p53-dependent chromatin relaxation is required for DNA double-strand break repair

Abstract

The tumor suppressor p53, an indispensable nuclear transcription factor, plays a central role in orchestrating cellular responses when DNA damage occurs. In this study, we demonstrate that in the initial phases of DNA double-strand break (DSB) repair, p53 is rapidly recruited to sites of damage and the surrounding chromatin, where it enhances DSB repair efficiency. This enhancement occurs through the modulation of chromatin dynamics and the promotion of a more relaxed chromatin configuration, a process influenced by p53 in response to DSB-inducing factors such as etoposide, ultraviolet radiation, and nucleases. These results underscore the pivotal function of p53 as a rapid responder to DSBs, delineating a significant departure from its traditionally recognized role as a downstream transcriptional regulator in DNA damage repair processes. This study emphasizes that the direct engagement of p53 in DNA repair through chromatin structure regulation extends beyond its established involvement in UV irradiation-induced nucleotide excision repair (NER), demonstrating analogous mechanistic attributes in the context of DSB repair. This newly illuminated perspective enhances our understanding of the multifaceted roles of p53 in genome stability and integrity.

1. Introduction

DNA, while being a comparatively stable organic molecule, is perpetually exposed to a multitude of endogenous and exogenous damaging agents. In response to this relentless challenge, cells have developed an intricate array of biochemical pathways designed to mitigate such threats [1,2]. This advanced regulatory network is collectively referred to as the DNA damage response (DDR), which orchestrates a variety of critical biological outcomes, such as cell cycle arrest, DNA repair, apoptosis, and cellular senescence [3]. The components of the DDR are strategically classified into sensors, signal transducers, and effectors to address cellular challenges efficiently. A DNA double-strand break (DSB) represents a significant challenge for cells, with accurate repair being crucial for maintaining cellular survival and preventing oncogenic translocations [4]. A paradigmatic example of the organizational structure of the DDR is the recruitment of the MRN (MRE11-RAD50-NBS1) complex to DSB sites. The MRN complex is instrumental in recruiting and activating the ataxia telangiectasia mutated (ATM) kinase, which is pivotal in the DDR. In response to DNA damage, ATM transitions from an inactive dimer to an active monomer through autophosphorylation at key serine residues, such as S1981 [5]. Once activated at a damage site, ATM phosphorylates H2AX at S139, producing γ-H2AX, which in turn recruits MDC1, establishing a feedback loop that enhances ATM signaling by attracting more MRN complexes and ATM molecules [6]. This activation cascade also involves other critical factors, such as 53BP1, BRCA1, and the ubiquitin ligases RNF8 and RNF168 [7,8]. Among the most prolific ATM substrates, the tumor suppressor p53 is at the top [9].

DSB repair involves two major pathways: homologous recombination (HR) and nonhomologous end-joining (NHEJ). The key difference between HR and NHEJ is that HR is recruited exclusively during the S and G2 phases of the cell cycle; in contrast, NHEJ can be activated during all cell cycle phases [10,11]. HR is an error-free repair mechanism that uses a homologous template to accurately reassemble damaged DNA strands. Key proteins in HR include the MRN complex, BRCA1/BRCA2, ATM, and ATR, which are essential for initiating and regulating this process [11]. Conversely, NHEJ lacks a homologous template, directly rejoining DNA ends and thus being inherently error prone. Critical NHEJ factors include the Ku70/Ku80 complex, DNA-dependent protein kinases, and XRCC4, which are pivotal in this repair pathway [10]. The eukaryotic DSB response takes place on chromatin near the break site; thus, the basic organization of eukaryotic chromatin needs to be considered. Chromatin relaxation is considered a fundamental pathway in the DNA damage response [12].

P53 was initially found to be associated with the T antigen of simian virus 40 in rodent cells that had undergone transformation [13], and its function as a tumor suppressor was first acknowledged in 1989 [14,15]. Under nonstressful conditions, the E3 ubiquitin ligase MDM2 maintains minimal p53 levels [16,17]. However, with the activation of cellular stress, p53 becomes phos...