Epigenetic telomere protection by Drosophila DNA damage response pathways

Analysis of terminal deletion chromosomes indicates that a sequence-independent mechanism regulates protection of Drosophila telomeres. Mutations in Drosophila DNA damage response genes such as atm/tefu, mre11, or rad50 disrupt telomere protection and localization of the telomere-associated proteins HP1 and HOAP, suggesting that recognition of chromosome ends contributes to telomere protection. However, the partial telomere protection phenotype of these mutations limits the ability to test if they act in the epigenetic telomere protection mechanism. We examined the roles of the Drosophila atm and atr-atrip DNA damage response pathways and the nbs homolog in DNA damage responses and telomere protection. As in other organisms, the atm and atr-atrip pathways act in parallel to promote telomere protection. Cells lacking both pathways exhibit severe defects in telomere protection and fail to localize the protection protein HOAP to telomeres. Drosophila nbs is required for both atm- and atr-dependent DNA damage responses and acts in these pathways during DNA repair. The telomere fusion phenotype of nbs is consistent with defects in each of these activities. Cells defective in both the atm and atr pathways were used to examine if DNA damage response pathways regulate telomere protection without affecting telomere specific sequences. In these cells, chromosome fusion sites retain telomere-specific sequences, demonstrating that loss of these sequences is not responsible for loss of protection. Furthermore, terminally deleted chromosomes also fuse in these cells, directly implicating DNA damage response pathways in the epigenetic protection of telomeres. We propose that recognition of chromosome ends and recruitment of HP1 and HOAP by DNA damage response proteins is essential for the epigenetic protection of Drosophila telomeres. Given the conserved roles of DNA damage response proteins in telomere function, related mechanisms may act at the telomeres of other organisms.     

Posttranslational control of telomere maintenance and the telomere damage response

Telomeres help maintain genome integrity by protecting natural chromosome ends from being recognized as damaged DNA. When telomeres become dysfunctional, they limit replicative lifespan and prevent outgrowth of potentially cancerous cells by activating a DNA damage response that forces cells into senescence or apoptosis. On the other hand, chromosome ends devoid of proper telomere protection are subject to DNA repair activities that cause end-to-end fusions and, when cells divide, extensive genomic instability that can promote cancer. While telomeres represent unique chromatin structures with important roles in cancer and aging, we have limited understanding of the way telomeres and the response to their malfunction are controlled at the level of chromatin. Accumulating evidence indicates that different types of posttranslational modifications act in both telomere maintenance and the response to telomere uncapping. Here, we discuss the latest insights on posttranslational control of telomeric chromatin, with emphasis on ubiquitylation and SUMOylation events.     

Activation of the DNA damage response by telomere attrition: a passage to cellular senescence

Critical telomere shortening induces senescence in many normal human cell types grown in culture. Recent data have revealed that dysfunctional telomeres can resemble certain forms of DNA damage, and point to a role for DNA damage signaling in the establishment and maintenance of telomere-initiated senescence. Here, we review these new observations and highlight potential avenues of future research. We consider the identities of the key DNA damage response factors involved in senescence and discuss a model for the molecular events occurring in pre-senescent cells that ultimately lead to a permanent cell cycle arrest phenotype.     

The DNA damage response: sensing and signaling

The protein kinases ATM and ATR are central components of the checkpoint mechanisms that signal the presence of damaged DNA and stalled replication forks. Recent studies have provided important new insights into how these kinases work together with their regulatory subunits, DNA repair proteins and adaptor proteins to sense abnormal DNA structures and implement the appropriate DNA damage response. These advances have provided a more detailed understanding of the interface between damaged DNA and the checkpoint sensor proteins.     

53BP1 deficiency combined with telomere dysfunction activates ATR-dependent DNA damage response

TRF1 protects mammalian telomeres from fusion and fragility. Depletion of TRF1 leads to telomere fusions as well as accumulation of γ-H2AX foci and activation of both the ataxia telangiectasia mutated (ATM)- and the ataxia telangiectasia and Rad3 related (ATR)-mediated deoxyribonucleic acid (DNA) damage response (DDR) pathways. 53BP1, which is also present at dysfunctional telomeres, is a target of ATM that accumulates at DNA double-strand breaks and favors nonhomologous end-joining (NHEJ) repair over ATM-dependent resection and homology-directed repair (homologous recombination [HR]). To address the role of 53BP1 at dysfunctional telomeres, we generated mice lacking TRF1 and 53BP1. 53BP1 deficiency significantly rescued telomere fusions in mouse embryonic fibroblasts (MEFs) lacking TRF1, but they showed evidence of a switch from the NHEJ- to HR-mediated repair of uncapped telomeres. Concomitantly, double-mutant MEFs showed evidence of hyperactivation of the ATR-dependent DDR. In intact mice, combined 53BP1/TRF1 deficiency in stratified epithelia resulted in earlier onset of DNA damage and increased CHK1 phosphorylation during embryonic development, leading to aggravation of skin phenotypes.     

The DNA damage response in the chromatin context: A coordinated process

In the cell nucleus, DNA damage signaling and repair machineries operate on a chromatin substrate, the integrity of which is critical for cell function and viability. Here, we review recent advances in deciphering the tight coordination between chromatin maintenance and the DNA damage response (DDR). We discuss how the DDR impacts chromatin marks, organization and mobility, and, in turn, how chromatin alterations actively contribute to the DDR, providing additional levels of regulation. We present our current knowledge of the molecular bases of these critical processes in physiological and pathological conditions, and also highlight open questions that emerge in this expanding field.     

DNA processing is not required for ATM-mediated telomere damage response after TRF2 deletion

Telomere attrition and other forms of telomere damage can activate the ATM kinase pathway. What generates the DNA damage signal at mammalian chromosome ends or at other double-strand breaks is not known. Telomere dysfunction is often accompanied by disappearance of the 3' telomeric overhang, raising the possibility that DNA degradation could generate the structure that signals. Here we address these issues by studying telomere structure after conditional deletion of mouse TRF2, the protective factor at telomeres. Upon removal of TRF2 from TRF2(F/-) p53-/- mouse embryo fibroblasts, a telomere damage response is observed at most chromosome ends. As expected, the telomeres lose the 3' overhang and are processed by the non-homologous end-joining pathway. Non-homologous end joining of telomeres was abrogated in DNA ligase IV-deficient (Lig4-/-) cells. Unexpectedly, the telomeres of TRF2-/- Lig4-/- p53-/- cells persisted in a free state without undergoing detectable DNA degradation. Notably, the telomeres retained their 3' overhangs, but they were recognized as sites of DNA damage, accumulating the DNA damage response factors 53BP1 and gamma-H2AX, and activating the ATM kinase. Thus, activation of the ATM kinase pathway at chromosome ends does not require overhang degradation or other overt DNA processing.     

Novel roles for A‐type lamins in telomere biology and the DNA damage response pathway

A‐type lamins are intermediate filament proteins that provide a scaffold for protein complexes regulating nuclear structure and function. Mutations in the LMNA gene are linked to a variety of degenerative disorders termed laminopathies, whereas changes in the expression of lamins are associated with tumourigenesis. The molecular pathways affected by alterations of A‐type lamins and how they contribute to disease are poorly understood. Here, we show that A‐type lamins have a key role in the maintenance of telomere structure, length and function, and in the stabilization of 53BP1, a component of the DNA damage response (DDR) pathway. Loss of A‐type lamins alters the nuclear distribution of telomeres and results in telomere shortening, defects in telomeric heterochromatin, and increased genomic instability. In addition, A‐type lamins are necessary for the processing of dysfunctional telomeres by non‐homologous end joining, putatively through stabilization of 53BP1. This study shows new functions for A‐type lamins in the maintenance of genomic integrity, and suggests that alterations of telomere biology and defects in DDR contribute to the pathogenesis of lamin‐related diseases.     

The DNA damage response: ten years after

The DNA damage response (DDR), through the action of sensors, transducers, and effectors, orchestrates the appropriate repair of DNA damage and resolution of DNA replication problems, coordinating these processes with ongoing cellular physiology. In the past decade, we have witnessed an explosion in understanding of DNA damage sensing, signaling, and the complex interplay between protein phosphorylation and the ubiquitin pathway employed by the DDR network to execute the response to DNA damage. These findings have important implications for aging and cancer.     

Chromatin and the DNA damage response

The impact of chromatin structure upon the DNA damage response is becoming increasingly apparent. We can reasonably expect many more papers showing how chromatin and chromatin modifications impact upon aspects of the DNA damage response. Here, we present our perspective on some recent developments in this exciting area of cell biology. We aim that this review will be of interest to those who study the DNA damage response, but not usually in the context of chromatin, and equally to those who study chromatin, but not the DNA damage response. It seems likely that these two communities will increasingly share common questions and interests.     

Ubiquitin and the DNA damage response

Comment on: Gatti M, et al. Cell Cycle 2012; 11:2538-44.     

Retroviral DNA integration and the DNA damage response

Retroviral DNA integration creates a discontinuity in the host cell chromatin and repair of this damage is required to complete the integration process. As integration and repair are essential for both viral replication and cell survival, it is possible that specific interactions with the host DNA repair systems might provide new cellular targets for human immunodeficiency virus therapy. Various genetic, pharmacological, and biochemical studies have provided strong evidence that postintegration DNA repair depends on components of the nonhomologous end-joining (NHEJ) pathway (DNA-PK (DNA-dependent protein kinase), Ku, Xrcc4, DNA ligase IV) and DNA damage-sensing pathways (Atr (Atm and Rad related), gamma-H2AX). Furthermore, deficiencies in NHEJ components result in susceptibility to apoptotic cell death following retroviral infection. Here, we review these findings and discuss other ways that retroviral DNA intermediates may interact with the host DNA damage signaling and repair pathways.