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PubMed2001

Frequent allelic imbalance at the ATM locus in DNA multiploid colorectal carcinomas.

DNA multiploidy may involve specific DNA ploidy states with respect to genetic alterations such as oncogenes, tumor suppressor gene mutation and microsatellite instability. To clarify the role of DNA multiploidy in colorectal cancer, we analysed allelic imbalance involving the ATM gene, localized to chromosome 11q22-23 and thought to be involved in genetic stability, in a series of multiploid colorectal carcinomas. In addition, p53 gene mutation (exons 5-8) and allelic imbalance at 11q24 loci distal to the ATM locus were also examined. The crypt isolation technique coupled with DNA cytometric sorting and polymerase chain reaction assay using 10 microsatellite markers tightly linked to the ATM gene were used to study ATM allelic imbalance in 55 colorectal carcinomas (15 diploid, 13 aneuploid, 27 multiploid). While allelic imbalance at the ATM locus was rarely observed in diploid and aneuploid carcinomas, multiploid carcinomas exhibited a high frequency of ATM allelic imbalance. In multiploid carcinoma samples, diploid subpopulations showed a smaller range of allelic imbalance at the loci tested compared to aneuploid subpopulations that demonstrated allelic imbalance over a relatively large region. Also, the frequency of AI at 11q24 showed a similar tendency to that at the ATM locus for each DNA ploidy state. An association between p53 gene mutation and ATM allelic imbalance in multiploid carcinoma was also observed. Our results suggest that ATM allelic imbalance and p53 gene mutations occur during the progression from diploid to aneuploid cell populations in multiploid colorectal carcinomas.

Allelic Imbalance↗

PubMed2004

Direct regulation of CREB transcriptional activity by ATM in response to genotoxic stress.

Ataxia-telangiectasia (A-T) is a syndrome of cancer susceptibility, immune dysfunction, and neurodegeneration that is caused by mutations in the A-T-mutated (ATM) gene. ATM has been implicated as a critical regulator of cellular responses to DNA damage, including the activation of cell cycle checkpoints and induction of apoptosis. Although defective cell cycle-checkpoint regulation and associated genomic instability presumably contribute to cancer susceptibility in A-T, the mechanism of neurodegeneration in A-T is not well understood. In addition, although ATM is required for the induction of the p53 transcriptional program in response to DNA damage, the identities of the relevant transcription factors that mediate ATM-dependent changes in gene expression remain largely undetermined. In this article, we describe a signal transduction pathway linking ATM directly to the Ca(2+)/cAMP response element-binding protein, CREB, a transcription factor that regulates cell growth, homeostasis, and survival. ATM phosphorylated CREB in vitro and in vivo in response to ionizing radiation (IR) and H(2)O(2) on a stress-inducible domain. IR-induced phosphorylation of CREB correlated with a decrease in CREB transactivation potential and reduced interaction between CREB and its transcriptional coactivator, CREB-binding protein (CBP). A CREB mutant containing Ala substitutions at ATM phosphorylation sites displayed enhanced transactivation potential, resistance to inhibition by IR, and increased binding to CBP. We propose that ATM-mediated phosphorylation of CREB in response to DNA damage modulates CREB-dependent gene expression and that dysregulation of the ATM-CREB pathway may contribute to neurodegeneration in A-T.

Amino Acid Sequence↗

PubMed2000

Mantle cell lymphoma is characterized by inactivation of the ATM gene.

In mantle cell lymphoma (MCL), the translocation t(11;14) is considered the cytogenetic hallmark of the disease. Recently, however, deletion of the chromosomal region 11q22-q23 has been identified as a frequent event in this type of cancer, indicating the existence of a pathogenically relevant tumor suppressor gene in this region. The deleted segment contains the ATM (ataxia telangiectasia mutated) gene. ATM is an interesting candidate as a tumor suppressor gene because constitutive inactivation of the gene predisposes ataxia telangiectasia patients to lymphoid malignancies. To assess the potential involvement of the gene in MCL lymphomagenesis, we performed mutation analysis of ATM in 12 sporadic cases of MCL, 7 of them with a deletion of one ATM gene copy, by using single-strand conformation polymorphism analysis of reverse transcription-PCR-amplified mRNA and subsequent DNA sequencing. In all seven cases containing a deletion of one ATM allele, a point mutation in the remaining allele was detected, which resulted in aberrant transcript splicing, truncation, or alteration of the protein. In addition, biallelic ATM mutations were identified in two MCLs that did not contain 11q deletions. Interestingly, in three cases analyzed, the ATM mutations detected in the tumor cells were not present in nonmalignant cells, demonstrating their somatic rather than germ-line origin. The inactivation of both alleles of the ATM gene by deletion and deleterious point mutation in the majority of cases analyzed indicates that ATM plays a role in the initiation and/or progression of MCL.

Alleles↗

PubMed2005

Drosophila ATM and ATR checkpoint kinases control partially redundant pathways for telomere maintenance.

In higher eukaryotes, the ataxia telangiectasia mutated (ATM) and ATM and Rad3-related (ATR) checkpoint kinases play distinct, but partially overlapping, roles in DNA damage response. Yet their interrelated function has not been defined for telomere maintenance. We discover in Drosophila that the two proteins control partially redundant pathways for telomere protection: the loss of ATM leads to the fusion of some telomeres, whereas the loss of both ATM and ATR renders all telomeres susceptible to fusion. The ATM-controlled pathway includes the Mre11 and Nijmegen breakage syndrome complex but not the Chk2 kinase, whereas the ATR-regulated pathway includes its partner ATR-interacting protein but not the Chk1 kinase. This finding suggests that ATM and ATR regulate different molecular events at the telomeres compared with the sites of DNA damage. This compensatory relationship between ATM and ATR is remarkably similar to that observed in yeast despite the fact that the biochemistry of telomere elongation is completely different in the two model systems. We provide evidence suggesting that both the loading of telomere capping proteins and normal telomeric silencing requires ATM and ATR in Drosophila and propose that ATM and ATR protect telomere integrity by safeguarding chromatin architecture that favors the loading of telomere-elongating, capping, and silencing proteins.

Animals↗

PubMed2003

Oligonucleotide microarrays demonstrate the highest frequency of ATM mutations in the mantle cell subtype of lymphoma.

Mutations have been described in the ataxia telangiectasia mutated (ATM) gene in small numbers of cases of lymphoid neoplasia. However, surveys of the ATM mutation status in lymphoma have been limited due to the large size (62 exons) and complex mutational spectrum of this gene. We have used microarray-based assays with 250,000 oligonucleotides to screen lymphomas from 120 patients for all possible ATM coding and splice junction mutations. The subtypes included were diffuse large B cell, mantle cell, immunoblastic large B cell, follicular, posttransplant lymphoproliferative disorder, and peripheral T cell lymphoma. We found the highest percentage of ATM mutations within the mantle cell (MCL) subtype (43%, 12 of 28 cases), followed by a lower level (10% of cases) in the other subtypes. A frame-shift ATM mutation was found in one peripheral T cell lymphoma patient. In six MCL cases examined, four ATM variants were due to somatic mutation in the tumor cells whereas two others seemed to be germ-line in origin. There was no difference in p53 mutation status in the ATM mutant and wild-type groups of MCL. There was no statistically significant difference in the median overall survival of patients with wild-type vs. mutated ATM in MCL. Additional mutational and functional analyses are needed to determine whether ATM mutations contribute to the development and progression of MCL or are just the consequence of genomic instability in MCL.

Ataxia Telangiectasia Mutated Proteins↗

PubMed2000

Abnormal development of Purkinje cells and lymphocytes in Atm mutant mice.

Motor incoordination, immune deficiencies, and an increased risk of cancer are the characteristic features of the hereditary disease ataxia-telangiectasia (A-T), which is caused by mutations in the ATM gene. Through gene targeting, we have generated a line of Atm mutant mice, Atm(y/y) mice. In contrast to other Atm mutant mice, Atm(y/y) mice show a lower incidence of thymic lymphoma and survive beyond a few months of age. Atm(y/y) mice exhibit deficits in motor learning indicative of cerebellar dysfunction. Even though we found no gross cerebellar degeneration in older Atm(y/y) animals, ectopic and abnormally differentiated Purkinje cells were apparent in mutant mice of all ages. These findings establish that some neuropathological abnormalities seen in A-T patients also are present in Atm mutant mice. In addition, we report a previously unrecognized effect of Atm deficiency on development or maintenance of CD4(+)8(+) thymocytes. We discuss these findings in the context of the hypothesis that abnormal development of Purkinje cells and lymphocytes contributes to the pathogenesis of A-T.

Animals↗

PubMed2001

Telomeric protein Pin2/TRF1 as an important ATM target in response to double strand DNA breaks.

ATM mutations are responsible for the genetic disease ataxia-telangiectasia (A-T). ATM encodes a protein kinase that is activated by ionizing radiation-induced double strand DNA breaks. Cells derived from A-T patients show many abnormalities, including accelerated telomere loss and hypersensitivity to ionizing radiation; they enter into mitosis and apoptosis after DNA damage. Pin2 was originally identified as a protein involved in G(2)/M regulation and is almost identical to TRF1, a telomeric protein that negatively regulates telomere elongation. Pin2 and TRF1, probably encoded by the same gene, PIN2/TRF1, are regulated during the cell cycle. Furthermore, up-regulation of Pin2 or TRF1 induces mitotic entry and apoptosis, a phenotype similar to that of A-T cells after DNA damage. These results suggest that ATM may regulate the function of Pin2/TRF1, but their exact relationship remains unknown. Here we show that Pin2/TRF1 coimmunoprecipitated with ATM, and its phosphorylation was increased in an ATM-dependent manner by ionizing DNA damage. Furthermore, activated ATM directly phosphorylated Pin2/TRF1 preferentially on the conserved Ser(219)-Gln site in vitro and in vivo. The biological significance of this phosphorylation is substantiated by functional analyses of the phosphorylation site mutants. Although expression of Pin2 and its mutants has no detectable effect on telomere length in transient transfection, a Pin2 mutant refractory to ATM phosphorylation on Ser(219) potently induces mitotic entry and apoptosis and increases radiation hypersensitivity of A-T cells. In contrast, Pin2 mutants mimicking ATM phosphorylation on Ser(219) completely fail to induce apoptosis and also reduce radiation hypersensitivity of A-T cells. Interestingly, the phenotype of the phosphorylation-mimicking mutants is the same as that which resulted from inhibition of endogenous Pin2/TRF1 in A-T cells by its dominant-negative mutants. These results demonstrate for the first time that ATM interacts with and phosphorylates Pin2/TRF1 and suggest that Pin2/TRF1 may be involved in the cellular response to double strand DNA breaks.

Amino Acid Sequence↗

PubMed2002

ATM is activated in response to N-methyl-N'-nitro-N-nitrosoguanidine-induced DNA alkylation.

p53 plays an important role in response to ionizing radiation by regulating cell cycle progression and triggering apoptosis. These activities are controlled, in part, by the phosphorylation of p53 by the protein kinase ATM. Recent evidence indicates that the monofunctional DNA alkylating agent N-methyl-N'-nitro-N- nitrosoguanidine (MNNG) also triggers up-regulation and phosphorylation of p53; however, the mechanism(s) responsible for this are unknown. We observed that in MNNG-treated normal human fibroblasts, up-regulation and phosphorylation of p53 was sensitive to the ATM kinase inhibitor wortmannin. ATM-deficient fibroblasts exhibited a delay in p53 up-regulation indicating a role for ATM in triggering the MNNG-induced response. Likewise, a mismatch repair (MMR)-deficient colorectal tumor line failed to show rapid up-regulation of p53. However, unlike ATM-deficient cells, these MMR-deficient cells displayed rapid phosphorylation of the p53 residue serine 15 after MNNG. In vitro kinase assays indicate that ATM is rapidly activated in both normal and MMR-deficient cells in response to MNNG. Using a number of morphological and biochemical approaches, we failed to observe MNNG-induced apoptosis in normal human fibroblasts, suggesting that apoptosis-induced DNA strand breaks are not required for the activation of ATM in response to MNNG. Comet assays indicated that strand breaks accumulated, and p53 up-regulation/phosphorylation occurred quite rapidly (within 30 min) after MNNG treatment, suggesting that DNA strand breaks that arise during the repair process activate ATM. These findings indicate that ATM activation is not limited to the ionizing radiation-induced response and potentially plays an important role in response to DNA alkylation.

Alkylation↗

PubMed2004

Double strand break repair by homologous recombination is regulated by cell cycle-independent signaling via ATM in human glioma cells.

To investigate double strand break (DSB) repair and signaling in human glioma cells, we stably transfected human U87 (ATM(+), p53(+)) glioma cells with a plasmid having a single I-SceI site within an inactive green fluorescent protein (GFP) expression cassette, allowing for the detection of homologous recombination repair (HRR) by GFP expression. HRR and nonhomologous end joining (NHEJ) were also determined by PCR. DSB repair was first detected at 12 h postinfection with an adenovirus expressing I-SceI with repair reaching plateau levels between 24 and 48 h. Within this time frame, NHEJ predominated over HRR in the range of 3-50-fold. To assess the involvement of ATM in DSB repair, we first examined whether ATM was associated with the DSB. Chromatin immunoprecipitation showed that ATM was present at the site of the DSB as early as 18 h postinfection. In cells treated with caffeine, an inhibitor of ATM, HRR was reduced, whereas NHEJ was not. In support of this finding, GFP flow cytometry demonstrated that caffeine reduced HRR by 90% under conditions when ATM kinase activity was inhibited. Dominant-negative ATM expressed from adenovirus inhibited HRR by 45%, also having little to no effect on NHEJ. Furthermore, HRR was inhibited by caffeine in serum-starved cells arrested in G(0)/G(1), suggesting that ATM is also important for HRR outside of the S and G(2) cell cycle phases. Altogether, these results demonstrate that HRR contributes substantially to DSB repair in human glioma cells, and, importantly, ATM plays a critical role in regulating HRR but not NHEJ throughout the cell cycle.

Adenoviridae↗

PubMed2006

Rapid activation of ATR by ionizing radiation requires ATM and Mre11.

The ataxia-telangiectasia-mutated (ATM) and ATM- and Rad3-related (ATR) protein kinases are crucial regulatory proteins in genotoxic stress response pathways that pause the cell cycle to permit DNA repair. Here we show that Chk1 phosphorylation in response to hydroxyurea and ultraviolet radiation is ATR-dependent and ATM- and Mre11-independent. In contrast, Chk1 phosphorylation in response to ionizing radiation (IR) is dependent on ATR, ATM, and Mre11. The ATR and ATM/Mre11 pathways are generally thought to be separate with ATM activation occurring early and ATR activation occurring as a late response to double strand breaks. However, we demonstrate that ATR is activated rapidly by IR, and ATM and Mre11 enhance ATR signaling. ATR-ATR-interacting protein recruitment to double strand breaks is less efficient in the absence of ATM and Mre11. Furthermore, IR-induced replication protein A foci formation is defective in ATM- and Mre11-deficient cells. Thus, ATM and Mre11 may stimulate the ATR signaling pathway by converting DNA damage generated by IR into structures that recruit and activate ATR.

Antineoplastic Agents↗

PubMed2003

Differing effects of breast cancer 1, early onset (BRCA1) and ataxia-telangiectasia mutated (ATM) mutations on cellular responses to ionizing radiation.

PURPOSE: The ataxia-telangiectasia mutated (ATM) gene encodes a protein kinase, the activation of which is an early event in the cellular response to ionizing radiation. One of the many substrates of ATM is BRCA1 (breast cancer 1, early onset gene), which has been associated with susceptibility to breast and ovarian cancer, and has been implicated in DNA repair processes. Various cellular responses to radiation were analysed in cells with mutations in ATM or BRCA1 in an attempt to clarify which effects of ATM can be mediated through BRCA1. MATERIALS AND METHODS: The response to radiation of cells with mutations in ATM or BRCA1 was examined, as were BRCA1-mutant tumour cells transfected with an exogenous wild-type BRCA1 allele. Assays included cell-survival curves, studies of potentially lethal damage repair, measurement of chromosomal aberrations and of G1 arrest, and Western blot analysis of lysates of irradiated cells to determine the phosphorylation of the product of the human Mdm2 gene (HDM2). RESULTS: Both ATM and BRCA1 mutations were associated with sensitivity to ionizing radiation, deficient repair of potentially lethal damage and markedly increased chromosomal aberrations. A BRCA1-mutated tumour cell line HCC1937, like ATM mutant cells, did not exhibit a normal G1 arrest but, unlike ATM mutant cells, did exhibit phosphorylation of HDM2. Expression of wild-type BRCA1 in HCC1937 cells partially restored radioresistance, restored repair of potentially lethal damage and markedly reduced radiation-induced chromosomal aberrations. G1 arrest, however, was not restored by expression of BRCA1. CONCLUSIONS: The results are consistent with a model in which ATM phosphorylation of BRCA1 regulates DNA repair functions, particularly those involved in potentially lethal damage repair and chromosomal integrity, but not other aspects of the cellular response to radiation such as G1 cell cycle arrest. To the authors' knowledge, this is the first demonstration of the ability of exogenously expressed BRCA1 to restore the ability to perform potentially lethal damage repair and maintain chromosomal integrity in irradiated cells.

Ataxia Telangiectasia Mutated Proteins↗

PubMed1998

Lack of correlation between ATM protein expression and tumour cell radiosensitivity.

PURPOSE: Cells derived from individuals in which the ataxia telangiectasia (ATM) gene is mutated are hypersensitive to ionizing radiation. Whether differences in ATM protein levels exist among human malignant glioma cell lines and whether such differences are correlated with cellular radiosensitivity were determined. MATERIALS AND METHODS: Polyclonal antibodies were raised to separate regions of the ATM protein. ATM protein expression in human malignant glioma cell lines, SV40 transformed normal human fibroblasts and SV40 transformed AT fibroblasts was analysed by Western blotting. Reverse transcriptase polymerase chain reaction (RT-PCR) was used to assess the presence of ATM transcript. RESULTS: While ATM protein was detected in all cell extracts, significant differences in the level of expression were observed. There was no apparent correlation between cellular radiosensitivity and differences in ATM protein levels in these human glioma cells. Extremely low levels of ATM protein were observed in M059J cells, which provide the only example of DNA-dependent protein kinase (DNA-PKcs) deficiency in a cell line of human origin. CONCLUSIONS: Variations in the levels of ATM protein are insufficient to explain the differences in cellular radiosensitivity observed in a panel of human malignant glioma cell lines.

Ataxia Telangiectasia↗

PubMed2002

Mutation of the ATM gene is not involved in the pathogenesis of either follicle center lymphoma or its transformation to higher-grade lymphoma.

Loss of function of the ataxia-telangiectasia mutated (ATM) gene, located on human chromosome 11q22-23, is the cause of ataxia-telangiectasia (A-T), which is associated with an extremely high risk for lymphoma. Abnormalities in 11q22-23, including deletions and mutations of the ATM gene, have been reported in T-cell prolymphocytic leukemias, B-CLL and in mantle cell lymphoma. In a survey of gene expression in follicle center lymphomas (FCL) and diffuse large B-cell lymphomas (DLBCL), almost all FCL expressed significant levels of ATM and the majority of DLBCL expressed low levels of ATM. This finding raised the possibility that the transformation of some FCL to DLBCL might be associated with inactivation of the ATM gene. Therefore, we analyzed biopsy specimens of 17 patients with FCL obtained at the time of diagnosis, four subsequent biopsies obtained at the time of FCL relapse and seven subsequent biopsies at the time of transformation to DLBCL. A comprehensive analysis of the ATM gene was performed by denaturing high performance liquid chromatography and sequencing. The analysis covered all of the 66 exons including the 9168 base pairs of ATM coding sequence as well as 16,676 base pairs of non-coding sequence. Twenty-eight known polymorphisms and rare sequence variants were observed, but no classic A-T mutations were detected. In 11 tumors, both tumor B-cells and normal T-cells were sorted for separate examination, and in each case, polymorphisms and rare variants were present in both tumor and normal cells. No new ATM gene mutations were associated with transformation from FCL to DLBCL. Thus, ATM gene mutations do not play a pivotal role either in the pathogenesis of FCL or in its transformation to DLBCL.

Ataxia Telangiectasia Mutated Proteins↗

PubMed2003

Loss of atm sensitises p53-deficient cells to topoisomerase poisons and antimetabolites.

BACKGROUND: Ataxia-telangiectasia is a pleiotropic autosomal recessive disorder caused by mutations in the ATM gene. In addition to a profound cancer predisposition, another hallmark of ataxia-telangiectasia is radiosensitivity. Recently, p53-null mouse fibroblasts have been reported to be radiosensitised by the concurrent loss of ATM. MATERIALS AND METHODS: We compared the sensitivity of atm(+/+)/p53(-/-) and atm(-/-)/p53(-/-) mouse embryonic fibroblasts to different classes of chemotherapeutic agents using the MTT assay, Trypan Blue exclusion and fluorescence-activated cell sorting for cell cycle and apoptosis analyses. RESULTS: Loss of ATM function in p53-deficient cells resulted in a 2- to 4-fold increase in sensitivity to the topoisomerase I poisons camptothecin and topotecan, to the topoisomerase II poisons doxorubicin, epirubicin and etoposide, and to the antimetabolites 5-fluorouracil and gemcitabine, but not to the platinum compounds cisplatin, carboplatin and oxaliplatin, the taxanes docetaxel and paclitaxel, or to busulfan. Loss of ATM function did not result in increased apoptosis, but resulted in increased Trypan Blue staining in response to epirubicin, suggesting that processes other than apoptosis may mediate cytotoxicity. ATM deficiency did not alter the extent of G(1)/S or G(2)/M cell cycle phase accumulation produced by epirubicin, suggesting that enhanced sensitivity was not due to failure of checkpoint activation. CONCLUSIONS: We provide further evidence that ATM is involved in regulating cellular defences against some cytotoxic agents in the absence of p53. Tumour-targeted functional inhibition of ATM may be a valuable strategy for increasing the efficacy of anticancer agents in the treatment of p53-mutant cancers.

Animals↗

PubMed2001

Heterozygosity for a mutation in Brca1 or Atm does not increase susceptibility to ENU-induced mammary tumors in Apc(Min)/+ mice.

The proteins encoded by BRCA1 and ATM may be important in DNA repair and maintenance of genomic integrity. Women heterozygous for a mutation in BRCA1 have an increased incidence of breast cancer. Some evidence also suggests that female carriers of ATM mutations may be susceptible to breast cancer. However, mice carrying one mutant allele of Brca1 or ATM are not highly susceptible to breast cancer. We proposed that heterozygosity for a mutant allele of Brca1 or ATM may confer a decreased ability to repair DNA damage. Such a defect might lead to a heightened sensitivity to tumor development in susceptible animal models. Therefore, mice predispose to mammary tumor development might show an increased susceptibility if they also carry an ATM or Brca1 mutation. C57BL/6J (B6) MIN/+ mice are predisposed to mammary and intestinal tumors and exposure to the point mutagen ethylnitrosourea (ENU) markedly increases mammary tumor multiplicity and incidence. To test our hypothesis, B6.MIN/+ male mice were crossed with 129S6/SvEvTac females heterozygous for a mutant allele of either Brca1 or ATM. Female progeny from each cross were treated with ENU and followed for tumor development. Only MIN/+ F1 females developed mammary tumors and heterozygosity for a mutant Brca1 or ATM allele had no effect on mammary or intestinal tumor incidence and multiplicity. These results suggest that heterozygosity for a mutation in Brca1 or ATM: does not affect MIN-induced tumorigenesis in mice under these conditions. Additionally, exposure to a somatic point mutagen does not increase tumor development in mice carrying Brca1 or ATM mutations.

Animals↗

PubMed2005

Induction and utilization of an ATM signaling pathway by polyomavirus.

Progression from G(1) to S is essential for polyomavirus DNA replication and depends on the interaction of large T with the retinoblastoma gene product pRb. This virus-induced replication pathway is accompanied by p53 activation resembling a DNA damage response (12). We sought to determine whether this pathway depends in part on activation of the ATM (ataxia telangiectasia mutated) kinase and whether the virus gains advantages from this pathway beyond that of entry into S. We show that polyomavirus infection activates the S- and G(2)-phase checkpoints in primary as well as established mouse cells. Infected cells undergo a prolonged S phase compared to uninfected serum-stimulated cells and show no evidence of a G(2)-->M transition before lytic death ensues. Infection is accompanied by increases in ATM activity in vitro and in the level of ATM-S1981-P in vivo. The incubation of infected cells with caffeine, a known ATM inhibitor, did not block entry into S but reduced the rate of viral compared to cellular DNA synthesis. Importantly, caffeine lowered the yields of viral DNA an average of 3- to 6-fold and those of infectious virus by as much as 10-fold. Virus yields were 10-fold lower in ATM (-/-) p53(-/-) than in ATM(+/+) p53(-/-) mouse embryo fibroblasts, indicating a p53-independent role of ATM in productive infection. Replacement of the normal SMC1 (structural maintenance of chromosomes, or cohesin) protein, a critical ATM substrate in the DNA repair pathway, with its phosphorylation mutant SMC1(S957AS966A) also lowered virus yields by roughly 90%. We suggest that polyomavirus activates and utilizes a component(s) of an ATM pathway of DNA repair to prolong S phase and aid its own replication.

Animals↗

PubMed1998

Upregulation of ATM in sclerosing adenosis of the breast.

The gene mutated in ataxia telangiectasia (ATM) has an established tumour suppressor role in breast cancer. ATM appears to be expressed in most normal cells, including breast epithelium, where it has been postulated to have a nuclear role in cell cycle regulation following DNA damage. However, ATM is not upregulated after DNA damage. In this study, we demonstrate an absence of immunohistologically detectable levels of ATM in the normally quiescent myoepithelial cells that line normal breast ducts. This contrasts dramatically with the significant expression of ATM in the proliferative myoepithelium of sclerosing adenosis (n = 7). This upregulation of ATM suggests that ATM expression is coupled to the proliferative status of the myoepithelium. Our results also indicate that there are factors other than ATM gene mutations that can dramatically influence ATM expression in the breast and that these factors should be considered for their possible implications in carcinogenesis.

Actins↗

PubMed2005

Deficiency in the catalytic subunit of DNA-dependent protein kinase causes down-regulation of ATM.

Previous reports have suggested a connection between reduced levels of the catalytic subunit of DNA-dependent protein kinases (DNA-PKcs), a component of the nonhomologous DNA double-strand breaks end-joining system, and a reduction in ATM. We studied this possible connection in other DNA-PKcs-deficient cell types, and following knockdown of DNA-PKcs with small interfering RNA, Chinese hamster ovary V3 cells, lacking DNA-PKcs, had reduced levels of ATM and hSMG-1, but both were restored after transfection with PRKDC. Atm levels were also reduced in murine scid cells. Reduction of ATM in a human glioma cell line lacking DNA-PKcs was accompanied by defective signaling through downstream substrates, post-irradiation. A large reduction of DNA-PKcs was achieved in normal human fibroblasts after transfection with two DNA-PKcs small interfering RNA sequences. This was accompanied by a reduction in ATM. These data were confirmed using immunocytochemical detection of the proteins. Within hours after transfection, a decline in PRKDC mRNA was seen, followed by a more gradual decline in DNA-PKcs protein beginning 1 day after transfection. No change in ATM mRNA was observed for 2 days post-transfection. Only after the DNA-PKcs reduction occurred was a reduction in ATM mRNA observed, beginning 2 days post-transfection. The amount of ATM began to decline, starting about 3 days post-treatment, then it declined to levels comparable to DNA-PKcs. Both proteins returned to normal levels at later times. These data illustrate a potentially important cross-regulation between the nonhomologous end-joining system for rejoining of DNA double-strand breaks and the ATM-dependent damage response network of pathways, both of which operate to maintain the integrity of the genome.

Animals↗