The DNA-dependent protein kinase

Smith, Graeme C.M.; Jackson, Stephen

doi:10.1101/gad.13.8.916

Cited by 798 publications

(639 citation statements)

References 226 publications

(254 reference statements)

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“…To test this possibility, we studied MEFs from mice lacking Ku80 or the DNA-dependent protein kinase catalytic subunit (DNA-PKcs), which are essential for DNA double-strand break repair by non-homologous end joining 20 . Ku80 −/− MEFs senesced rapidly in both 20% and 3% oxygen (Fig.…”

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confidence: 99%

Oxygen sensitivity severely limits the replicative lifespan of murine fibroblasts

et al. 2003

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Most mammalian cells do not divide indefinitely, owing to a process termed replicative senescence. In human cells, replicative senescence is caused by telomere shortening, but murine cells senesce despite having long stable telomeres 1 . Here, we show that the phenotypes of senescent human fibroblasts and mouse embryonic fibroblasts (MEFs) differ under standard culture conditions, which include 20% oxygen. MEFs did not senesce in physiological (3%) oxygen levels, but underwent a spontaneous event that allowed indefinite proliferation in 20% oxygen. The proliferation and cytogenetic profiles of DNA repair-deficient MEFs suggested that DNA damage limits MEF proliferation in 20% oxygen. Indeed, MEFs accumulated more DNA damage in 20% oxygen than 3% oxygen, and more damage than human fibroblasts in 20% oxygen. Our results identify oxygen sensitivity as a critical difference between mouse and human cells, explaining their proliferative differences in culture, and possibly their different rates of cancer and ageing.Replicative senescence limits the proliferation of many cell types in culture and in vivo 2 .Human cells senesce replicatively largely because telomeres -DNA structures that cap chromosome ends -shorten and malfunction when replicated in the absence of telomerase, which most human cells do not express 1 . Senescent cells adopt a distinctive morphology and pattern of gene expression 2,3 , a phenotype also induced by telomere-independent events 2,4 such as DNA damage and activation of certain oncogenes. The senescence response is thought to suppress cancer 2 .MEFs are used widely in the study of replicative senescence, despite notable differences between human and rodent cells 1,2 . MEFs spontaneously overcome replicative senescence (immortalization) 5,6 , whereas human fibroblasts rarely do so. Moreover, MEF senescence relies predominantly on the p19 ARF /p53 tumour suppressor pathway, whereas human fibroblasts require loss of both p53 and retinoblastoma (Rb) tumour suppressor functions for Correspondence should be addressed to: J.C. (jcampisi@lbl.gov). Note: Supplementary Information is available on the Nature Cell Biology website. COMPETING FINANCIAL INTERESTSThe authors declare that they have no competing financial interests. Here, we compared the phenotypes of senescent mouse and human fibroblasts. We cultured MEFs from C57Bl/6 (or FVB; data not shown) mice under standard conditions, which include atmospheric (20%) oxygen (Fig. 1a). As expected 5 , the cells grew well for approximately 1 week (2-3 population doublings) before proliferation began to decline. After 4-5 weeks (8-10 population doublings), the cultures senesced (arrow), showing no change in cell number for more than 1 week and a senescent morphology. Proliferation eventually resumed, owing to outgrowth of immortal variants 5 . HHS Public AccessWe monitored the capacity for DNA synthesis at each passage by labelling cells with 3 Hthymidine for 3 days and counting radiolabelled nuclei. As reported 9 , the percentage of labelled nuclei d...

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Oxygen sensitivity severely limits the replicative lifespan of murine fibroblasts

et al. 2003

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“…DNA double-strand breaks (DSBs), which can be produced by ionizing radiation (IR) or replication inhibition (Jeggo et al, 1995;Rothstein et al, 2000;Saintigny et al, 2001), are highly toxic lesions which can lead to profound genome rearrangements. On the other hand, DSBs can also be advantageously used to generate genetic diversity in physiological processes such as meiosis or V(D)J recombination (Kleckner, 1996;Smith and Jackson, 1999;Cohen and Pollard, 2001;Jung and Alt, 2004).…”

Section: Introductionmentioning

confidence: 99%

XRCC4 in G1 suppresses homologous recombination in S/G2, in G1 checkpoint-defective cells

et al. 2006

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Non-homologous end joining (NHEJ) and homologous recombination (HR) are two pathways that can compete or cooperate for DNA double-strand break (DSB) repair. NHEJ was previously shown to act throughout the cell cycle whereas HR is restricted to late S/G2. Paradoxically, we show here that defect in XRCC4 (NHEJ) leads to over-stimulation of HR when cells were irradiated in G1, not in G2. However, XRCC4 defect did not modify the strict cell cycle regulation for HR (i.e. in S/G2) as attested by (i) the formation of Rad51 foci in late S/G2 whatever the XRCC4 status, and (ii) the fact that neither Rad51 foci nor HR (gene conversion plus single-strand annealing) events induced by ionizing radiation were detected when cells were maintained blocked in G1. Finally, both c-H2AX analysis and pulse field gel electrophoresis showed that following irradiation in G1, some DSBs reached S/G2 in NHEJ-defective cells. Taken together, our results show that when cells are defective in G1/S arrest, DSB produced in G1 and left unrepaired by XRCC4 can be processed by HR but in late S/G2.

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“…DNA-PK and PARP1 have important functions in DNA double strand break repair, recombination, and telomere maintenance but are also involved in chromatin and transcriptional control 36– 38 . For example, Ku proteins associate with a series of homeodomain proteins (HOXC4, OCT1, OCT2, DLX2) thereby recruiting them to DNA ends where they are phosphorylated by DNA-PK 39 .…”

Section: Resultsmentioning

confidence: 99%

Systems analysis of the prostate tumor suppressor NKX3.1 supports roles in DNA repair and luminal cell differentiation

Yang

Chung²,

et al. 2014

F1000Res

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NKX3.1 is a homeobox transcription factor whose function as a prostate tumor suppressor remains insufficiently understood because neither the transcriptional program governed by NKX3.1, nor its interacting proteins have been fully revealed. Using affinity purification and mass spectrometry, we have established an extensive NKX3.1 interactome which contains the DNA repair proteins Ku70, Ku80, and PARP, thus providing a molecular underpinning to previous reports implicating NKX3.1 in DNA repair. Transcriptomic profiling of NKX3.1-negative prostate epithelial cells acutely expressing NKX3.1 revealed a rapid and complex response that is a near mirror image of the gene expression signature of human prostatic intraepithelial neoplasia (PIN). Pathway and network analyses suggested that NKX3.1 actuates a cellular reprogramming toward luminal cell differentiation characterized by suppression of pro-oncogenic c-MYC and interferon-STAT signaling and activation of tumor suppressor pathways. Consistently, ectopic expression of NKX3.1 conferred a growth arrest depending on TNFα and JNK signaling. We propose that the tumor suppressor function of NKX3.1 entails a transcriptional program that maintains the differentiation state of secretory luminal cells and that disruption of NKX3.1 contributes to prostate tumorigenesis by permitting luminal cell de-differentiation potentially augmented by defects in DNA repair.

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The DNA-dependent protein kinase

Cited by 798 publications

References 226 publications

Oxygen sensitivity severely limits the replicative lifespan of murine fibroblasts

Oxygen sensitivity severely limits the replicative lifespan of murine fibroblasts

XRCC4 in G1 suppresses homologous recombination in S/G2, in G1 checkpoint-defective cells

Systems analysis of the prostate tumor suppressor NKX3.1 supports roles in DNA repair and luminal cell differentiation

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