Profiles of global gene expression in ionizing-radiation-damaged human diploid fibroblasts reveal synchronization behind the G1 checkpoint in a G0-like state of quiescence.
Profiles of global gene expression in ionizing-radiation-damaged human diploid fibroblasts reveal synchronization behind the G1 checkpoint in a G0-like state of quiescence.
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DOI：
10.1289/ehp.8026
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发表时间：
2006-04
影响因子：
10.4
通讯作者：
Kaufmann WK
中科院分区：
文献类型：
--
作者：
Zhou T;Chou JW;Simpson DA;Zhou Y;Mullen TE;Medeiros M;Bushel PR;Paules RS;Yang X;Hurban P;Lobenhofer EK;Kaufmann WK
表达端粒酶的人二倍体成纤维细胞对电离辐射（IR）诱导的DNA损伤的细胞周期阻滞和刻板转录反应进行了定量研究。细胞毒性分析表明，1.5 Gy IR使3种成纤维细胞系的集落形成失活40-45%；该剂量用于所有后续分析。在ir后2小时，成纤维细胞从G2到M的进展阻滞率为90%，在ir后6和12小时，从G1到S的进展阻滞率同样严重。ir后6和12小时的正常DNA合成和有丝分裂率导致S和M室在24小时空出bbb70 %。在ir处理的细胞中分析全局基因表达。微阵列分析算法，EPIG，鉴定了三种成纤维细胞系共有的9种ir应答基因表达模式，包括显性p53依赖性G1检查点应答。许多p53靶基因，如CDKN1A、GADD45、BTG2和PLK3，在ir后2小时显著上调。许多受E2F家族转录因子调控的基因，包括CDK2、CCNE1、CDC6、CDC2、MCM2，在ir后24小时显著下调。在阻滞的成纤维细胞中，许多参与DNA代谢的基因也明显受到抑制，这显然是G1检查点后细胞同步的结果。然而，基因表达的聚类和主成分分析显示，ir后24小时的基因表达谱与G0生长静止相似。结果揭示了人类二倍体成纤维细胞对IR的高度刻板的反应模式，主要反映了G1检查点后的同步性，但突出地诱导了G0静止的其他标记，如GAS1。
Cell cycle arrest and stereotypic transcriptional responses to DNA damage induced by ionizing radiation (IR) were quantified in telomerase-expressing human diploid fibroblasts. Analysis of cytotoxicity demonstrated that 1.5 Gy IR inactivated colony formation by 40–45% in three fibroblast lines; this dose was used in all subsequent analyses. Fibroblasts exhibited > 90% arrest of progression from G2 to M at 2 hr post-IR and a similarly severe arrest of progression from G1 to S at 6 and 12 hr post-IR. Normal rates of DNA synthesis and mitosis 6 and 12 hr post-IR caused the S and M compartments to empty by > 70% at 24 hr. Global gene expression was analyzed in IR-treated cells. A microarray analysis algorithm, EPIG, identified nine IR-responsive patterns of gene expression that were common to the three fibroblast lines, including a dominant p53-dependent G1 checkpoint response. Many p53 target genes, such as CDKN1A, GADD45, BTG2, and PLK3, were significantly up-regulated at 2 hr post-IR. Many genes whose expression is regulated by E2F family transcription factors, including CDK2, CCNE1, CDC6, CDC2, MCM2, were significantly down-regulated at 24 hr post-IR. Numerous genes that participate in DNA metabolism were also markedly repressed in arrested fibroblasts apparently as a result of cell synchronization behind the G1 checkpoint. However, cluster and principal component analyses of gene expression revealed a profile 24 hr post-IR with similarity to that of G0 growth quiescence. The results reveal a highly stereotypic pattern of response to IR in human diploid fibroblasts that reflects primarily synchronization behind the G1 checkpoint but with prominent induction of additional markers of G0 quiescence such as GAS1.