53BP1-dependent robust localized KAP-1 phosphorylation is essential for heterochromatic DNA double-strand break repair

Noon, Angela T.; Shibata, Atsushi; Rief, Nicole; Löbrich, Markus; Stewart, Grant S.; Jeggo, Penelope A.; Goodarzi, Aaaron A

doi:10.1038/ncb2017

Cited by 291 publications

(379 citation statements)

References 36 publications

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“…Using this cutoff value, a total of 433 siRNAs were identified that potentially sensitize SW1575 to radiation (Supplementary Table S1). Interestingly, several genes involved in the DNA damage response were amongst these hits, such as RAD51, TOPBP1, HTATIP, RNF168, PNKP, and TRIM28 (15)(16)(17)(18)(19). A more detailed pathway analysis of the hits using Ingenuity Pathway Analysis software (www.ingenuity.com) showed that proteins belonging to specific cellular functions, such as cell-cycle control as well as cell death and survival, were significantly overrepresented in the hit list ( Table 1).…”

Section: Resultsmentioning

confidence: 99%

Genome-wide siRNA Screen Identifies the Radiosensitizing Effect of Downregulation of MASTL and FOXM1 in NSCLC

Nagel

Walsum

Buijze

et al. 2015

Molecular Cancer Therapeutics

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Lung cancer is the most common cancer worldwide and on top of that has a very poor prognosis, which is reflected by a 5-year survival rate of 5% to 15%. Radiotherapy is an integral part of most treatment regimens for this type of tumor, often combined with radiosensitizing cytotoxic drugs. In this study, we identified many genes that could potentially be exploited for targeted radiosensitization using a genome-wide siRNA screen in non-small cell lung cancer (NSCLC) cells. The screen identified 433 siRNAs that potentially sensitize lung cancer cells to radiation. Validation experiments showed that knockdown of expression of Forkhead box M1 (FOXM1) or microtubuleassociated serine/threonine kinase-like (MASTL) indeed causes radiosensitization in a panel of NSCLC cells. Strikingly, this effect was not observed in primary human fibroblasts, suggesting that the observed radiosensitization is specific for cancer cells. Phosphoproteomics analyses with and without irradiation showed that a number of cell-cycle-related proteins were significantly less phosphorylated after MASTL knockdown in comparison to the control, while there were no changes in the levels of phosphorylation of DNA damage response proteins. Subsequent analyses showed that MASTL knockdown cells respond differently to radiation, with a significantly shortened G 2 -M phase arrest and defects in cytokinesis, which are followed by a cell-cycle arrest. In summary, we have identified many potential therapeutic targets that could be used for radiosensitization of NSCLC cells, with MASTL being a very promising and druggable target to combine with radiotherapy.

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Section: Resultsmentioning

confidence: 99%

Genome-wide siRNA Screen Identifies the Radiosensitizing Effect of Downregulation of MASTL and FOXM1 in NSCLC

Nagel

Walsum

Buijze

et al. 2015

Molecular Cancer Therapeutics

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“…Although DDR signaling was suppressed, our qRT-PCR analysis, however, revealed that the NHEJ pathway of DNA repair was not significantly downregulated (Figure 2a), as shown for DNA-PKcs, KU70, KU80, ligase 4 (LIG4), XRCC4 and 53BP1 gene expression levels -the latter was recently reported to be involved also in DNA repair. 14 We then comparatively studied the ATM-dependent DDR signaling in irradiated NSC and astrocytes by the means of western blot analyses. In NSC, irradiation lead to ATM autophosphorylation and a mobility shift of CHK2, suggesting its DNA-damage-induced phosphorylation, whereas in astrocytes phospho-ATM was not detectable at 1 or 24 h after irradiation and CHK2 protein was completely absent (Figure 2b), in agreement with their gene expression profiles (Figure 2a).…”

Section: Resultsmentioning

confidence: 99%

Terminally differentiated astrocytes lack DNA damage response signaling and are radioresistant but retain DNA repair proficiency

2011

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The impact and consequences of damage generation into genomic DNA, especially in the form of DNA double-strand breaks, and of the DNA-damage response (DDR) pathways that are promptly activated, have been elucidated in great detail. Most of this research, however, has been performed on proliferating, often cancerous, cell lines. In a mammalian body, the majority of cells are terminally differentiated (TD), and derives from a small pool of self-renewing somatic stem cells. Here, we comparatively studied DDR signaling and radiosensitivity in neural stem cells (NSC) and their TD-descendants, astrocytes -the predominant cells in the mammalian brain. Astrocytes have important roles in brain physiology, development and plasticity. We discovered that NSC activate canonical DDR upon exposure to ionizing radiation. Strikingly, astrocytes proved radioresistant, lacked functional DDR signaling, with key DDR genes such as ATM being repressed at the transcriptional level. Nevertheless, astrocytes retain the expression of non-homologous end-joining (NHEJ) genes and indeed they are DNA repair proficient. Unlike in NSC, in astrocytes DNA-PK seems to be the PI3K-like protein kinase responsible for cH2AX signal generation upon DNA damage. We also demonstrate the lack of functional DDR signaling activation in vivo in astrocytes of irradiated adult mouse brains, although adjacent neurons activate the DDR.

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“…DNA repair is no exception, and dynamic changes in chromatin condensation and accompanying histone marks have been recognized as inevitable DDR pathways [15,[75][76][77][78][79][80]. Here, too, the DDR relies on recruitment of existing players in the chromatin organization arena [32][33][34][75][76][77][78][79][81][82][83][84][85]. H2B monoubiquitylation was previously implicated in the DDR in the budding yeast [86][87][88].…”

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confidence: 99%

RNF20–RNF40: A ubiquitin‐driven link between gene expression and the DNA damage response

et al. 2011

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a b s t r a c tThe DNA damage response (DDR) is emerging as a vast signaling network that temporarily modulates numerous aspects of cellular metabolism in the face of DNA lesions, especially critical ones such as the double strand break (DSB). The DDR involves extensive dynamics of protein post-translational modifications, most notably phosphorylation and ubiquitylation. The DSB response is mobilized primarily by the ATM protein kinase, which phosphorylates a plethora of key players in its various branches. It is based on a core of proteins dedicated to the damage response, and a cadre of proteins borrowed temporarily from other cellular processes to help meet the challenge. A recently identified novel component of the DDR pathway -histone H2B monoubiquitylationexemplifies this principle. In mammalian cells, H2B monoubiquitylation is driven primarily by an E3 ubiquitin ligase composed of the two RING finger proteins RNF20 and RNF40. Generation of monoubiquitylated histone H2B (H2Bub) has been known to be coupled to gene transcription, presumably modulating chromatin decondensation at transcribed regions. New evidence indicates that the regulatory function of H2Bub on gene expression can selectively enhance or suppress the expression of distinct subsets of genes through a mechanism involving the hPAF1 complex and the TFIIS protein. This delicate regulatory process specifically affects genes that control cell growth and genome stability, and places RNF20 and RNF40 in the realm of tumor suppressor proteins. In parallel, it was found that following DSB induction, the H2B monoubiquitylation module is recruited to damage sites where it induces local H2Bub, which in turn is required for timely recruitment of DSB repair protein and, subsequently, timely DSB repair. This pathway represents a crossroads of the DDR and chromatin organization, and is a typical example of how the DDR calls to action functional modules that in unprovoked cells regulate other processes. Ó 2011 Federation of European Biochemical Societies. Published by Elsevier B.V. The DNA damage response: borrowing functional modules in an emergencyCellular metabolism is controlled by numerous, interlocked signaling networks. These networks are constantly responding to internal and external stimuli related to the cell's normal life cycle and functions, and to threats to its well being. A major threat to cellular homeostasis is subversion of its genomic stability, which may lead to undue cell death or to neoplasia [1,2]. DNA damage caused by internal or external damaging agents is a major danger to the integrity of the cellular genome. The cellular defense system against this threat is the DNA damage response (DDR) -an elaborate signaling network activated by DNA damage, which swiftly modulates many physiological processes [3,4]. A strong trigger of the DDR is the DNA double-strand break (DSB) [5,6]. DSBs are induced by ionizing radiation, radiomimetic chemicals, reactive oxygen species formed in the course of normal metabolism, and can also result from replic...

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53BP1-dependent robust localized KAP-1 phosphorylation is essential for heterochromatic DNA double-strand break repair

Cited by 291 publications

References 36 publications

Genome-wide siRNA Screen Identifies the Radiosensitizing Effect of Downregulation of MASTL and FOXM1 in NSCLC

Genome-wide siRNA Screen Identifies the Radiosensitizing Effect of Downregulation of MASTL and FOXM1 in NSCLC

Terminally differentiated astrocytes lack DNA damage response signaling and are radioresistant but retain DNA repair proficiency

RNF20–RNF40: A ubiquitin‐driven link between gene expression and the DNA damage response

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