Yang C R, Wilson-Van Patten C, Planchon S M, Wuerzberger-Davis S M, Davis T W, Cuthill S, Miyamoto S, Boothman D A
Departments of Radiation Oncology and Pharmacology and the Ireland Comprehensive Cancer Center, Laboratory of Molecular Stress Responses, Case Western Reserve University, Cleveland, Ohio 44106-4942, USA.
FASEB J. 2000 Feb;14(2):379-90. doi: 10.1096/fasebj.14.2.379.
Regulation of transcriptional responses in growth-arrested human cells under conditions that promote potentially lethal damage repair after ionizing radiation (IR) is poorly understood. Sp1/retinoblastoma control protein (RCP) DNA binding increased within 30 min and peaked at 2-4 h after IR (450-600 cGy) in confluent radioresistant human malignant melanoma (U1-Mel) cells. Increased phosphorylation of Sp1 directly corresponded to Sp1/RCP binding and immediate-early gene induction, whereas pRb remained hypophosphorylated. Transfection of U1-Mel cells with the human papillomavirus E7 gene abrogated Sp1/RCP induction and G(0)/G(1) cell cycle checkpoint arrest responses, increased apoptosis and radiosensitivity, and augmented genetic instability (i.e., increased polyploidy cells) after IR. Increased NF-kappaB DNA binding in U1-Mel cells after IR treatment lasted much longer (i.e., >20 h). U1-Mel cells overexpressing dominant-negative IkappaBalpha S32/36A mutant protein were significantly more resistant to IR exposure and retained both G(2)/M and G(0)/G(1) cell cycle checkpoint responses without significant genetic instability (i.e., polyploid cell populations were not observed). Nuclear p53 protein levels and DNA binding activity increased only after high doses of IR (>1200 cGy). Disruption of p53 responses in U1-Mel cells by E6 transfection also abrogated G(0)/G(1) cell cycle checkpoint arrest responses and increased polyploidy after IR, but did not alter radiosensitivity. These data suggest that abrogation of individual components of this coordinate IR-activated transcription factor response may lead to divergent alterations in cell cycle checkpoints, genomic instability, apoptosis, and survival. Such coordinate transcription factor activation in human cancer cells is reminiscent of prokaryotic SOS responses, and further elucidation of these events should shed light on the initial molecular events in the chromosome instability phenotype.-Yang, C.-R., Wilson-Van Patten, C., Planchon, S. M., Wuerzberger-Davis, S. M., Davis, T. W., Cuthill, C., Miyamoto, S., Boothman, D. A. Coordinate modulation of Sp1, NF-kappa B, and p53 in confluent human malignant melanoma cells after ionizing radiation.
在促进电离辐射（IR）后潜在致死性损伤修复的条件下，生长停滞的人类细胞中转录反应的调控机制目前还知之甚少。在汇合的耐辐射人类恶性黑色素瘤（U1-Mel）细胞中，Sp1/视网膜母细胞瘤控制蛋白（RCP）的DNA结合在IR（450-600 cGy）后30分钟内增加，并在2-4小时达到峰值。Sp1磷酸化增加直接对应于Sp1/RCP结合和立即早期基因诱导，而pRb仍处于低磷酸化状态。用人乳头瘤病毒E7基因转染U1-Mel细胞可消除Sp1/RCP诱导和G(0)/G(1)细胞周期检查点停滞反应，增加凋亡和放射敏感性，并增强IR后的遗传不稳定性（即多倍体细胞增加）。IR处理后U1-Mel细胞中NF-κB DNA结合增加持续时间更长（即>20小时）。过表达显性负性IκBα S32/36A突变蛋白的U1-Mel细胞对IR暴露的抗性明显更强，并保留G(2)/M和G(0)/G(1)细胞周期检查点反应，且无明显遗传不稳定性（即未观察到多倍体细胞群体）。仅在高剂量IR（>1200 cGy）后，核p53蛋白水平和DNA结合活性才增加。通过E6转染破坏U1-Mel细胞中的p53反应也可消除G(0)/G(1)细胞周期检查点停滞反应，并增加IR后的多倍体，但不改变放射敏感性。这些数据表明，消除这种协调的IR激活转录因子反应的单个成分可能导致细胞周期检查点、基因组不稳定性、凋亡和存活方面的不同改变。人类癌细胞中的这种协调转录因子激活让人联想到原核生物的SOS反应，对这些事件的进一步阐明应有助于揭示染色体不稳定性表型中的初始分子事件。-杨，C.-R.，威尔逊-范·帕滕，C.，普兰雄，S.M.，伍尔兹伯格-戴维斯，S.M.，戴维斯，T.W.，卡思卡特，C.，宫本，S.，布斯曼，D.A.电离辐射后汇合的人类恶性黑色素瘤细胞中Sp1、NF-κB和p53的协调调节。
