Cloning and characterization of mammalian SMC1 and SMC3 genes and proteins, components of the DNA recombination complexes RC-1

Stursberg, S.; Riwar, Brigitte; Jessberger, Rolf

doi:10.1016/s0378-1119(99)00021-9

Cited by 41 publications

(40 citation statements)

References 36 publications

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“…The functional ramifications of Smc1 phosphorylation in this pathway have not been elucidated. Another intriguing potential link of Smc1 function to radiosensitivity relates to its apparent role in DNA recombination via its presence in the RC-1 complex (Jessberger et al 1996;Stursberg et al 1999). Whether phosphorylation of Smc1 by Atm affects its function in recombination events needs to be explored.…”

Section: Discussionmentioning

confidence: 99%

“…The proteins appear to function as heterodimers with the Smc1-Smc3 heterodimer implicated in sister chromatid cohesion (also known as the cohesin complex) and the Smc2-Smc4 heterodimer implicated in chromosome condensation (also known as the condensin complex; Hirano 1999). The Smc1-Smc3 heterodimer has also been found in a mammalian protein complex, called RC-1, that promotes DNA recombination (Jessberger et al 1996;Stursberg et al 1999). The RC-1 complex, which appears to constitute only a minor fraction of the total Smc1-Smc3 complexes in normal cells (Strunnikov and Jessberger 1999), promotes repair of gaps and deletions through recombination between otherwise homologous DNA molecules (Jessberger et al 1993).…”

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confidence: 99%

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Involvement of the cohesin protein, Smc1, in Atm-dependent and independent responses to DNA damage

Kim

Xu²,

Kastan³

2002

Genes Dev.

457

358

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Structural maintenance of chromosomes (SMC) proteins play important roles in sister chromatid cohesion, chromosome condensation, sex-chromosome dosage compensation, and DNA recombination and repair. Protein complexes containing heterodimers of the Smc1 and Smc3 proteins have been implicated specifically in both sister chromatid cohesion and DNA recombination. Here, we show that the protein kinase, Atm, which belongs to a family of phosphatidylinositol 3-kinases that regulate cell cycle checkpoints and DNA recombination and repair, phosphorylates Smc1 protein after ionizing irradiation. Atm phosphorylates Smc1 on serines 957 and 966 in vitro and in vivo, and expression of an Smc1 protein mutated at these phosphorylation sites abrogates the ionizing irradiation-induced S phase cell cycle checkpoint. Optimal phosphorylation of these sites in Smc1 after ionizing irradiation also requires the presence of the Atm substrates Nbs1 and Brca1. These same sites in Smc1 are phosphorylated after treatment with UV irradiation or hydroxyurea in an Atm-independent manner, thus demonstrating that another kinase must be involved in responses to these cellular stresses. Yeast containing hypomorphic mutations in SMC1 and human cells overexpressing Smc1 mutated at both of these phosphorylation sites exhibit decreased survival following ionizing irradiation. These results demonstrate that Smc1 participates in cellular responses to DNA damage and link Smc1 to the Atm signal transduction pathway. Signal transduction pathways activated by DNA damage and other cellular stresses are critical determinants of cell survival and cellular transformation. The product of the ATM gene, which is mutated in the cancer-prone disorder, Ataxia-telangiectasia (A-T), appears to be a major regulator of cellular responses to ionizing irradiation . Cells from A-T patients exhibit a variety of abnormalities, including genetic instability, radiosensitivity, cell cycle checkpoint defects in the G 1 , S, and G 2 phases of the cell cycle, and poor growth in culture. Atm is a protein kinase that is activated by ionizing irradiation (Banin et al. 1998;Canman et al. 1998) and that phosphorylates a number of different substrates following activation, including p53, Mdm2, Chk2, Nbs1, Brca1, and Rad17 (Kastan and Lim 2000; Zhou and Elledge 2000; Shiloh and Kastan 2001). The functional consequences of some of these phosphorylation events have been elucidated. For example, Atm phosphorylation of p53 (Banin et al. 1998;Canman et al. 1998), Chk2 (Matsuoka et al. 1998Ahn et al. 2000;Melchionna et al. 2000;, and Mdm2 (Maya et al. 2001) appears to contribute to the arrest of cells in G 1 following ionizing irradiation whereas phosphorylation of Nbs1, Brca1, and Rad17 is required for transient S phase (Nbs1 and Brca1) or G 2 (Brca1 and Rad17) arrests after ionizing irradiation Zhao et al. 2000;Bao et al. 2001;Xu et al. 2001). However, not all of the phenotypic abnormalities in A-T patients and cells from A-T patients can be explained by these known phosphorylation events, t...

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Involvement of the cohesin protein, Smc1, in Atm-dependent and independent responses to DNA damage

Kim

Xu²,

Kastan³

2002

Genes Dev.

457

358

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“…There is also a possibility that the mSMCB-mSMCD-PW29-based complex is involved in a process other than sister chromatid cohesion since no functional studies of its role in mammals have been carried out. There is, however, an indication that the SMC components of this complex are shared with the RC1 complex, involved in genetic recombination (Stursberg et al, 1999).…”

Section: Discussionmentioning

confidence: 99%

“…This complex is termed the 'cohesin' complex. Interestingly, the mammalian multiprotein recombination complex RC-1 is able to catalyze renaturation of single-stranded DNA, and includes bovine homologs of XSMC1 and XSMC3 (Jessberger et al, 1996;Stursberg et al, 1999). Homologs of SMC1 and SMC3, hSMC1 and hSMC3, have also been identified in humans (Schmiesing et al, 1998).…”

Section: Introductionmentioning

confidence: 99%

Characterization of the components of the putative mammalian sister chromatid cohesion complex

Darwiche

Freeman

Strunnikov

1999

Gene

109

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Establishing and maintaining proper sister chromatid cohesion throughout the cell cycle are essential for maintaining genome integrity. To understand how sister chromatid cohesion occurs in mammals, we have cloned and characterized mouse orthologs of proteins known to be involved in sister chromatid cohesion in other organisms. The cDNAs for the mouse orthologs of SMC1 S.c. and SMC3 S.c. , mSMCB and mSMCD respectively, were cloned and the corresponding transcripts and proteins were characterized. mSMCB and mSMCD are transcribed at similar levels in adult mouse tissues except in testis, which has an excess of mSMCD transcripts. The mSMCB and mSMCD proteins, as well as the PW29 protein, a mouse homolog of Mcd1p S.c./ Rad21 S.p. , form a complex similar to cohesin in X. laevis. mSMCB, mSMCD and PW29 protein levels show no significant cellcycle dependence. The bulk of the mSMCB, mSMCD and PW29 proteins undergo redistribution from the chromosome vicinity to the cytoplasm during prometaphase and back to the chromatin in telophase. This pattern of intracellular localization suggests a complex role for this group of SMC proteins in chromosome dynamics. The PW29 protein and PCNA, which have both been implicated in sister chromatid cohesion, do not colocalize, indicating that these proteins may not function in the same cohesion pathway. Overexpression of a PW29-GFP fusion protein in mouse fibroblasts leads to inhibition of proliferation, implicating this protein and its complex with SMC proteins in the control of mitotic cycle progression.

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“…Protein Domains-Cloning of the full-length cDNAs for bSMC1 and bSMC3 was described before (24). For expression of individual SMC protein domains, the respective cDNA fragments were cloned into the appropriate vectors of the pQE30 -32 series (Qiagen, Inc.), placing a His 6 tag at the N terminus of the proteins.…”

Section: Cloning Overexpression and Purification Of Bsmc1 And Bsmc3mentioning

confidence: 99%

Mammalian SMC3 C-terminal and Coiled-coil Protein Domains Specifically Bind Palindromic DNA, Do Not Block DNA Ends, and Prevent DNA Bending

Akhmedov¹,

Gross²,

Jessberger³

1999

Journal of Biological Chemistry

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The C-terminal domains of yeast structural maintenance of chromosomes (SMC) proteins were previously shown to bind double-stranded DNA, which generated the idea of the antiparallel SMC heterodimer, such as the SMC1/3 dimer, bridging two DNA molecules. Analysis of bovine SMC1 and SMC3 protein domains now reveals that not only the C-terminal domains, but also the coiled-coil region, binds DNA, while the N terminus is inactive. Duplex DNA and DNA molecules with secondary structures are highly preferred substrates for both the C-terminal and coiled-coil domains. Contrasting other cruciform DNA-binding proteins like HMG1, the SMC3 C-terminal and coiled-coil domains do not bend DNA, but rather prevent bending in ring closure assays. Phosphatase, exonuclease, and ligase assays showed that neither domain renders DNA ends inaccessible for other enzymes. These observations allow modifications of models for SMC-DNA interactions.

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Cloning and characterization of mammalian SMC1 and SMC3 genes and proteins, components of the DNA recombination complexes RC-1

Cited by 41 publications

References 36 publications

Involvement of the cohesin protein, Smc1, in Atm-dependent and independent responses to DNA damage

Involvement of the cohesin protein, Smc1, in Atm-dependent and independent responses to DNA damage

Characterization of the components of the putative mammalian sister chromatid cohesion complex

Mammalian SMC3 C-terminal and Coiled-coil Protein Domains Specifically Bind Palindromic DNA, Do Not Block DNA Ends, and Prevent DNA Bending

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