Telomere repeat–binding factor 2 binds extensively to extra-telomeric G-quadruplexes and regulates the epigenetic status of several gene promoters

Ananda Kishore Mukherjee,Shalu Sharma,Sulochana Bagri,Rintu Kutum,Pankaj Kumar,Asgar Hussain,Prateek Singh,Dhurjhoti Saha,Anirban Kar,Debasis Dash,Shantanu Chowdhury

doi:10.1074/jbc.ra119.008687

Abstract

The role of the telomere repeat-binding factor 2 (TRF2) in telomere maintenance is well-established. However, recent findings suggest that TRF2 also functions outside telomeres, but relatively little is known about this function. Herein, using genome-wide ChIP-Seq assays of TRF2-bound chromatin from HT1080 fibrosarcoma cells, we identified thousands of TRF2-binding sites within the extra-telomeric genome. In light of this observation, we asked how TRF2 occupancy is organized within the genome. Interestingly, we found that extra-telomeric TRF2 sites throughout the genome are enriched in potential G-quadruplex–forming DNA sequences. Furthermore, we validated TRF2 occupancy at several promoter G-quadruplex motifs, which did adopt quadruplex forms in solution. TRF2 binding altered expression and the epigenetic state of several target promoters, indicated by histone modifications resulting in transcriptional repression of eight of nine genes investigated here. Furthermore, TRF2 occupancy and target gene expression were also sensitive to the well-known intracellular G-quadruplex–binding ligand 360A. Together, these results reveal an extensive genome-wide association of TRF2 outside telomeres and that it regulates gene expression in a G-quadruplex–dependent fashion.

Highlights

The role of the telomere repeat-binding factor 2 (TRF2) in telomere maintenance is well-established
To check the overall distribution of the aligned reads, the entire genome was divided into 50-bp bins; significant enrichment of TRF2 reads over input was found in 2.79% of the bins, suggesting selective distribution of TRF2 occupancy at a genome-wide level
To test interaction of TRF2 with G4 motifs, we focused on nine promoters that had at least one potential G4 (PG4) motif within 500 bp of transcription start sites (TSS) and overlapped with the TRF2HC peak (Fig. 3A)

Summary

Introduction

The role of the telomere repeat-binding factor 2 (TRF2) in telomere maintenance is well-established. TRF2 occupancy and target gene expression were sensitive to the well-known intracellular G-quadruplex– binding ligand 360A Together, these results reveal an extensive genome-wide association of TRF2 outside telomeres and that it regulates gene expression in a G-quadruplex– dependent fashion. TRF2 plays a critical role in how telomere ends evade detection as damaged DNA by blocking the ATM5 and ATR kinases from triggering DNA damage response [10] For this and other telomererelated functions (10 –12), TRF2 has been studied as a telomeric factor [13, 14], where it is known to associate with double-stranded telomeric DNA as a homodimer from dimerization of the C-terminal MYB domains of TRF2 [10, 11].

Methods

Results

Conclusion