2118-P: Regulation of Ataxia-Telangiectasia and Rad3-Related (ATR)–Dependent DNA Damage Response by Nitric Oxide in ß-Cells

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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INTS3 controls the hSSB1-mediated DNA damage response

The Journal of Cell Biology ◽

10.1083/jcb.200907026 ◽

2009 ◽

Vol 187 (1) ◽

pp. 25-32 ◽

Cited By ~ 54

Author(s):

Jeffrey R. Skaar ◽

Derek J. Richard ◽

Anita Saraf ◽

Alfredo Toschi ◽

Emma Bolderson ◽

...

Keyword(s):

Dna Damage ◽

Signaling Pathway ◽

Dna Damage Response ◽

Ataxia Telangiectasia ◽

Binding Protein ◽

Ataxia Telangiectasia Mutated ◽

Uncharacterized Protein ◽

Damage Response ◽

Atm Signaling Pathway ◽

Atm Signaling

Human SSB1 (single-stranded binding protein 1 [hSSB1]) was recently identified as a part of the ataxia telangiectasia mutated (ATM) signaling pathway. To investigate hSSB1 function, we performed tandem affinity purifications of hSSB1 mutants mimicking the unphosphorylated and ATM-phosphorylated states. Both hSSB1 mutants copurified a subset of Integrator complex subunits and the uncharacterized protein LOC58493/c9orf80 (henceforth minute INTS3/hSSB-associated element [MISE]). The INTS3–MISE–hSSB1 complex plays a key role in ATM activation and RAD51 recruitment to DNA damage foci during the response to genotoxic stresses. These effects on the DNA damage response are caused by the control of hSSB1 transcription via INTS3, demonstrating a new network controlling hSSB1 function.

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Sensitivity of tumor cells deficient in the fanconi anemia pathway to inhibition of ataxia telangiectasia mutated (ATM)

Journal of Clinical Oncology ◽

10.1200/jco.2007.25.18_suppl.10509 ◽

2007 ◽

Vol 25 (18_suppl) ◽

pp. 10509-10509

Author(s):

R. D. Kennedy ◽

P. Stuckert ◽

E. Archila ◽

M. De LaVega ◽

C. Chen ◽

...

Keyword(s):

Dna Damage ◽

Cell Lines ◽

Dna Damage Response ◽

Fanconi Anemia ◽

Ataxia Telangiectasia ◽

Pancreatic Head ◽

Genotoxic Stress ◽

Ataxia Telangiectasia Mutated ◽

Chromosomal Breakage ◽

Damage Response

10509 Loss of the fanconi anemia (FA) pathway function has been described in a number of sporadic tumor types including breast, ovarian, pancreatic, head and neck and hematological malignancies. Functionally, the FA pathway responds to stalled DNA replication following DNA damage. Given the importance of the FA pathway in the response to DNA damage, we hypothesized that cells deficient in this pathway may become hyper-dependent on alternative DNA damage response pathways in order to respond to endogenous genotoxic stress such as occurs during metabolism. Therefore, targeting these alternative pathways could offer therapeutic strategies in FA pathway deficient tumors. To identify new therapeutic targets we treated FA pathway competent and deficient cells with a DNA damage response siRNA library, that individually knocked out 230 genes. We identified a number of gene targets that were specifically toxic to FA pathway deficient cells, amongst which was the DNA damage response kinase Ataxia Telangiectasia Mutated (ATM). To test the requirement for ATM in FA pathway deficient cells, we interbred Fancg ± Atm± mice. Consistent with the siRNA screen result, Fancg-/- Atm-/- mice were non viable and Fancg± Atm-/- and Fancg-/- Atm ± progeny were less frequent that would have been expected. Several human cell lines with FA gene mutations were observed to have constitutive activation of ATM which was markedly reduced on correction with the appropriate wild-type FA gene. Interestingly, FA pathway deficient cells, including the FANCC mutant and FANCG mutant pancreatic cancer cell lines, were selectively sensitive to monotherapy with the ATM inhibitor KU55933, as measured by dose inhibition and colony count assays. FA pathway deficient cells also demonstrated an increased level of chromosomal breakage, cell cycle arrest and apoptosis following KU55933 treatment when compared to FA pathway corrected cells. We conclude that FA pathway deficient cells have an increased requirement for ATM activation in order to respond to sporadic DNA damage. This offers the possibility that monotherapy with ATM inhibitors could be a therapeutic strategy for tumors that are deficient for the FA pathway. No significant financial relationships to disclose.

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ATM regulates a DNA damage response posttranscriptional RNA operon in lymphocytes

Blood ◽

10.1182/blood-2010-09-310987 ◽

2011 ◽

Vol 117 (8) ◽

pp. 2441-2450 ◽

Cited By ~ 27

Author(s):

Krystyna Mazan-Mamczarz ◽

Patrick R. Hagner ◽

Yongqing Zhang ◽

Bojie Dai ◽

Elin Lehrmann ◽

...

Keyword(s):

Gene Expression ◽

Dna Damage ◽

Dna Damage Response ◽

Ataxia Telangiectasia ◽

Rna Binding ◽

Critical Role ◽

Cell Cycle Checkpoints ◽

Dependent Manner ◽

Damage Response ◽

Multiple Cell

Abstract Maintenance of genomic stability depends on the DNA damage response, a biologic barrier in early stages of cancer development. Failure of this response results in genomic instability and high predisposition toward lymphoma, as seen in patients with ataxia-telangiectasia mutated (ATM) dysfunction. ATM activates multiple cell-cycle checkpoints and DNA repair after DNA damage, but its influence on posttranscriptional gene expression has not been examined on a global level. We show that ionizing radiation modulates the dynamic association of the RNA-binding protein HuR with target mRNAs in an ATM-dependent manner, potentially coordinating the genotoxic response as an RNA operon. Pharmacologic ATM inhibition and use of ATM-null cells revealed a critical role for ATM in this process. Numerous mRNAs encoding cancer-related proteins were differentially associated with HuR depending on the functional state of ATM, in turn affecting expression of encoded proteins. The findings presented here reveal a previously unidentified role of ATM in controlling gene expression posttranscriptionally. Dysregulation of this DNA damage response RNA operon is probably relevant to lymphoma development in ataxia-telangiectasia persons. These novel RNA regulatory modules and genetic networks provide critical insight into the function of ATM in oncogenesis.

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