Dataset Information

UFMylation orchestrates chromatin engagement of core NHEJ components to promote DNA double-strand break repair

ABSTRACT: DNA double-strand breaks (DSBs) are highly cytotoxic lesions that – if misrepaired – can lead to genomic instability, cancer, and developmental disorders. While ubiquitin and select ubiquitin-like proteins (UBLs) are established regulators of DSB repair, many UBLs remain understudied. Here, through systematic UBL screening, we identify UFM1 as a novel regulator of non-homologous end-joining (NHEJ). Using a photo-crosslinkable UFM1 probe, we uncover core NHEJ components, including XRCC4, as downstream UFM1 receptors. Complementary proximity screening reveals UFMylated Ku70 as a key upstream target recognised by XRCC4. Mechanistically, UFMylation facilitates chromatin recruitment and assembly of NHEJ factors, establishing a central role in end-joining repair. Our findings link hereditary UFMylation disorders to defective NHEJ processes, contributing to our molecular understanding of diseases biology. Beyond this, our bottom-up screening strategy reveals broader networks of UFM1-regulated factors, featuring diverse roles in genome maintenance and cellular homeostasis.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Petra Beli

LAB HEAD: Petra Beli

PROVIDER: PXD069185 | Pride | 2026-07-31

REPOSITORIES: Pride

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Dataset's files

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			Action	DRS
	20191213_QEP1_PBE_FC_U2OS_UFC1_APEX_rep1_1.raw	Raw
	20191213_QEP1_PBE_FC_U2OS_UFC1_APEX_rep1_2.raw	Raw
	20191213_QEP1_PBE_FC_U2OS_UFC1_APEX_rep1_3.raw	Raw
	20191213_QEP1_PBE_FC_U2OS_UFC1_APEX_rep1_4.raw	Raw
	20191213_QEP1_PBE_FC_U2OS_UFC1_APEX_rep1_5.raw	Raw

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Publications

Systematic discovery of UFM1 receptors reveals a regulatory module in DNA repair directing non-homologous end-joining.

Wang Zijuan Z Foster Benjamin M BM da Costa Isabelle C IC Wu Yue Y Behera Deepak D Conte Francesca F Trotter Eleanor W EW Lopezcolorado Felicia Wednesday FW Cabello-Lobato Maria Jose MJ Choudhary Shweta S Wiener Reuven R Beli Petra P Smith Duncan L DL Banks William H WH Bagley Steven S McKee Shane S Minnis Meenakshi M Meyer Stefan S Chaplin Amanda K AK Dörner Wolfgang W Mootz Henning D HD Hagan Iain M IM Galanty Yaron Y Stark Jeremy M JM Larrosa Igor I Cliff Matthew J MJ Schmidt Christine K CK

Nature communications 20260615 1

Posttranslational modifications with ubiquitin-like modifiers (UBLs) are critical for genome maintenance, yet many remain mechanistically uncharacterised. Here, we identify UFM1 as a key regulator of non-homologous end-joining (NHEJ), a major DNA double-strand break repair pathway. Using a structure-guided chemical biology approach, we develop a photo-crosslinkable UFM1 probe and, in combination with NMR, map non-canonical UFM1-binding interfaces in core NHEJ factors, including the disordered XR ...[more]

PMID: 42297806

Similar Datasets

Project description:Genetic variants that hinder post-translational protein modifications by UFM1, UFMylation, cause encephalopathies. UFMylation regulates endoplasmic reticulum (ER) homeostasis, but how UFMylation-deficiencies cause selective neurological defects is unknown. Using Ufm1 knock-out mice, we investigated two types of UFMylation pathologies, UFM1 loss and expression of a pathogenic UFM1-R81C variant. We found that UFM1-deficiency confounds neuron development and synapse function. Mechanistically, UFM1 loss is associated with induction of ER stress, activation of the PERK-UPR pathway, and reduced protein translation. These defects are rescued by wild-type UFM1, but only partially by UFM1-R81C. UFM1-deficient and UFM1-R81Cexpressing neurons display distinct responses to ER stress, indicating that UFM1-R81C is not merely a loss-of-function variant. Exploring therapeutic options, we show that the PERK-UPR inhibitor Trazodone restores protein translation solely in UFM1-R81C-expressing neurons, and increases synapse number. Our study unveils a pivotal role for UFMylation in neuronal development, provides a molecular understanding of the signaling mechanisms altered in UFM1- associated encephalopathies, and offers important insights into potential treatments for these Genetic variants that hinder post-translational protein modifications by UFM1, UFMylation, cause encephalopathies. UFMylation regulates endoplasmic reticulum (ER) homeostasis, but how UFMylation-deficiencies cause selective neurological defects is unknown. Using Ufm1 knock-out mice, we investigated two types of UFMylation pathologies, UFM1 loss and expression of a pathogenic UFM1-R81C variant. We found that UFM1-deficiency confounds neuron development and synapse function. Mechanistically, UFM1 loss is associated with induction of ER stress, activation of the PERK-UPR pathway, and reduced protein translation. These defects are rescued by wild-type UFM1, but only partially by UFM1-R81C. UFM1-deficient and UFM1-R81C-expressing neurons display distinct responses to ER stress, indicating that UFM1-R81C is not merely a loss-of-function variant. Exploring therapeutic options, we show that the PERK-UPR inhibitor Trazodone restores protein translation solely in UFM1-R81C-expressing neurons, initiating an increase in synapse number. Our study unveils a pivotal role for UFMylation in neuronal development and provides a molecular understanding of the signaling mechanisms altered in UFM1-associated encephalopathies, offering valuable insights into potential treatments for these disorders.

Project description:Classical non-homologous end-joining (C-NHEJ) is a major mammalian DNA double strand break (DSB) repair pathway. Core C-NHEJ factors, such as XRCC4, are required for joining DSB intermediates of the G1 phase-specific V(D)J recombination reaction in progenitor lymphocytes. Core factors also contribute to joining DSBs in cycling mature B-lineage cells, including DSBs generated during antibody class switch recombination (CSR) and DSBs generated by ionizing radiation (IR). The XLF C-NHEJ factor is dispensable for V(D)J recombination in normal cells, but, due to functional redundancy, is absolutely required for this process in cells deficient for the ATM DSB response factor. The recently identified PAralogue of XRCC4 and XLF (PAXX) factor has homology to these two proteins and variably contributes to IR-induced DSB repair in human and chicken cells. We now report that PAXX is dispensable for joining V(D)J recombination DSBs in G1-arrested mouse pro-B cell lines, dispensable for joining CSR-associated DSBs in a cycling mouse B cell line, and dispensable for normal IR-resistance in both G1-arrested and cycling pro-B lines. However, we find that combined deficiency for PAXX and XLF in G1-arrested pro-B lines abrogates DSB joining during V(D)J recombination and sensitizes the cells to IR exposure. Thus, PAXX provides core C-NHEJ factor-associated functions in the absence of XLF and vice versa in G1-arrested Pro-B cell lines. Finally, we also find that PAXX-deficiency has no impact on V(D)J recombination DSB joining in ATM-deficient pro-B lines. We discuss implications of these findings with respect to potential PAXX and XLF functions in C-NHEJ. Examination of CSR Switch mu-to-alpha junctions from mu bait DSBs using LAM-HTGTS and Illumina Miseq. Two clones of PAXX-/- and XLF-/- and 1 clone of Ligase4-/- were derived from the parental CH12F3 line; the second Ligase4-/- clone was acquired from Keifei Yu. Two clones of XLF-/-PAXX-/-CH12 cells were derived from one of the PAXX-/- clones. Three biological replicates were analyzed for each clone.