Transcriptional networks and cellular senescence in human mammary fibroblasts
Transcriptional networks and cellular senescence in human mammary fibroblasts
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DOI：
10.1091/mbc.e04-05-0392
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发表时间：
2005-02-01
影响因子：
3.3
通讯作者：
Jat, PS
中科院分区：
文献类型：
--
作者：
Hardy, K;Mansfield, L;Jat, PS
衰老是限制细胞有限增殖潜力的分子程序，是保护身体免受癌症侵害的重要屏障。用于测量转录组变化和调节其表达的技术表明，有可能解剖复杂细胞过程的转录网络。HMF 3A细胞是条件永生化的人乳腺成纤维细胞，其可以被诱导经历协调衰老。在这里，我们将这些细胞与微阵列，RNA干扰和计算机启动子分析相结合，以促进负责调节细胞衰老的转录网络的解剖。我们首先确定了HMF 3A细胞衰老时转录组中的突变，然后将其与原代人乳腺成纤维细胞复制性衰老时观察到的突变进行比较。除了DUSP 1和已知的p53和E2 F靶点外，许多基因如PHLDA 1，NR 4A 3和STAC的新剪接变体也与衰老有关。然后通过RNA沉默和微阵列分析来分析它们在衰老中的作用。所有差异基因的计算机启动子分析预测，核因子-kappaB和C/EBP转录因子被激活衰老后，我们证实了这一点，电泳迁移率变动分析。这些结果提示了细胞衰老的一个假定信号网络。
Senescence, the molecular program that limits the finite proliferative potential of a cell, acts as an important barrier to protect the body from cancer. Techniques for measuring transcriptome changes and for modulating their expression suggest that it may be possible to dissect the transcriptional networks underlying complex cellular processes. HMF3A cells are conditionally immortalized human mammary fibroblasts that can be induced to under-go coordinated Senescence. Here, we used these cells in conjunction with microarrays, RNA interference, and in silico promoter analysis to promote the dissection of the transcriptional networks responsible for regulating cellular senescence. We first identified chancres in the transcriptome when HMF3A cells undergo senescence and then compared them with those observed upon replicative senescence in primary human mammary fibroblasts. In addition to DUSP1 and known p53 and E2F targets, a number of genes such as PHLDA1, NR4A3, and a novel splice variant of STAC were implicated in senescence. Their role in senescence was then analyzed by RNA silencing followed by microarray analysis. In silico promoter analysis of all differential genes predicted that nuclear factor-kappaB and C/EBP transcription factors are activated upon senescence, and we confirmed this by electrophoretic mobility shift assay. The results suggest a putative signaling network for cellular senescence.