Important role of Ser 219 phosphorylation of TRF1 in regulation of cell cycle

Abstract

To investigate the role of Ser 219 phosphorylation of TRF1 (telomere repeat binding factor 1) in regulation of cell cycle. The mimicking phosphorylation mutant (TRF1S219D-GFP) and the non-phosphorylatable mutant (TRF1S219A-GFP) were constructed; the mutant genes and corresponding proteins were checked by sequencing and Western blot, respectively. Immunofluorescence staining was performed to detect the localization of mutants in HeLa cells. Cell cycle was analyzed by flow cytometry and ATM level was evaluated by immunoblotting. The mutant genes were verified by direct sequencing and protein expression of GFP-tagged mutants was confirmed by immunoblotting.TRF1S219A-GFP and TRF1S219D-GFP were both localized in telomere of HeLa cells. Moreover, overexpression of TRF1-GFP or TRF1S219A-GFP resulted in an accumulation of HeLa cells in G2/M (P<0.05). The protein level of ATM was increased when overexpression the wide type or mutants. The Ser 219 phosphorylation of TRF1 by ATM could result in cell cycle arrest in G2/M, which is related to overexpression of TRF1.