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Results for “Structural Maintenance of Chromosome Protein 1”

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At least 19 recordsLinked to original sources

PubMed2005

Post-irradiation phosphorylation of structural maintenance chromosome 1 (SMC1) is independent of the Fanconi protein pathway.

PURPOSE: To confirm the sensitivity of cells from patients with Fanconi anemia (FA) to ionizing radiation, and to determine whether the phosphorylation of structural maintenance chromosome 1 (SMC1) was associated with radiosensitivity, as it is in other DNA repair disorders. METHODS AND MATERIALS: Using lymphoblastoid cell lines from FA patients before and after exposure to ionizing radiation, the colony survival assay, radioresistant DNA synthesis, and SMC1 phosphorylation were measured. FA lymphoblastoid cell lines that had been transfected with the wild-type FANC gene were used as controls. RESULTS: Cells from FA patients of six complementation groups were radiosensitive. Despite this, SMC1 phosphorylation was normal in each case; radioresistant DNA synthesis, a measure of S phase checkpoint integrity, was defective in FANCD2 lymphoblastoid cell lines and was corrected in FANCD2 + D2 cells. CONCLUSIONS: The data indicate that the FANC pathway proteins play a major role in the cellular responses to ionizing radiation, but not in SMC1 phosphorylation or in the S phase checkpoint of FANCD2-deficient cells. Thus, SMC1 activation is not a common denominator of radiosensitivity, as has been suggested by radiation responses of cells from ataxia-telangiectasia, Nijmegen breakage syndrome, or Mre11 deficiency patients.

Cell Cycle Proteins↗

PubMed2006

Cohesin protein SMC1 represses the nuclear receptor CAR-mediated synergistic activation of a human P450 gene by xenobiotics.

CAR (constitutive active/androstane receptor) regulates both the distal enhancer PBREM (phenobarbital-responsive enhancer module) and the proximal element OARE [OA (okadaic acid) response element] to synergistically up-regulate the endogenous CYP2B6 (where CYP is cytochrome P450) gene in HepG2 cells. In this up-regulation, CAR acts as both a transcription factor and a co-regulator, directly binding to and enhancing PBREM upon activation by xenobiotics such as TCPOBOP {1,4-bis-[2-(3,5-dichloropyridyloxy)]benzene} and indirectly associating with the OARE in response to OA [Swales, Kakizaki, Yamamoto, Inoue, Kobayashi and Negishi (2005) J. Biol. Chem. 280, 3458-3466]. We have now identified the cohesin protein SMC1 (structural maintenance of chromosomes 1) as a CAR-binding protein and characterized it as a negative regulator of OARE activity, thus repressing synergy. Treatment with SMC1 small interfering RNA augmented the synergistic up-regulation of CYP2B6 expression 20-fold in HepG2 cells, while transient co-expression of spliced form of SMC1 abrogated the synergistic activation of a 1.8 kb CYP2B6 promoter. SMC1 indirectly binds to a 19 bp sequence (-236/-217) immediately downstream from the OARE in the CYP2B6 promoter. Both DNA affinity and chromatin immunoprecipitation assays showed that OA treatment dissociates SMC1 from the CYP2B6 promoter, reciprocating the indirect binding of CAR to OARE. These results are consistent with the conclusion that SMC1 binding represses OARE activity and its dissociation allows the recruitment of CAR to the OARE, synergizing PBREM activity and the expression of the CYP2B6 gene.

Animals↗

PubMed2005

RPGR-ORF15, which is mutated in retinitis pigmentosa, associates with SMC1, SMC3, and microtubule transport proteins.

Mutations in the retinitis pigmentosa GTPase regulator (RPGR) gene account for almost 20% of patients with retinitis pigmentosa. Most mutations are detected in alternatively spliced RPGR-ORF15 isoform(s), which are primarily but not exclusively expressed in the retina. We show that, in addition to the axoneme, the RPGR-ORF15 protein is localized to the basal bodies of photoreceptor connecting cilium and to the tip and axoneme of sperm flagella. Mass spectrometric analysis of proteins that were immunoprecipitated from the retinal axoneme-enriched fraction using an anti-ORF15 antibody identified two chromosome-associated proteins, structural maintenance of chromosomes (SMC) 1 and SMC3. Using pulldown assays, we demonstrate that the interaction of RPGR with SMC1 and SMC3 is mediated, at least in part, by the RCC1-like domain of RPGR. This interaction was not observed with phosphorylation-deficient mutants of SMC1. Both SMC1 and SMC3 localized to the cilia of retinal photoreceptors and Madin-Darby canine kidney cells, suggesting a broader physiological relevance of this interaction. Additional immunoprecipitation studies revealed the association of RPGR-ORF15 isoform(s) with the intraflagellar transport polypeptide IFT88 as well as microtubule motor proteins, including KIF3A, p150Glued, and p50-dynamitin. Inhibition of dynein function by overexpressing p50 abrogated the localization of RPGR-ORF15 to basal bodies. Taken together, these results provide novel evidence for the possible involvement of RPGR-ORF15 in microtubule organization and regulation of transport in primary cilia.

Animals↗

PubMed2004

The ATM-SMC1 pathway is essential for activation of the chromium[VI]-induced S-phase checkpoint.

Hexavalent chromium (Cr[VI]) is a common industrial waste product, an environmental pollutant, and a recognized human carcinogen. Following cellular uptake, Cr[VI] can cause DNA damage, however, the mechanisms by which mammalian cells respond to Cr-induced DNA damage remain to be elucidated. Using single cell gel electrophoresis (e.g., Comet Assay) and immunofluoresence microscopy to detect the presence of gamma-H2AX foci, we find that Cr[VI] induces DNA double-strand breaks similar to ionizing radiation (IR). We also demonstrated that ataxia telangiectasia mutated (ATM) is activated in response to Cr[VI] and exposure to Cr[VI] triggers a dose and ATM-dependent S-phase arrest. Further, we document that ATM is required for phosphorylation of the structural maintenance of chromosome protein 1 (SMC1). Finally, we find that ATM-dependent phosphorylation of SMC1 is required to facilitate S-phase cell-cycle arrest in response to Cr[VI] exposure. Collectively, these results indicate that the ATM-SMC1 pathway plays a critical role in cellular response to Cr[VI].

Ataxia Telangiectasia Mutated Proteins↗

PubMed2004

Chromatin association of rad17 is required for an ataxia telangiectasia and rad-related kinase-mediated S-phase checkpoint in response to low-dose ultraviolet radiation.

Activation of the S-phase checkpoint results in an inhibition of DNA synthesis in response to DNA damage. This is an active cellular response that may enhance cell survival and limit heritable genetic abnormalities. While much attention has been paid to elucidating signal transduction pathways regulating the ionizing radiation-induced S-phase checkpoint, less is known about whether UV radiation initiates the process and the mechanism controlling it. Here, we demonstrate that low-dose UV radiation activates an S-phase checkpoint that requires the ataxia telangiectasia and Rad-related kinase (ATR). ATR regulates the S-phase checkpoint through phosphorylation of the downstream target structural maintenance of chromosomal protein 1. Furthermore, the ATPase activity of Rad17 is crucial for its chromatin association and for the functional effects of ATR activation in response to low-dose UV radiation. These results suggest that low-dose UV radiation activates an S-phase checkpoint requiring ATR-mediated signal transduction pathway.

Adenosine Triphosphatases↗

PubMed2006

ATR regulates hexavalent chromium-induced S-phase checkpoint through phosphorylation of SMC1.

Hexavalent chromium (Cr[VI]) is an industrial waste product known to cause nasal and lung cancer in exposed workers. Intracellularly, Cr[VI] undergoes a series of enzymatic reductions resulting in the formation of reactive chromate intermediates and oxygen free radicals. These metabolites react with DNA to cause numerous types of genomic lesions, but the cellular response to these genotoxic insults is poorly understood. Recently, we demonstrated that in response to DNA damage induced by Cr[VI], an ataxia-telangiectasia mutated (ATM) and structural maintenance of chromosomal protein 1 (SMC1)-dependent S-phase checkpoint is activated. Interestingly, this checkpoint response was only ATM-dependent in cells exposed to low doses of Cr[VI], we demonstrate that the ATM and Rad3 related kinase, ATR, is required to activate the S-phase checkpoint. In response to all doses of Cr[VI], ATR is activated and phosphorylates SMC1 to facilitate the checkpoint. Further, chromatin binding ability of Rad17 is required for this process. Taken together, these results indicate that the Rad17-ATR-SMC1 pathway is essential for Cr[VI]-induced S-phase checkpoint activation.

Ataxia Telangiectasia Mutated Proteins↗

PubMed2002

The coiled coil region (amino acids 129-250) of the tumor suppressor protein adenomatous polyposis coli (APC). Its structure and its interaction with chromosome maintenance region 1 (Crm-1).

The APC (adenomatous polyposis coli) tumor suppressor protein has many different intracellular functions including a nuclear export activity. Only little is known about the molecular architecture of the 2843-amino acid APC protein. Guided by secondary structure predictions we identified a fragment close to the N-terminal end, termed APC-(129-250), as a soluble and protease-resistant domain. We solved the crystal structure of APC-(129-250), which is monomeric and consists of three alpha-helices forming two separate antiparallel coiled coils. APC-(129-250) includes the nuclear export signal NES-(165-174) at the C-terminal end of the first helix. Surprisingly, the conserved hydrophobic amino acids of NES-(165-174) are buried in one of the coiled coils and are thus not accessible for interaction with other proteins. We demonstrate the direct interaction of APC-(129-250) with the nuclear export factor chromosome maintenance region 1 (Crm-1). This interaction is enhanced by the small GTPase Ran in its activated GTP-bound form and also by a double mutation in APC-(129-250), which deletes two amino acids forming two of the major interhelical interactions within the coiled coil. These observations hint to a regulatory mechanism of the APC nuclear export activity by NES masking.

Adenomatous Polyposis Coli Protein↗

PubMed2006

Direct interaction between cohesin complex and DNA replication machinery.

Structural maintenance of chromosome 1 (Smc1) is a multifunctional protein, which has been implicated in sister chromatid cohesion, DNA recombination and repair, and the activation of cell cycle checkpoints by ionizing radiation, ultraviolet light, and other genotoxic agents. In order to identify the proteins that interact with Smc1, we conducted the Tandem affinity purification (TAP) technique and analyzed the Smc1-interacting proteins via MALDI-TOF mass spectrometry. We identified minichromosome maintenance 7 (Mcm7), an essential component of the pre-replication complex, as a novel Smc1-interacting protein. Co-immunoprecipitation revealed an interaction occurring between Smc1 and Mcm7, both in vitro and in vivo. Using a GST pull-down assay, we determined that Smc1 interacts physically with Mcm7 via its N-terminal and hinge regions, and Mcm7 interacts with Smc1 via its middle region. Interestingly, we also discovered that Smc1 interacts with other DNA replication proteins, including Mcm6, RFC1, and DNA polymerase alpha. These results suggest that a functional link exists between the cohesin complex and DNA replication proteins.

Cell Cycle Proteins↗

PubMed2004

Search for epithelial-specific mRNAs in peripheral blood of patients with colon cancer by RT-PCR.

Research has widely supported the efficacy of screening for colorectal cancer in reducing mortality. A blood-based assay potentially represents a more accessible early detection tool for the identification of solid tumor cells originating from a primary tumor site in the body. We demonstrate a relatively easy and highly reproducible technique for the detection of mRNA expression of genes as markers of malignancy in blood samples of patients with colon cancer. The present study aims to identify a set of specific mRNAs expressed in epithelial cells but not in blood cells, which may be useful as markers for early detection of circulating colon cancer cells by a simple, qualitative RT-PCR assay following semi-automated RNA extraction from peripheral blood samples. Our approach includes a systematic search for candidate markers using digital differential display, search on UniGene colon EST libraries and analysis of published data on colon cancer gene expression. A final list included the following genes: bone morphogenetic protein 4 (BMP4), cyclin D (CycD), family with sequence similarity 3, member D (FAM3D), gastrin (GAS), glycoprotein A33 transmembrane (GPA33), glutathione peroxidase 2 gastrointestinal (GPX2), galactoside-binding, soluble, 4 (galectin 4) (LGALS4), non-SMC, structural maintenance of chromosomes, element 1 protein (NSE1), tumor-associated calcium signal transducer 1 (TACSTD1), telomerase reverse transcriptase (hTERT), trefoil factor 3 intestinal (TFF3), transmembrane 4 superfamily member 3 (TM4SF3), UDP glycosyltransferase 1 family, polypeptide A9 (UGT1A9), villin 1 (VIL1), and the novel gene FLJ20127. The mRNA expression of these genes was evaluated in a pool of 16 samples from subjects diagnosed with colon cancer and from 16 normal-controls. We observed expression in 13 of the 15 investigated genes from the blood samples of the vast majority of patients considered, but also in a certain percentage of the controls (from 14.3 to 100%). This finding confirms that the extreme sensitivity of RT-PCR is able to detect minimal amounts of mRNA expressed in a non tissue-specific manner ('illegitimate transcription'). On the contrary, NSE1 and GAS mRNAs were not detected either in patient or in control blood samples; however, they were abundantly expressed in normal and cancerous colon mucosa, encouraging further search for useful markers able to detect epithelial cells in peripheral blood.

Aged↗

PubMed1996

SMC proteins constitute two subunits of the mammalian recombination complex RC-1.

Recombination protein complex RC-1, purified from calf thymus nuclear extracts, catalyzes cell-free DNA strand transfer and repair of gaps and deletions through DNA recombination. DNA polymerase E, DNA ligase III and a DNA structure-specific endonuclease co-purify with the five polypeptide complex. Here we describe the identification of two hitherto unknown subunits of RC-1. N-terminal amino acid sequences of the 160 and 130 kDa polypeptides display up to 100% identity to proteins of the structural maintenance of chromosomes (SMC) subfamilies 1 and 2. SMC proteins are involved in mitotic chromosome segregation and condensation, as well as in certain DNA repair pathways in fission (rad18 gene) and budding (RHC18 gene) yeast. The assignment was substantiated by immuno-cross-reactivity of the RC-1 subunits with polyclonal antibodies specific for Xenopus laevis SMC proteins. These antibodies, and polyclonal antibodies directed against the bovine 160 and 130 kDa polypeptides, named BSMC1 and BSMC2 (bovine SMC), inhibited RC-1-mediated DNA transfer, indicating that the SMC proteins are necessary components of the reaction. Two independent assays revealed DNA reannealing activity of RC-1, which resides in its BSMC subunits, thereby demonstrating a novel function of these proteins. To our knowledge, this is the first evidence for the association of mammalian SMC proteins with a multiprotein complex harboring, among others, DNA recombination, DNA ligase and DNA polymerase activities.

Adenosine Triphosphatases↗

PubMed2006

Spatial organization of the mammalian genome surveillance machinery in response to DNA strand breaks.

We show that DNA double-strand breaks (DSBs) induce complex subcompartmentalization of genome surveillance regulators. Chromatin marked by gamma-H2AX is occupied by ataxia telangiectasia-mutated (ATM) kinase, Mdc1, and 53BP1. In contrast, repair factors (Rad51, Rad52, BRCA2, and FANCD2), ATM and Rad-3-related (ATR) cascade (ATR, ATR interacting protein, and replication protein A), and the DNA clamp (Rad17 and -9) accumulate in subchromatin microcompartments delineated by single-stranded DNA (ssDNA). BRCA1 and the Mre11-Rad50-Nbs1 complex interact with both of these compartments. Importantly, some core DSB regulators do not form cytologically discernible foci. These are further subclassified to proteins that connect DSBs with the rest of the nucleus (Chk1 and -2), that assemble at unprocessed DSBs (DNA-PK/Ku70), and that exist on chromatin as preassembled complexes but become locally modified after DNA damage (Smc1/Smc3). Finally, checkpoint effectors such as p53 and Cdc25A do not accumulate at DSBs at all. We propose that subclassification of DSB regulators according to their residence sites provides a useful framework for understanding their involvement in diverse processes of genome surveillance.

Animals↗

PubMed2005

Human Rad9 is required for the activation of S-phase checkpoint and the maintenance of chromosomal stability.

In response to DNA damage or replication block, cells activate a battery of checkpoint signaling cascades to control cell cycle progression and elicit DNA repair in order to maintain genomic stability and integrity. Identified as a homolog of its fission yeast counterpart, human Rad9 was proposed to form a Rad9-Hus1-Rad1 protein complex to mediate checkpoint signals. However, the precise function of Rad9 in the process of checkpoint activation is not fully understood. Using the RNA interference technique, we investigated the role of Rad9 in the genotoxic stress-induced activation of S-phase checkpoint and the maintenance of chromosomal stability. We found that Rad9 knockdown reduced the phosphorylation of Rad17, Chk1 and Smc1 in response to DNA replication block and certain types of DNA damage. Immunofluorescence studies showed that the removal of Rad9 disrupted the foci formation of phosphorylated Chk1, but not ATR. Moreover, Rad9 knockdown resulted in radioresistant DNA synthesis and reduced cell viability under replication stress. Finally, removal of Rad9 by RNAi led to increased accumulation of spontaneous chromosomal aberrations. Taken together, these results suggest a critical and specific role of Rad9 in the activation of S-phase checkpoint and the maintenance of chromosome stability.

Ataxia Telangiectasia Mutated Proteins↗

PubMed1998

Structural maintenance of chromosomes protein C-terminal domains bind preferentially to DNA with secondary structure.

Structural maintenance of chromosomes (SMC) proteins interact with DNA in chromosome condensation, sister chromatid cohesion, DNA recombination, and gene dosage compensation. How individual SMC proteins and their functional domains bind DNA has not been described. We demonstrate the ability of the C-terminal domains of Saccharomyces cerevisiae SMC1 and SMC2 proteins, representing two major subfamilies with different functions, to bind DNA in an ATP-independent manner. Three levels of DNA binding specificity were observed: 1) a >100-fold preference for double-stranded versus single-stranded DNA; 2) a high affinity for DNA fragments able to form secondary structures and for synthetic cruciform DNA molecules; and 3) a strong preference for AT-rich DNA fragments of particular types. These include fragments from the scaffold-associated regions, and an alternating poly(dA-dT)-poly(dT-dA) synthetic polymer, as opposed to a variety of other polymers. Reannealing of complementary DNA strands is also promoted primarily by the C-terminal domains. Consistent with their in vitro DNA binding activity, we show that overexpression of the SMC C termini increases plasmid loss without altering viability or cell cycle progression.

Base Composition↗

PubMed2005

A carboxyl leucine-rich region of parathyroid hormone-related protein is critical for nuclear export.

PTHrP is an oncofetal protein with distinct proliferative and antiapoptotic roles that are affected by nucleocytoplasmic shuttling. The protein's nuclear export is sensitive to leptomycin B, consistent with a chromosome region maintenance protein 1-dependent pathway. We determined that the 109-139 region of PTHrP was involved in its nuclear export by demonstrating that a C-terminal truncation mutant, residues 1-108, exports at a reduced rate, compared with the wild-type 139 amino acid isoform. We searched for potential nuclear export sequences within the 109-139 region, which is leucine rich. Comparisons with established nuclear export sequences identified a putative consensus signal at residues 126-136. Deletion of this region resulted in nuclear export characteristics that closely matched those of the C-terminal truncation mutant. Confocal microscopic analyses of transfected 293, COS-1, and HeLa cells showed that steady-state nuclear levels of the truncated and deletion mutants were significantly greater than levels of wild-type PTHrP and were unaffected by leptomycin B, unlike the wild-type protein. In addition, both mutants demonstrated greatly reduced nuclear export with assays using nuclear preparations and intact cells. Based on these results, we conclude that the 126-136 amino acid sequence closely approximates the structure of a chromosome region maintenance protein 1-dependent leucine-rich nuclear export signal and is critical for nuclear export of PTHrP.

Active Transport, Cell Nucleus↗

PubMed2004

ATM-dependent CHK2 activation induced by anticancer agent, irofulven.

Irofulven (6-hydroxymethylacylfulvene, HMAF, MGI 114) is one of a new class of anticancer agents that are semisynthetic derivatives of the mushroom toxin illudin S. Preclinical studies and clinical trials have demonstrated that irofulven is effective against several tumor types. Mechanisms of action studies indicate that irofulven induces DNA damage, MAPK activation, and apoptosis. In this study we found that in ovarian cancer cells, CHK2 kinase is activated by irofulven while CHK1 kinase is not activated even when treated at higher concentrations of the drug. By using GM00847 human fibroblast expressing tetracycline-controlled, FLAG-tagged kinase-dead ATR (ATR.kd), it was demonstrated that ATR kinase does not play a major role in irofulven-induced CHK2 activation. Results from human fibroblasts proficient or deficient in ATM function (GM00637 and GM05849) indicated that CHK2 activation by irofulven is mediated by the upstream ATM kinase. Phosphorylation of ATM on Ser(1981), which is critical for kinase activation, was observed in ovarian cancer cell lines treated with irofulven. RNA interference results confirmed that CHK2 activation was inhibited after introducing siRNA for ATM. Finally, experiments done with human colon cancer cell line HCT116 and its isogenic CHK2 knockout derivative; and experiments done by expressing kinase-dead CHK2 in an ovarian cancer cell line demonstrated that CHK2 activation contributes to irofulven-induced S phase arrest. In addition, it was shown that NBS1, SMC1, and p53 were phosphorylated in an ATM-dependent manner, and p53 phosphorylation on serine 20 is dependent on CHK2 after irofulven treatment. In summary, we found that the anticancer agent, irofulven, activates the ATM-CHK2 DNA damage-signaling pathway, and CHK2 activation contributes to S phase cell cycle arrest induced by irofulven.

Antineoplastic Agents, Alkylating↗

PubMed2007

Incidence and clinical features of X-linked Cornelia de Lange syndrome due to SMC1L1 mutations.

Cornelia de Lange syndrome (CdLS) is a multisystem developmental disorder characterized by facial dysmorphism, growth and mental retardation, microcephaly, and various malformations. Heterozygous mutations in the NIPBL gene have been detected in approximately 45% of affected individuals. Recently, a second CdLS gene, mapping to the X chromosome, has been identified: SMC1L1 (structural maintenance of chromosomes 1-like 1; or SMC1A). In order to estimate the incidence and refine the clinical presentation of X-linked CdLS, we have screened a series of 11 CdLS boys carrying no NIPBL anomaly. We have identified two novel de novo SMC1L1 missense mutations (c.587G>A [p.Arg196His] and c.3254A>G [p.Tyr1085Cys]). Our results confirm that SMC1L1 mutations cause CdLS and support the view that SMC1L1 accounts for a significant fraction of boys with unexplained CdLS. Furthermore, we suggest that SMC1L1 mutations have milder effects than NIPBL mutations with respect to pre- and postnatal growth retardation and associated malformations. If confirmed, these data may have important implications for directing mutation screening in CdLS.

Adolescent↗

PubMed2005

Inhibition of human Chk1 causes increased initiation of DNA replication, phosphorylation of ATR targets, and DNA breakage.

Human checkpoint kinase 1 (Chk1) is an essential kinase required to preserve genome stability. Here, we show that Chk1 inhibition by two distinct drugs, UCN-01 and CEP-3891, or by Chk1 small interfering RNA (siRNA) leads to phosphorylation of ATR targets. Chk1-inhibition triggered rapid, pan-nuclear phosphorylation of histone H2AX, p53, Smc1, replication protein A, and Chk1 itself in human S-phase cells. These phosphorylations were inhibited by ATR siRNA and caffeine, but they occurred independently of ATM. Chk1 inhibition also caused an increased initiation of DNA replication, which was accompanied by increased amounts of nonextractable RPA protein, formation of single-stranded DNA, and induction of DNA strand breaks. Moreover, these responses were prevented by siRNA-mediated downregulation of Cdk2 or the replication initiation protein Cdc45, or by addition of the CDK inhibitor roscovitine. We propose that Chk1 is required during normal S phase to avoid aberrantly increased initiation of DNA replication, thereby protecting against DNA breakage. These results may help explain why Chk1 is an essential kinase and should be taken into account when drugs to inhibit this kinase are considered for use in cancer treatment.

Ataxia Telangiectasia Mutated Proteins↗

PubMed2005

Rad50 depletion impacts upon ATR-dependent DNA damage responses.

The Mre11/Rad50/NBS1 (MRN) complex is mutated in inherited genomic instability syndromes featuring cancer predisposition, mental retardation and immunodeficiency. It functions both in DNA double-strand break repair and in controlling the ataxia telangiectasia mutated (ATM) kinase during the response to these lesions. Patients inheriting homozygosity for an NBS1 hypomorphic allele display reduced phosphorylation of signaling factors such as Chk1, but not of chromatin-associated factor H2AX, after stresses that activate the ATM-related kinase, ATR. Therefore, we tested whether MRN has a global controlling role over the ATR kinase through the study of MRN deficiencies generated via RNA interference. We show for the first time that MRN is required for ATR-dependent phosphorylation of structural maintenance of chromosomes 1 (Smc1), which acts within chromatin to ensure sister chromatid cohesion and to effect several DNA damage responses. We have uncovered novel phenotypes caused by MRN deficiency that support a functional link between this complex, ATR and Smc1, including hypersensitivity to UV exposure, a defective UV responsive intra-S phase checkpoint and a specific pattern of genomic instability. In addition, certain ATR-dependent responses do not require MRN. These studies demonstrate that there is indeed a controlling role for MRN over the ATR kinase and have established that the downstream events under this control are broad, including both chromatin-associated and diffuse signaling factors, but may not be universal. These studies contribute to our understanding of the central role that MRN plays in damage detection and signaling, which serve to maintain genomic stability and resist neoplastic transformation.

Acid Anhydride Hydrolases↗