Proteomic Profiling Reveals a Specific Role for Translesion DNA Polymerase η in the Alternative Lengthening of Telomeres

Laura Garcia-Exposito,Elodie Bournique,Valérie Bergoglio,Arindam Bose,Jonathan Barroso-Gonzalez,Sufang Zhang,Justin L Roncaioli,Marietta Lee,Callen T Wallace,Simon C Watkins,Patricia L Opresko,Jean-Sébastien Hoffmann,Roderick J O’Sullivan

doi:10.1016/j.celrep.2016.10.048

Abstract

SUMMARYCancer cells rely on the activation of telomerase or the alternative lengthening of telomeres (ALT) pathways for telomere maintenance and survival. ALT involves homologous recombination (HR)-dependent exchange and/or HR-associated synthesis of telomeric DNA. Utilizing proximity-dependent biotinylation (BioID), we sought to determine the proteome of telomeres in cancer cells that employ these distinct telomere elongation mechanisms. Our analysis reveals that multiple DNA repair networks converge at ALT telomeres. These include the specialized translesion DNA synthesis (TLS) proteins FANCJ-RAD18-PCNA and, most notably, DNA polymerase eta (Polη). We observe that the depletion of Polη leads to increased ALT activity and late DNA polymerase δ (Polδ)-dependent synthesis of telomeric DNA in mitosis. We propose that Polη fulfills an important role in managing replicative stress at ALT telomeres, maintaining telomere recombination at tolerable levels and stimulating DNA synthesis by Polδ.

Highlights

Telomere maintenance mechanisms are normally restrained to prevent tumorigenesis but are hijacked by cancer cells
We show that Polh is initially recruited to telomeres via RAD18-dependent ubiquitination of PCNA and partly via an association with the shelterin subunit TRF1
We verified that the stable expression of myc-BirA-TRF1 does not adversely affect the binding of endogenous TRF1 at telomeres and that the induction of BirA-mediated biotinylation does not perturb cell-cycle progression or provoke DNA damage checkpoint signaling in U2OS cells (Figure S1B–S1D)

Summary

Introduction

Telomere maintenance mechanisms are normally restrained to prevent tumorigenesis but are hijacked by cancer cells. Telomerase is suppressed in a significant number of cancers that maintain telomere length by engaging the alternative lengthening of telomeres (ALT) mechanism (Bryan et al, 1997). ALT involves the inter-chromosomal exchange of telomeric DNA and is regulated by DNA repair and core recombination factors. DNA repair factors frequently co-localize within clusters of telomeric DNA and specialized structures termed ALT-associated PML bodies (APBs) (reviewed in Cesare and Reddel, 2010). Elegant time-lapse imaging of telomeres in living cells has revealed that telomeric recombination and, by implication, so-called ALT activity can be stimulated by telomeric DNA damage, which triggers a homology search over distances of several microns in the cell and the clustering of several telomeres within PML bodies together with DNA repair and homologous recombination (HR) factors (Cho et al, 2014)

Results

Discussion

Conclusion