Abstract
The human MRE11/RAD50/NBS1 (MRN) complex plays a crucial role in sensing and repairing DNA DSB. MRE11 possesses dual 3′−5′ exonuclease and endonuclease activity and forms the core of the multifunctional MRN complex. We previously identified a C-terminally truncated form of MRE11 (TR-MRE11) associated with post-translational MRE11 degradation. Here we identified SPRTN as the essential protease for the formation of TR-MRE11 and characterised the role of this MRE11 form in its DNA damage response (DDR). Using tandem mass spectrometry and site-directed mutagenesis, the SPRTN-dependent cleavage site for MRE11 was identified between 559 and 580 amino acids. Despite the intact interaction of TR-MRE11 with its constitutive core complex proteins RAD50 and NBS1, both nuclease activities of truncated MRE11 were dramatically reduced due to its deficient binding to DNA. Furthermore, lack of the MRE11 C-terminal decreased HR repair efficiency, very likely due to abolished recruitment of TR-MRE11 to the sites of DNA damage, which consequently led to increased cellular radiosensitivity. The presence of this DNA repair-defective TR-MRE11 could explain our previous finding that the high MRE11 protein expression by immunohistochemistry correlates with improved survival following radical radiotherapy in bladder cancer patients.
Highlights
MRE11 is a component of the conservedMRE11–RAD50–NBS1 (MRN) complex, a key player in the early stages of the DNA damage response (DDR)
In cell-free extracts taken from primary bladder tumours at transurethral tumour resection, we found the presence of the truncation in tumour samples from bladder cancer patients, but this was not associated with clinical stage (Ta, T1 and T2) (Fig. 1B)
The reason why high MRE11 protein expression, by immunohistochemistry using an antibody with an epitope spanning amino acids 182–582, is associated with better survival rate in bladder cancer patients after radiotherapy[34,35] has not yet been explained
Summary
MRE11 is a component of the conservedMRE11–RAD50–NBS1 (MRN) complex, a key player in the early stages of the DNA damage response (DDR). MRN is responsible for the recognition, repair, and signalling of DNA double-strand breaks (DSB)[1,2] and is required for homologous recombination (HR), classical non-homologous end-joining (C-NHEJ) and alternative non-homologous end-joining (A-NHEJ) pathways in detection and signalling of DSBs3. Its effects are mediated via its 3′−5′ exonuclease and single-stranded (ss) and DNA hairpin endonuclease activities[4,5,6,7]. Absence of any of its components leads to embryonic lethality in mammals[4]. The 3′−5′ exo- and endonuclease activities of MRE11 have specialised roles in HR repair and in supporting ataxia-telangiectasia-mutated and Rad3- related (ATR) kinase activation[8,9]. Abrogation of MRE11 nuclease activity causes a striking array of phenotypes, indistinguishable from the absence of MRN, including early embryonic lethality and marked genomic instability[2,9]
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