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      The roles of the human SETMAR (Metnase) protein in illegitimate DNA recombination and non-homologous end joining repair

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          Highlights

          • Full length SETMAR expression has no effect on DNA repair and integration in vivo.

          • SETMAR putative nuclease activity is not required in vivo.

          • Separate expression of the SET and MAR domains affects DNA repair and integration.

          • SETMAR isoform with a truncated SET-domain is specific to species containing the MAR domain.

          Abstract

          SETMAR is a fusion between a SET-domain methyltransferase gene and a mariner-family transposase gene, which is specific to anthropoid primates. However, the ancestral SET gene is present in all other mammals and birds. SETMAR is reported to be involved in transcriptional regulation and a diverse set of reactions related to DNA repair. Since the transcriptional effects of SETMAR depend on site-specific DNA binding, and are perturbed by inactivating the methyltransferase, we wondered whether we could differentiate the effects of the SET and MAR domains in DNA repair assays. We therefore generated several stable U2OS cell lines expressing either wild type SETMAR or truncation or point mutant variants. We tested these cell lines with in vivo plasmid-based assays to determine the relevance of the different domains and activities of SETMAR in DNA repair. Contrary to previous reports, we found that wild type SETMAR had little to no effect on the rate of cell division, DNA integration into the genome or non-homologous end joining. Also contrary to previous reports, we failed to detect any effect of a strong active-site mutation that should have knocked out the putative nuclease activity of SETMAR.

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          Most cited references30

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          Regulation of MLL1 H3K4 methyltransferase activity by its core components.

          Histone H3 Lys4 (H3K4) methylation is a prevalent mark associated with transcription activation. A common feature of several H3K4 methyltransferase complexes is the presence of three structural components (RbBP5, Ash2L and WDR5) and a catalytic subunit containing a SET domain. Here we report the first biochemical reconstitution of a functional four-component mixed-lineage leukemia protein-1 (MLL1) core complex. This reconstitution, combined with in vivo assays, allows direct analysis of the contribution of each component to MLL1 enzymatic activity and their roles in transcriptional regulation. Moreover, taking clues from a crystal structure analysis, we demonstrate that WDR5 mediates interactions of the MLL1 catalytic unit both with the common structural platform and with the histone substrate. Mechanistic insights gained from this study can be generalized to the whole family of SET1-like histone methyltransferases in mammals.
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            Comprehensive Mapping of Histone Modifications at DNA Double-Strand Breaks Deciphers Repair Pathway Chromatin Signatures

            Summary Double-strand breaks (DSBs) are extremely detrimental DNA lesions that can lead to cancer-driving mutations and translocations. Non-homologous end joining (NHEJ) and homologous recombination (HR) represent the two main repair pathways operating in the context of chromatin to ensure genome stability. Despite extensive efforts, our knowledge of DSB-induced chromatin still remains fragmented. Here, we describe the distribution of 20 chromatin features at multiple DSBs spread throughout the human genome using ChIP-seq. We provide the most comprehensive picture of the chromatin landscape set up at DSBs and identify NHEJ- and HR-specific chromatin events. This study revealed the existence of a DSB-induced monoubiquitination-to-acetylation switch on histone H2B lysine 120, likely mediated by the SAGA complex, as well as higher-order signaling at HR-repaired DSBs whereby histone H1 is evicted while ubiquitin and 53BP1 accumulate over the entire γH2AX domains.
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              Methylation of histone H3 lysine 36 enhances DNA repair by nonhomologous end-joining.

              Given its significant role in the maintenance of genomic stability, histone methylation has been postulated to regulate DNA repair. Histone methylation mediates localization of 53BP1 to a DNA double-strand break (DSB) during homologous recombination repair, but a role in DSB repair by nonhomologous end-joining (NHEJ) has not been defined. By screening for histone methylation after DSB induction by ionizing radiation we found that generation of dimethyl histone H3 lysine 36 (H3K36me2) was the major event. Using a novel human cell system that rapidly generates a single defined DSB in the vast majority of cells, we found that the DNA repair protein Metnase (also SETMAR), which has a SET histone methylase domain, localized to an induced DSB and directly mediated the formation of H3K36me2 near the induced DSB. This dimethylation of H3K36 improved the association of early DNA repair components, including NBS1 and Ku70, with the induced DSB, and enhanced DSB repair. In addition, expression of JHDM1a (an H3K36me2 demethylase) or histone H3 in which K36 was mutated to A36 or R36 to prevent H3K36me2 formation decreased the association of early NHEJ repair components with an induced DSB and decreased DSB repair. Thus, these experiments define a histone methylation event that enhances DNA DSB repair by NHEJ.
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                Author and article information

                Contributors
                Journal
                DNA Repair (Amst)
                DNA Repair (Amst.)
                DNA Repair
                Elsevier
                1568-7864
                1568-7856
                1 August 2019
                August 2019
                : 80
                : 26-35
                Affiliations
                [0005]School of Life Sciences, University of Nottingham, Queen’s Medical Centre, Nottingham, NG7 2UH, UK
                Author notes
                [* ]Corresponding author. ronald.chalmers@ 123456nottingham.ac.uk
                [1]

                Present address: Sir William Dunn School of Pathology, University of Oxford, Oxford, OX1 3RE, UK.

                Article
                S1568-7864(19)30092-8
                10.1016/j.dnarep.2019.06.006
                6715855
                31238295
                b27d06ab-987d-4d5c-8f1c-d25495ec6898
                © 2019 The Authors

                This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

                History
                : 18 March 2019
                : 18 June 2019
                : 18 June 2019
                Categories
                Article

                Genetics
                dsb, dna double-strand break,nhej, non-homologous end joining,setmar,dna integration,non-homologous end joining,dna double-strand break repair,metnase

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