Research Article: The Telomeric Protein TRF2 Binds the ATM Kinase and Can Inhibit the ATM-Dependent DNA Damage Response

Date Published: August 17, 2004

Publisher: Public Library of Science

Author(s): Jan Karlseder, Kristina Hoke, Olga K Mirzoeva, Christopher Bakkenist, Michael B Kastan, John H. J Petrini, Titia de Lange

Abstract: The telomeric protein TRF2 is required to prevent mammalian telomeres from activating DNA damage checkpoints. Here we show that overexpression of TRF2 affects the response of the ATM kinase to DNA damage. Overexpression of TRF2 abrogated the cell cycle arrest after ionizing radiation and diminished several other readouts of the DNA damage response, including phosphorylation of Nbs1, induction of p53, and upregulation of p53 targets. TRF2 inhibited autophosphorylation of ATM on S1981, an early step in the activation of this kinase. A region of ATM containing S1981 was found to directly interact with TRF2 in vitro, and ATM immunoprecipitates contained TRF2. We propose that TRF2 has the ability to inhibit ATM activation at telomeres. Because TRF2 is abundant at chromosome ends but not elsewhere in the nucleus, this mechanism of checkpoint control could specifically block a DNA damage response at telomeres without affecting the surveillance of chromosome internal damage.

Partial Text: Telomeres prevent the recognition of natural chromosome ends as double-stranded breaks (DSBs). When telomeres become dysfunctional due to shortening or loss of protective factors, chromosome ends activate a DNA damage response mediated (in part) by the ATM kinase (Karlseder et al. 1999; Takai et al. 2003). A major challenge in telomere biology is to define the mechanism by which functional telomeres prevent these events. Here we show that the human telomere-associated protein TRF2 is an inhibitor of the ATM kinase, suggesting a mechanism by which the telomeric protein complex prevents the activation of this DNA damage response transducer.

TRF2 was overexpressed in IMR90 primary fibroblasts using a retroviral vector. Under these conditions, TRF2 saturates its telomeric binding sites and is present in the nucleoplasm. While control IMR90 cells showed the expected reduction in mitotic index after ionizing radiation (IR), TRF2 overexpression partially abrogated this checkpoint response, increasing the percentage of cells entering mitosis from 0.3% to 8% (Figure 1). The inappropriate entry into mitosis is indicative of a failure of the cell cycle checkpoints. Since cell cycle arrest after IR largely depends on ATM (reviewed in Shiloh 2003), we asked whether TRF2 was acting on this kinase. Caffeine, an inhibitor of ATM and the related kinase ATR, suppressed IR-induced arrest to a similar extent as TRF2 (Figure 1). Furthermore, caffeine had no additional effect on the ability of TRF2-overexpressing cells to bypass the DSB checkpoint, suggesting that TRF2 and caffeine target the same step in the response pathway.

Natural chromosome ends require mechanisms to prevent the activation of the DNA damage response. Inhibition of the ATM kinase at human telomeres is particularly important since the telomeric complex contains the Mre11 complex, one of the DNA damage sensors of the ATM pathway (Carson et al. 2003; Petrini and Stracker 2003; Uziel et al. 2003). The telomeric protein TRF2 appears to play a central role in preventing telomeres from activating ATM. Removal of TRF2 from telomeres results in the localization of the active, phosphorylated form of ATM to unprotected chromosome ends (Takai et al. 2003) and induces ATM-dependent apoptosis (Karlseder et al. 1999). The data reported here are consistent with the hypothesis that TRF2 protects telomeres through a direct interaction with ATM that blocks its activation. As a result, TRF2 abrogates the downstream outcomes of the ATM-dependent DNA damage response, including phosphorylation of various ATM targets and cell cycle arrest.



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