1FC3.3 The amounts of ATM protein and residual kinase activity predict the phenotype in Ataxia-Telangiectasia: A genotype – phenotype study

Ataxia telangiectasia–mutated gene (ATM) is a 350-kDa protein whose function is defective in the autosomal recessive disorder ataxia telangiectasia (AT). Affinity-purified polyclonal antibodies were used to characterize ATM. Steady-state levels of ATM protein varied from undetectable in most AT cell lines to highly expressed in HeLa, U2OS, and normal human fibroblasts. Subcellular fractionation showed that ATM is predominantly a nuclear protein associated with the chromatin and nuclear matrix. ATM protein levels remained constant throughout the cell cycle and did not change in response to serum stimulation. Ionizing radiation had no significant effect on either the expression or distribution of ATM. ATM immunoprecipitates from HeLa cells and the human DNA-dependent protein kinase null cell line MO59J, but not from AT cells, phosphorylated the 34-kDa subunit of replication protein A (RPA) complex in a single-stranded and linear double-stranded DNA–dependent manner. Phosphorylation of p34 RPA occurred on threonine and serine residues. Phosphopeptide analysis demonstrates that the ATM-associated protein kinase phosphorylates p34 RPA on similar residues observed in vivo. The DNA-dependent protein kinase activity observed for ATM immunocomplexes, along with the association of ATM with chromatin, suggests that DNA damage can induce ATM or a stably associated protein kinase to phosphorylate proteins in the DNA damage response pathway.

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Presence of ATM protein and residual kinase activity correlates with the phenotype in ataxia-telangiectasia: A genotype-phenotype study

Human Mutation ◽

10.1002/humu.22016 ◽

2012 ◽

Vol 33 (3) ◽

pp. 561-571 ◽

Cited By ~ 81

Author(s):

Mijke M. M. Verhagen ◽

James I. Last ◽

Frans B. L. Hogervorst ◽

Dominique F. C. M. Smeets ◽

Nel Roeleveld ◽

...

Keyword(s):

Ataxia Telangiectasia ◽

Kinase Activity ◽

Atm Protein

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Stable Brain ATM Message and Residual Kinase-Active ATM Protein in Ataxia-Telangiectasia

Journal of Neuroscience ◽

10.1523/jneurosci.0778-11.2011 ◽

2011 ◽

Vol 31 (20) ◽

pp. 7568-7577 ◽

Cited By ~ 17

Author(s):

J. Li ◽

J. Chen ◽

H. V. Vinters ◽

R. A. Gatti ◽

K. Herrup

Keyword(s):

Ataxia Telangiectasia ◽

Atm Protein

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Development of an Immunohistochemical Assay to Detect the Ataxia-Telangiectasia Mutated (ATM) Protein in Gastric Carcinoma

Applied Immunohistochemistry & Molecular Morphology ◽

10.1097/pai.0000000000000786 ◽

2020 ◽

Vol 28 (4) ◽

pp. 303-310 ◽

Cited By ~ 2

Author(s):

Rachel M. Miller ◽

Chinedu Nworu ◽

Laurel McKee ◽

Denis Balcerzak ◽

Linh Pham ◽

...

Keyword(s):

Gastric Carcinoma ◽

Ataxia Telangiectasia ◽

Ataxia Telangiectasia Mutated ◽

Immunohistochemical Assay ◽

Atm Protein

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Correction of ATM mutations in iPS cells from two ataxia-telangiectasia patients restores DNA damage and oxidative stress responses

Human Molecular Genetics ◽

10.1093/hmg/ddaa023 ◽

2020 ◽

Vol 29 (6) ◽

pp. 990-1001 ◽

Cited By ~ 5

Author(s):

Dmitry A Ovchinnikov ◽

Sarah L Withey ◽

Hannah C Leeson ◽

U Wang Lei ◽

Ashmitha Sundarrajan ◽

...

Keyword(s):

Oxidative Stress ◽

Dna Damage ◽

Stress Responses ◽

Ataxia Telangiectasia ◽

Compound Heterozygous ◽

Gene Correction ◽

Atm Kinase ◽

Atm Protein ◽

Induced Apoptosis ◽

And Oxidative Stress

Abstract Patients with ataxia-telangiectasia (A-T) lack a functional ATM kinase protein and exhibit defective repair of DNA double-stranded breaks and response to oxidative stress. We show that CRISPR/Cas9-assisted gene correction combined with piggyBac (PB) transposon-mediated excision of the selection cassette enables seamless restoration of functional ATM alleles in induced pluripotent stem cells from an A-T patient carrying compound heterozygous exonic missense/frameshift mutations, and from a patient with a homozygous splicing acceptor mutation of an internal coding exon. We show that the correction of one allele restores expression of ~ 50% of full-length ATM protein and ameliorates DNA damage-induced activation (auto-phosphorylation) of ATM and phosphorylation of its downstream targets, KAP-1 and H2AX. Restoration of ATM function also normalizes radiosensitivity, mitochondrial ROS production and oxidative-stress-induced apoptosis levels in A-T iPSC lines, demonstrating that restoration of a single ATM allele is sufficient to rescue key ATM functions. Our data further show that despite the absence of a functional ATM kinase, homology-directed repair and seamless correction of a pathogenic ATM mutation is possible. The isogenic pairs of A-T and gene-corrected iPSCs described here constitute valuable tools for elucidating the role of ATM in ageing and A-T pathogenesis.

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Ataxia telangiectasia mutated interacts with Parkin and induces mitophagy independent of kinase activity. Evidence from mantle cell lymphoma

Haematologica ◽

10.3324/haematol.2019.234385 ◽

2020 ◽

pp. haematol.2019.234385 ◽

Cited By ~ 2

Author(s):

Aloke Sarkar ◽

Christine M. Stellrecht ◽

Hima V. Vangapandu ◽

Mary Ayres ◽

Benny A. Kaipparettu ◽

...

Keyword(s):

Mantle Cell Lymphoma ◽

Ataxia Telangiectasia ◽

Kinase Activity ◽

Cell Lymphoma ◽

Ataxia Telangiectasia Mutated ◽

Mantle Cell

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DNA-PKcs promotes chromatin decondensation to facilitate initiation of the DNA damage response

Nucleic Acids Research ◽

10.1093/nar/gkz694 ◽

2019 ◽

Vol 47 (18) ◽

pp. 9467-9479 ◽

Cited By ~ 10

Author(s):

Huiming Lu ◽

Janapriya Saha ◽

Pauline J Beckmann ◽

Eric A Hendrickson ◽

Anthony J Davis

Keyword(s):

Dna Damage ◽

Dna Damage Response ◽

Ataxia Telangiectasia ◽

Cellular Response ◽

Kinase Activity ◽

Human Cell Line ◽

Cell Cycle Checkpoint ◽

Chromatin Decondensation ◽

Damage Site ◽

Damage Response

Abstract The DNA damage response (DDR) encompasses the cellular response to DNA double-stranded breaks (DSBs), and includes recognition of the DSB, recruitment of numerous factors to the DNA damage site, initiation of signaling cascades, chromatin remodeling, cell-cycle checkpoint activation, and repair of the DSB. Key drivers of the DDR are multiple members of the phosphatidylinositol 3-kinase-related kinase family, including ataxia telangiectasia mutated (ATM), ataxia telangiectasia and Rad3-related (ATR), and the DNA-dependent protein kinase catalytic subunit (DNA-PKcs). ATM and ATR modulate multiple portions of the DDR, but DNA-PKcs is believed to primarily function in the DSB repair pathway, non-homologous end joining. Utilizing a human cell line in which the kinase domain of DNA-PKcs is inactivated, we show here that DNA-PKcs kinase activity is required for the cellular response to DSBs immediately after their induction. Specifically, DNA-PKcs kinase activity initiates phosphorylation of the chromatin factors H2AX and KAP1 following ionizing radiation exposure and drives local chromatin decondensation near the DSB site. Furthermore, loss of DNA-PKcs kinase activity results in a marked decrease in the recruitment of numerous members of the DDR machinery to DSBs. Collectively, these results provide clear evidence that DNA-PKcs activity is pivotal for the initiation of the DDR.

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