ATM regulates proteasome-dependent subnuclear localization of TRF1, which is important for telomere maintenance

Abstract

Ataxia telangiectasia mutated (ATM), a PI-3 kinase essential for maintaining genomic stability, has been shown to regulate TRF1, a negative mediator of telomerase-dependent telomere extension. However, little is known about ATM-mediated TRF1 phosphorylation site(s) in vivo. Here, we report that ATM phosphorylates S367 of TRF1 and that this phosphorylation renders TRF1 free of chromatin. We show that phosphorylated (pS367)TRF1 forms distinct non-telomeric subnuclear foci and that these foci occur predominantly in S and G2 phases, implying that their formation is cell cycle regulated. We show that phosphorylated (pS367)TRF1-containing foci are sensitive to proteasome inhibition. We find that a phosphomimic mutation of S367D abrogates TRF1 binding to telomeric DNA and renders TRF1 susceptible to protein degradation. In addition, we demonstrate that overexpressed TRF1-S367D accumulates in the subnuclear domains containing phosphorylated (pS367)TRF1 and that these subnuclear domains overlap with nuclear proteasome centers. Taken together, these results suggest that phosphorylated (pS367)TRF1-containing foci may represent nuclear sites for TRF1 proteolysis. Furthermore, we show that TRF1 carrying the S367D mutation is unable to inhibit telomerase-dependent telomere lengthening or to suppress the formation of telomere doublets and telomere loss in TRF1-depleted cells, suggesting that S367 phosphorylation by ATM is important for the regulation of telomere length and stability.

Cdk-dependent phosphorylation regulates TRF1 recruitment to PML bodies and promotes C-circle production in ALT cells

“…Immunofluorescence was performed as described (Mitchell and Zhu, 2014;Zhu et al, 2003). FISH analysis was carried out essentially as described (McKerlie et al, 2012). For triple staining, blocked coverslips were incubated with both rabbit anti-pT371 (at 1:500) and mouse anti-PML (at 1:200) antibodies in blocking buffer (1 mg/ml BSA, 3% goat serum, 0.1% Triton X-100 and 1 mM EDTA in PBS) at room temperature for 2 h. After three washes in PBS, coverslips were incubated with both TRITC-conjugated donkey anti-rabbit (1:250; Cat.…”

“…715-156-150, Jackson Laboratories) at room temperature for 30 min. Subsequently, coverslips were processed as described (McKerlie et al, 2012). Cell images were then recorded on a Zeiss Axioplan 2 microscope with a Hammamatsu C4742-95 camera and processed in Open Lab.…”

Telomeres and Telomerase: The Switch of Tumor Genesis in Vivo

“…In addition, TRF1 deficiency results in lower cellular multiplication, increasing G2 arrest and mitotic inactivity [45]. It is further demonstrated that TRF1 abraogation inhibits the growth of aggressive lung carcinomas independently of telomere length and p53 status [45].…”

“…It is characteristic that TRF1 deletion results in the accumulation of γ-H2AX at telomeric DNA, leading to the instability of chromosomes [43,44]. In addition, TRF1 deficiency results in lower cellular multiplication, increasing G2 arrest and mitotic inactivity [45]. It is further demonstrated that TRF1 abraogation inhibits the growth of aggressive lung carcinomas independently of telomere length and p53 status [45].…”

Posttranslational control of telomere maintenance and the telomere damage response

“…93,[98][99][100][101] While phosphorylation of TRF1 by Plk1 and CK2 increases TRF1-telomere interaction, ATMmediated phosphorylation appears to inhibit association of TRF1 with telomeres, thereby promoting telomere elongation, such that ATM inhibition leads to telomere shortening. The latter is consistent with the elevated rate of telomere shortening observed in cells from patients with ataxia telangiectasia (AT), a disease caused by ATM mutation.…”