1 regulation. In contrast, TRF2 targeting and telomerase inhibition have additive effects. The possibility that TRF2 can activate a telomeric degradation pathway was directly tested in human primary cells that do not express telomerase. In these cells, overexpression of full-length TRF2 leads to an increased rate of telomere shortening.

We thank A. Belmont, T. de Lange, A. Pollice, J. Pulitzer, J. Shay, and M. P. Zibella for plasmids. We are grateful to P. A. Defossez and S. Marcand for helpful comments on the manuscript.

The work in the Gilson laboratory is supported by La Ligue Nationale contre le Cancer and the Région Rhône-Alps (contract 00816045). The work in the Sabatier laboratory is supported by CEC funding (FIGH-CT-1999-0009). K.A. is a Ph.D. fellow from La Ligue National contre le Cancer, M.B. is supported by the CEC (HPRN-CT-2000-00089), and S.B. is supported by a postdoctoral fellowship from the Centre Léon Bérard (Lyon, France).