Key Spec Table
|Species Reactivity||Key Applications||Host||Format||Antibody Type|
|H, M||IP, WB||M||Purified||Monoclonal Antibody|
|Description||Anti-Rad50 Antibody, clone 13B3/2C6|
|Presentation||10mM PBS, pH 7.4|
|Application||Anti-Rad50 Antibody, clone 13B3/2C6 is a high quality Mouse Monoclonal Antibody for the detection of Rad50 & has been validated in IP & WB.|
|Safety Information according to GHS|
|Storage and Shipping Information|
|Storage Conditions||2 years at -20°C|
|Material Size||100 µg|
|Anti-Rad50, clone 13B3/2C6||3026666|
|Anti-Rad50, clone 13B3/2C6||2883130|
|Anti-Rad50, clone 13B3/2C6 - 2424648||2424648|
|Anti-Rad50, clone 13B3/2C6 - 1955409||1955409|
|Anti-Rad50, clone 13B3/2C6 - 1958296||1958296|
|Anti-Rad50, clone 13B3/2C6 - 2043183||2043183|
|Anti-Rad50, clone 13B3/2C6 - 2204863||2204863|
|Anti-Rad50, clone 13B3/2C6 - 2326276||2326276|
|Anti-Rad50, clone 13B3/2C6 - 2519318||2519318|
|Anti-Rad50, clone 13B3/2C6 - DAM1641085||DAM1641085|
|Reference overview||Application||Pub Med ID|
|Hyperthermia alters the interaction of proteins of the Mre11 complex in irradiated cells.|
Gerashchenko, BI; Gooding, G; Dynlacht, JR
Cytometry. Part A : the journal of the International Society for Analytical Cytology 77 940-52 2010
Radiosensitization of mammalian cells by heat is believed to involve the inhibition of repair of DNA double-strand breaks (DSBs). The Mre11 complex (composed of Mre11, Rad50, and Nbs1) is involved in DSB repair and forms foci at sites of radiation-induced DSBs. Heat induces the translocation of a significant amount of Mre11, Rad50, and Nbs1 from the nucleus to the cytoplasm, but little is known about how heat affects the integrity of the proteins still remaining in nuclei, or alters kinetics of formation/disappearance of DNA repair foci in heated, irradiated cells. Here, we show that hyperthermia alters the interaction between proteins of the Mre11 complex in irradiated human melanoma cells and inhibits the formation of repair foci. At various times after X-irradiation and/or heating (2 h at 41.5 or 42.5 °C), the cells were fixed and stained for Mre11, Rad50, and Nbs1. Colocalization of proteins and formation and disappearance of nuclear foci in heated and/or irradiated cells, determined using confocal microscopy, were compared. In heated, irradiated cells, focus formation was inhibited for 2-8 h, and colocalization of the proteins of the Mre11 complex was reduced for 12-24 h post-treatment. Colocalization was recovered in irradiated cells within 24 h after heating at 41.5 °C, but was inhibited longer after heating at 42.5 °C. The decreased colocalization in heated, irradiated cells suggests that there is a decrease in protein interaction, and Mre11 complexes in nuclei disassemble after heating. Such changes could be involved, at least in part, in heat radiosensitization and inhibition of DSB repair. Also, the kinetics of disassembly and reassembly of Mre11 complexes appears to be dependent upon treatment temperature.
|A tool for enhancement and scoring of DNA repair foci.|
Gerashchenko, BI; Dynlacht, JR
Cytometry. Part A : the journal of the International Society for Analytical Cytology 75 245-52 2009
Upon induction of DNA double-strand breaks (DSBs), Mre11 and Rad50 proteins of the Mre11 DNA repair complex accumulate at the sites of DSBs and form discrete nuclear foci. Precision in scoring of Mre11/Rad50-containing foci depends upon detection of those foci, some of which have a fluorescence staining intensity that is too close to the fluorescence staining intensity of the remaining Mre11 and Rad50 proteins that have not been incorporated into foci. Human U-1 melanoma cells in exponential growth were irradiated with various doses of X-rays (0-12 Gy) to induce the formation of repair foci. Four hours after irradiation, cells were simultaneously labeled for Mre11 and Rad50 proteins, using a two-color immunofluorescence staining technique. Laser scanning confocal microscopy was used to collect the composite images of randomly selected cell nuclei. Intensity correlation analysis (ICA) of equally intense fluorescence signals from Mre11 and Rad50 proteins was performed to obtain the regions with correlated pixels. ICA permitted enhanced detection of low level fluorescence of Mre11/Rad50 foci ("hidden" foci) that can be barely detected upon imaging of only one protein. For example, while imaging of only one protein (either Mre11 or Rad50) in the nucleus of a 6 Gy-irradiated cell revealed 9 foci, imaging of two proteins with ICA revealed 11 foci. ICA permitted an evaluation of the dose dependence of nuclear foci in cells irradiated with various doses of X-rays, with focus formation increasing up to a dose of 6 Gy. Our data accumulated using two-color immunofluorescence staining of Mre11 and Rad50 proteins and ICA of these two target proteins provide a basis for enhanced detection and accuracy in the scoring of DNA repair foci.
|Functional role for senataxin, defective in ataxia oculomotor apraxia type 2, in transcriptional regulation.|
Suraweera, A; Lim, Y; Woods, R; Birrell, GW; Nasim, T; Becherel, OJ; Lavin, MF
Human molecular genetics 18 3384-96 2009
Ataxia oculomotor apraxia type 2 (AOA2) is an autosomal recessive neurodegenerative disorder characterized by cerebellar ataxia and oculomotor apraxia. The gene mutated in AOA2, SETX, encodes senataxin, a putative DNA/RNA helicase which shares high homology to the yeast Sen1p protein and has been shown to play a role in the response to oxidative stress. To investigate further the function of senataxin, we identified novel senataxin-interacting proteins, the majority of which are involved in transcription and RNA processing, including RNA polymerase II. Binding of RNA polymerase II to candidate genes was significantly reduced in senataxin deficient cells and this was accompanied by decreased transcription of these genes, suggesting a role for senataxin in the regulation/modulation of transcription. RNA polymerase II-dependent transcription termination was defective in cells depleted of senataxin in keeping with the observed interaction of senataxin with poly(A) binding proteins 1 and 2. Splicing efficiency of specific mRNAs and alternate splice-site selection of both endogenous genes and artificial minigenes were altered in senataxin depleted cells. These data suggest that senataxin, similar to its yeast homolog Sen1p, plays a role in coordinating transcriptional events, in addition to its role in DNA repair.
|Mutations in the MRE11, RAD50, XRS2, and MRE2 genes alter chromatin configuration at meiotic DNA double-stranded break sites in premeiotic and meiotic cells.|
Ohta, K, et al.
Proc. Natl. Acad. Sci. U.S.A., 95: 646-51 (1998) 1998
In the yeast Saccharomyces cerevisiae, meiotic recombination is initiated by DNA double-stranded breaks (DSBs) occurring in micrococcal nuclease (MNase)-hypersensitive regions of the chromatin. MNase-sensitive sites also undergo meiosis-specific alterations in chromatin structure prior to the appearance of DSBs. DSB formation requires the products of numerous genes. Herein we have examined the effects of mutations in four such genes, MRE11, RAD50, XRS2, and MRE2, on MNase sensitivity at DSB sites in premeiotic and meiotic cells. Disruption mutations in each of four genes confer greater than wild-type levels of MNase sensitivity in premeiotic cells. In meiotic prophase, all of these mutations affect MNase sensitivity at DSB sites and fall into two distinct phenotypic classes. The type 1 mutations (mre2 and mre11) confer a reduction in MNase sensitivity relative to the wild-type level. The type 2 mutations (rad50 and xrs2) permit a meiotic increase in the MNase sensitivity to reach a final level higher than that observed in wild-type cells. An mre11 disruption mutation (type 1) is epistatic to a rad50 null mutation (type 2) with respect to its meiotic effects on MNase sensitivity, suggesting that the events observed in the type 2 mutants during meiosis are dependent upon type 1 functions. One interpretation of these results is that Mre11, Rad50, Xrs2, and possibly Mer2 (whose splicing is Mre2-dependent) form a complex at recombination hot spots and establish a chromatin/DNA configuration favorable for the induction of DSBs.
|Nuclease activities in a complex of human recombination and DNA repair factors Rad50, Mre11, and p95.|
Trujillo, K M, et al.
J. Biol. Chem., 273: 21447-50 (1998) 1998
Genetic studies in yeast have indicated a role of the RAD50 and MRE11 genes in homologous recombination, telomere length maintenance, and DNA repair processes. Here, we purify from nuclear extract of Raji cells a complex consisting of human Rad50, Mre11, and another protein factor with a size of about 95 kDa (p95), which is likely to be Nibrin, the protein encoded by the gene mutated in Nijmegen breakage syndrome. We show that the Rad50-Mre11-p95 complex possesses manganese-dependent single-stranded DNA endonuclease and 3' to 5' exonuclease activities. These nuclease activities are likely to be important for recombination, repair, and genomic stability.
|Human Rad50 is physically associated with human Mre11: identification of a conserved multiprotein complex implicated in recombinational DNA repair.|
Dolganov, G M, et al.
Mol. Cell. Biol., 16: 4832-41 (1996) 1996
In this report, we describe the identification and molecular characterization of a human RAD50 homolog, hRAD50. hRAD50 was included in a collection of cDNAs which were isolated by a direct cDNA selection strategy focused on the chromosomal interval spanning 5q23 to 5q31. Alterations of the 5q23-q31 interval are frequently observed in myelodysplasia and myeloid leukemia. This strategy was thus undertaken to create a detailed genetic map of that region. Saccharomyces cerevisiae RAD50 (ScRAD50) is one of three yeast RAD52 epistasis group members (ScRAD50, ScMRE11, and ScXRS2) in which mutations eliminate meiotic recombination but confer a hyperrecombinational phenotype in mitotic cells. The yeast Rad50, Mre11, and Xrs2 proteins appear to act in a multiprotein complex, consistent with the observation that the corresponding mutants confer essentially identical phenotypes. In this report, we demonstrate that the human Rad50 and Mre11 proteins are stably associated in a protein complex which may include three other proteins. hRAD50 is expressed in all tissues examined, but mRNA levels are significantly higher in the testis. Other human RAD52 epistasis group homologs exhibit this expression pattern, suggesting the involvement of human RAD52 epistasis group proteins in meiotic recombination. Human RAD52 epistasis group proteins are highly conserved and act in protein complexes that are analogous to those of their yeast counterparts. These findings indicate that the function of the RAD52 epistasis group is conserved in human cells.