{"doi":"10.1016/j.jbc.2022.101672","title":"Oligomerization of DNA replication regulatory protein RADX is essential to maintain replication fork stability","abstract":"Genome integrity requires complete and accurate DNA replication once per cell division cycle. Replication stress poses obstacles to this process that must be overcome to prevent replication fork collapse. An important regulator of replication fork stability is the RAD51 protein, which promotes replication fork reversal and protects nascent DNA strands from nuclease-mediated degradation. Many regulatory proteins control these RAD51 activities, including RADX, which binds both ssDNA and RAD51 at replication forks to ensure that fork reversal is confined to stalled forks. Many ssDNA-binding proteins function as hetero- or homo-oligomers. In this study, we addressed whether this is also the case for RADX. Using biochemical and genetic approaches, we found that RADX acts as a homo-oligomer to control replication fork stability. RADX oligomerizes using at least two different interaction surfaces, including one mapped to a C-terminal region. We demonstrate that mutations in this region prevent oligomerization and prevent RADX function in cells, and that addition of a heterologous dimerization domain to the oligomerization mutants restored their ability to regulate replication. Taken together, our results demonstrate that like many ssDNA-binding proteins, oligomerization is essential for RADX-mediated regulation of genome stability. Genome integrity requires complete and accurate DNA replication once per cell division cycle. Replication stress poses obstacles to this process that must be overcome to prevent replication fork collapse. An important regulator of replication fork stability is the RAD51 protein, which promotes replication fork reversal and protects nascent DNA strands from nuclease-mediated degradation. Many regulatory proteins control these RAD51 activities, including RADX, which binds both ssDNA and RAD51 at replication forks to ensure that fork reversal is confined to stalled forks. Many ssDNA-binding proteins function as hetero- or homo-oligomers. In this study, we addressed whether this is also the case for RADX. Using biochemical and genetic approaches, we found that RADX acts as a homo-oligomer to control replication fork stability. RADX oligomerizes using at least two different interaction surfaces, including one mapped to a C-terminal region. We demonstrate that mutations in this region prevent oligomerization and prevent RADX function in cells, and that addition of a heterologous dimerization domain to the oligomerization mutants restored their ability to regulate replication. Taken together, our results demonstrate that like many ssDNA-binding proteins, oligomerization is essential for RADX-mediated regulation of genome stability. The human genome is replicated with high fidelity once per cell division cycle. A highly regulated ensemble of proteins involved in DNA synthesis, chromatin deposition, DNA repair, and replication stress responses ensure replication fork stability and completion of genome duplication in a timely manner. Fork reversal is one of the mechanisms of replication stress tolerance that facilitates the repair or bypass of DNA damage and other replication stresses (1Berti M. Cortez D. Lopes M. The plasticity of DNA replication forks in response to clinically relevant genotoxic stress.Nat. Rev. Mol. Cell Biol. 2020; 21: 633-651Google Scholar, 2Cortez D. Replication-coupled DNA repair.Mol. Cell. 2019; 74: 866-876Google Scholar). RAD51, the recombinase in homology-directed double-strand break repair, works in cooperation with ATP-dependent motor proteins like SMARCAL1, ZRANB3, HLTF, and FBH1 to catalyze fork reversal (3Bai G. Kermi C. Stoy H. Schiltz C.J. Bacal J. Zaino A.M. Hadden M.K. Eichman B.F. Lopes M. Cimprich K.A. HLTF promotes fork reversal, limiting replication stress resistance and preventing multiple mechanisms of unrestrained DNA synthesis.Mol. Cell. 2020; 78: 1237-1251.e7Google Scholar, 4Betous R. Mason A.C. Rambo R.P. Bansbach C.E. Badu-Nkansah A. Sirbu B.M. Eichman B.F. Cortez D. SM","journal":"Journal of Biological Chemistry","year":2022,"id":278464,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":6,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9544,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2022-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":388842,"name":"Madison B. Adolph","orcid":"0000-0002-4761-7388","position":1,"is_corresponding":false},{"id":252498,"name":"David Cortez","orcid":"0000-0003-0154-140X","position":2,"is_corresponding":false},{"id":698133,"name":"Taha M. Mohamed","orcid":"0000-0002-2752-9663","position":0,"is_corresponding":true}],"reference_count":21,"raw_metadata":null,"created_at":"2026-07-19T00:28:47.357993Z","pmid":"35120927","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}