Dataset Information

RAD51AP1 and RAD54L Can Underpin Two Distinct RAD51-Dependent Routes of DNA Damage Repair via Homologous Recombination.

ABSTRACT: Homologous recombination DNA repair (HR) is a complex DNA damage repair pathway and an attractive target of inhibition in anti-cancer therapy. To help guide the development of efficient HR inhibitors, it is critical to identify compensatory HR sub-pathways. In this study, we describe a novel synthetic interaction between RAD51AP1 and RAD54L, two structurally unrelated proteins that function downstream of the RAD51 recombinase in HR. We show that concomitant deletion of RAD51AP1 and RAD54L further sensitizes human cancer cell lines to treatment with olaparib, a Poly (adenosine 5'-diphosphate-ribose) polymerase inhibitor, to the DNA inter-strand crosslinking agent mitomycin C, and to hydroxyurea, which induces DNA replication stress. We also show that the RAD54L paralog RAD54B compensates for RAD54L deficiency, although, surprisingly, less extensively than RAD51AP1. These results, for the first time, delineate RAD51AP1- and RAD54L-dependent sub-pathways and will guide the development of inhibitors that target HR stimulators of strand invasion.

SUBMITTER: Selemenakis P

PROVIDER: S-EPMC9149245 | biostudies-literature | 2022

REPOSITORIES: biostudies-literature

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RAD51AP1 and RAD54L Can Underpin Two Distinct RAD51-Dependent Routes of DNA Damage Repair via Homologous Recombination.

Selemenakis Platon P Sharma Neelam N Uhrig Mollie E ME Katz Jeffrey J Kwon Youngho Y Sung Patrick P Wiese Claudia C

Frontiers in cell and developmental biology 20220516

Homologous recombination DNA repair (HR) is a complex DNA damage repair pathway and an attractive target of inhibition in anti-cancer therapy. To help guide the development of efficient HR inhibitors, it is critical to identify compensatory HR sub-pathways. In this study, we describe a novel synthetic interaction between RAD51AP1 and RAD54L, two structurally unrelated proteins that function downstream of the RAD51 recombinase in HR. We show that concomitant deletion of RAD51AP1 and RAD ...[more]

PMID: 35652094

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Project description:ObjectiveThe current study aims to identify the dysregulated pathway involved in carcinogenesis and the essential survival-related dysregulated genes among this pathway in the early stage of lung adenocarcinoma (LUAD).Patients and methodsData from The Cancer Genome Atlas (TCGA) including 526 tumor tissues of LUAD and 59 healthy lung tissues were analyzed to gain differentially expressed genes (DEGs). Gene ontology (GO) analysis was conducted with DAVID, while the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of DEGs was performed, followed by gene set enrichment analysis (GSEA) methods. Survival analysis was implemented in TCGA dataset and validated in Gene Expression Omnibus (GEO) cohort GSE50081, which includes 127 patients with stage I LUAD.ResultsGSEA enrichment analysis suggested that homologous recombination repair (HRR) pathway was significantly enriched. Subsequent KEGG pathway enrichment analysis indicated the significant up-regulation of HRR pathway in patients with T1 stage LUAD. Retrieved in Gene database, RAD54L is involved in HRR pathway and were recognized to be significantly differentially expressed in T1 stage LUAD in our study. The survival analysis indicated that high expression of RAD54L was significantly related to worse overall survival in patients with T1 stage LUAD (TCGA cohort: HR=2.10, 95% CI [1.47-2.98], P = 0.001; GSE50081 validation cohort: HR = 2.61, 95% CI [1.51-4.52], P = 0.002). Multivariate cox regression analysis indicated that RAD54L is an independent prognostic factor in the early-stage LUAD.ConclusionHRR pathway is up-regulated in LUAD, among which the expression of RAD54L was found to be significantly differentially expressed in T1 stage tumor tissue. Patients with high expression of RAD54L were associated with worse overall survival in the TCGA cohort and validation cohort. This study suggests a potential mechanism of lung cancer progression and provide a budding prognostic factor and treatment target in early-stage LUAD.

Project description:BackgroundRAD51-dependent homologous recombination (HR) is one of the most important pathways for repairing DNA double-strand breaks (DSBs), and its regulation is crucial to maintain genome integrity. Elp1 gene encodes IKAP/ELP1, a core subunit of the Elongator complex, which has been implicated in translational regulation. However, how ELP1 contributes to genome maintenance is unclear.MethodsTo investigate the function of Elp1, Elp1-deficient mouse embryonic fibroblasts (MEFs) were generated. Metaphase chromosome spreading, immunofluorescence, and comet assays were used to access chromosome abnormalities and DSB formation. Functional roles of Elp1 in MEFs were evaluated by cell viability, colony forming capacity, and apoptosis assays. HR-dependent DNA repair was assessed by reporter assay, immunofluorescence, and western blot. Polysome profiling was used to evaluate translational efficiency. Differentially expressed proteins and signaling pathways were identified using a label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS) proteomics approach.ResultsHere, we report that Elp1 depletion enhanced genomic instability, manifested as chromosome breakage and genotoxic stress-induced genomic DNA fragmentation upon ionizing radiation (IR) exposure. Elp1-deficient cells were hypersensitive to DNA damage and exhibited impaired cell proliferation and defective HR repair. Moreover, Elp1 depletion reduced the formation of IR-induced RAD51 foci and decreased RAD51 protein levels. Polysome profiling analysis revealed that ELP1 regulated RAD51 expression by promoting its translation in response to DNA damage. Notably, the requirement for ELP1 in DSB repair could be partially rescued in Elp1-deficient cells by reintroducing RAD51, suggesting that Elp1-mediated HR-directed repair of DSBs is RAD51-dependent. Finally, using proteome analyses, we identified several proteins involved in cancer pathways and DNA damage responses as being differentially expressed upon Elp1 depletion.ConclusionsOur study uncovered a molecular mechanism underlying Elp1-mediated regulation of HR activity and provides a novel link between translational regulation and genome stability.

Dataset Information

RAD51AP1 and RAD54L Can Underpin Two Distinct RAD51-Dependent Routes of DNA Damage Repair via Homologous Recombination.

Publications

RAD51AP1 and RAD54L Can Underpin Two Distinct RAD51-Dependent Routes of DNA Damage Repair <i>via</i> Homologous Recombination.

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