Mirzayans Razmik, Andrais Bonnie, Scott April, Wang Ying W, Weiss Robert H, Murray David
Department of Oncology, University of Alberta, Cross Cancer Institute, Edmonton, AB T6G 1Z2, Canada.
Division of Nephrology, Department of Internal Medicine, University of California, Davis, CA 95616, USA.
Int J Mol Sci. 2015 May 20;16(5):11609-28. doi: 10.3390/ijms160511609.
Phosphorylation of H2AX on Ser139 (γH2AX) after exposure to ionizing radiation produces nuclear foci that are detectable by immunofluorescence microscopy. These so-called γH2AX foci have been adopted as quantitative markers for DNA double-strand breaks. High numbers of spontaneous γH2AX foci have also been reported for some human solid tumor-derived cell lines, but the molecular mechanism(s) for this response remains elusive. Here we show that cancer cells (e.g., HCT116; MCF7) that constitutively express detectable levels of p21WAF1 (p21) exhibit low numbers of γH2AX foci (<3/nucleus), whereas p21 knockout cells (HCT116p21-/-) and constitutively low p21-expressing cells (e.g., MDA-MB-231) exhibit high numbers of foci (e.g., >50/nucleus), and that these foci are not associated with apoptosis. The majority (>95%) of cells within HCT116p21-/- and MDA-MB-231 cultures contain high levels of phosphorylated p53, which is localized in the nucleus. We further show an inverse relationship between γH2AX foci and nuclear accumulation of WIP1, an oncogenic phosphatase. Our studies suggest that: (i) p21 deficiency might provide a selective pressure for the emergence of apoptosis-resistant progeny exhibiting genomic instability, manifested as spontaneous γH2AX foci coupled with phosphorylation and nuclear accumulation of p53; and (ii) p21 might contribute to positive regulation of WIP1, resulting in dephosphorylation of γH2AX.
暴露于电离辐射后，丝氨酸139位点的H2AX磷酸化（γH2AX）会产生可通过免疫荧光显微镜检测到的核灶。这些所谓的γH2AX灶已被用作DNA双链断裂的定量标志物。也有报道称一些人类实体瘤衍生细胞系存在大量自发γH2AX灶，但这种反应的分子机制仍不清楚。在这里我们表明，组成性表达可检测水平p21WAF1（p21）的癌细胞（如HCT116；MCF7）显示出低数量的γH2AX灶（<3/细胞核），而p21基因敲除细胞（HCT116p21-/-）和组成性低表达p21的细胞（如MDA-MB-231）显示出高数量的灶（如>50/细胞核），并且这些灶与细胞凋亡无关。HCT116p21-/-和MDA-MB-231培养物中的大多数细胞（>95%）含有高水平的磷酸化p53，其定位于细胞核。我们进一步表明γH2AX灶与致癌磷酸酶WIP1的核积累之间存在负相关关系。我们的研究表明：（i）p21缺陷可能为表现出基因组不稳定的抗凋亡后代的出现提供选择压力，表现为自发γH2AX灶以及p53的磷酸化和核积累；（ii）p21可能有助于WIP1的正向调节，导致γH2AX去磷酸化。
