Accumulation of p21 proteins at DNA damage sites independent of p53 and core NHEJ factors following irradiation

Koike, Manabu; Yutoku, Yasutomo; Koike, Aki

doi:10.1016/j.bbrc.2011.07.032

Cited by 40 publications

(41 citation statements)

References 27 publications

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“…While this manuscript was in preparation, Koike and collaborators reported the accumulation of CDKN1A at local sites of damaged DNA introduced by UV-A laser irradiation [47]. Similar to our experiments, these investigators describe the inability of ectopically expressed truncated CDKN1A that is unable to interact with PCNA, to accumulate into foci after UV-A laser irradiation.…”

Section: Discussionsupporting

confidence: 80%

“…Importantly, differences in the DNA damage spectra and the extremely high local dose from UV-A microlasers can lead to the recruitment of specific DNA-binding proteins and repair factors, which, after densely ionizing charged particle irradiation, do not localize to the sites of particle traversal [48,49]. In addition, while Koike and collaborators [47], in line with our results for high LET radiation, show that rapidly recruited EGFP-CDKN1A colocalizes with a DSB marker after UV-A laser irradiation, our new results in the present study clearly delineate that this is not the case after sparsely ionizing X-irradiation.…”

Section: Discussionmentioning

confidence: 99%

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PCNA-dependent accumulation of CDKN1A into nuclear foci after ionizing irradiation

et al. 2012

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Section: Discussionsupporting

confidence: 80%

Section: Discussionmentioning

confidence: 99%

PCNA-dependent accumulation of CDKN1A into nuclear foci after ionizing irradiation

et al. 2012

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“…The control mechanisms underlying the subcellular local- ization of Ku70 and Ku80 play a key role in regulating the physiological function of not only Ku itself, but also other NHEJ core factors [6,11,17,20]. Recently, using Ku70 −/− MLE cells, we have shown that Ku70 is essential for the accumulations of XRCC4 and XLF, but not those of Artemis and p21, at DSBs in the early stage following irradiation of lung epithelial cells [10,11]. Thus, the accumulation of the NHEJ core factors XRCC4 and XLF is dependent on Ku; however, that of another NHEJ factor, Artemis, which plays a role in both the NHEJ and HR pathways, is independent of Ku in MLE cells.…”

Section: Discussionmentioning

confidence: 99%

The Defect of Ku70 Affects Sensitivity to X-Ray and Radiation-Induced Caspase-Dependent Apoptosis in Lung Cells

Koike¹,

Yutoku

Koike³

2013

The Journal of Veterinary Medical Science

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ABSTRACT. The DNA repair protein Ku70 is a key player in chemoresistance to anticancer agents (e.g., etoposide) or radioresistance. The responses of different organs to radiation vary widely and likely depend on the cell population in the organs. Previously, we established and characterized Ku70-deficient murine lung epithelial (Ku70 −/− MLE) cells and found that these cells are more sensitive than Ku70 +/− MLE cells (control cells) to X-irradiation, as determined by clonogenic survival assay; however, the mechanism underlying this sensitivity remains unclear. In this study, we examined the mechanism by which X-irradiation triggers the death of Ku70 −/− MLE cells. Our results showed that Ku70 −/− MLE cells were more sensitive to radiation-induced apoptosis than control cells, although X-irradiation activated caspase-3 and caspase-7, and cleaved PARP in both cell lines. We also examined the expression level of phosphorylated H2AX (γH2AX), which is a marker of DSB, and observed the phosphorylation of H2AX and the elimination of γH2AX in both cell lines after X-irradiation. The elimination in Ku70 −/− MLE cells was slower than that in control cells, suggesting that DSB repair activity in the Ku70 −/− MLE cells is lower than that in control cells. These findings suggest that Ku70 might play a key role in the inhibition of apoptosis through the DSB repair pathway in lung epithelial cells. Our findings also suggest that these cell lines might be useful for the study of Ku70 functions and the Ku70-dependent DSB repair pathway in lung epithelial cells. KEY WORDS: anticancer treatment, epithelial cell, γH2AX, Ku70, Ku80.

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“…Western blot analysis: The extraction of total lysates and Western blot analysis were performed as described previously with modifications [19,21]. Briefly, the lysates were electrophoresed on 5-20% SDS-polyacrylamide gels (Wako, Osaka, Japan).…”

Section: Cell Lines and Culturesmentioning

confidence: 99%

“…DNA was stained with 0.025 µg/ml 4,6-diamino-2-phenylindole (DAPI) fluorescent dye (boehringer Mannheim, Mannheim, Germany), and the cells were examined under an Olympus IX 70 fluorescence microscope (Olympus) to determine the localization of proteins. Images were acquired using an FV300 confocal laser scanning microscope (Olympus) as previously described [17,19,21].…”

Section: Cell Lines and Culturesmentioning

confidence: 99%

Establishment of Hamster Cell Lines with EGFP-Tagged Human XRCC4 and Protection from Low-Dose X-Ray Radiation

Koike

Yutoku

Koike

2012

The Journal of Veterinary Medical Science

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AbSTRAcT. In clinical settings, cellular resistance to chemotherapy and radiotherapy is a significant component of tumor treatment failure. The mechanisms underlying the control of localization of DNA repair proteins play a key role in the regulation of DNA repair activity. The DNA repair protein XRCC4, which is a regulator of DNA ligase IV activity, might be a key contributor to not only chemoresistance to anticancer agents, e.g., etoposide, but also radioresistance. However, it remains unclear whether XRCC4, which is a key player in nonhomologous DNA-end-joining (NHEJ), plays a role in low-dose radioresistance. In this study, we confirmed that human XRCC4 tagged with the enhanced green fluorescent protein (EGFP-XRCC4), as well as the DNA damage sensor Ku80 tagged with EGFP, mainly localized in the nuclei and its accumulation at DNA damaged sites began immediately after microirradiation. Moreover, we generated and characterized cell lines expressing EGFP-XRCC4 in XRCC4-deficient cells, i.e., XR-1 cells derived from the Chinese hamster ovary. Our findings showed that XR-1 cells were more sensitive than controls (CHO-K1) to low-dose X-irradiation (<0.5 Gy), whereas the radiosensitive phenotype of XR-1 cells was rescued by the expression of EGFP-XRCC4. We also confirmed that EGFP-XRCC4 expressed stably in XR-1 cells stabilizes DNA ligase IV. Altogether, these cell lines might be useful for the study of not only the dynamics and function of XRCC4, but also the molecular mechanism underlying the cellular resistance via the NHEJ pathway to low-dose radiation in mammalian cells.

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Accumulation of p21 proteins at DNA damage sites independent of p53 and core NHEJ factors following irradiation

Cited by 40 publications

References 27 publications

PCNA-dependent accumulation of CDKN1A into nuclear foci after ionizing irradiation

PCNA-dependent accumulation of CDKN1A into nuclear foci after ionizing irradiation

The Defect of Ku70 Affects Sensitivity to X-Ray and Radiation-Induced Caspase-Dependent Apoptosis in Lung Cells

Establishment of Hamster Cell Lines with EGFP-Tagged Human XRCC4 and Protection from Low-Dose X-Ray Radiation

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