Isolation of unique STAT5 targets by chromatin immunoprecipitation-based gene identification
Isolation of unique STAT5 targets by chromatin immunoprecipitation-based gene identification
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DOI：
10.1074/jbc.m408464200
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发表时间：
2004-12-24
影响因子：
4.8
通讯作者：
Frank, DA
中科院分区：
文献类型：
--
作者：
Nelson, EA;Walker, SR;Frank, DA
STAT 5a和STAT 5 b是两个高度相关的转录因子，控制着细胞的基本功能。几个STAT 5靶点是已知的，尽管大多数可能仍然没有特征。为了鉴定一组更完整的STAT 5调控基因，我们使用了染色质免疫沉淀方法的修改，该方法不预先假定有关这些靶点的任何信息。利用STAT 5被白细胞介素-3激活的Ba/f3细胞，我们发现了新的STAT 5结合位点，这些位点可能是附近基因的调节区域。这些位点通常远离转录起始位点，并且大多数不含CpG岛，表明它们不在传统的启动子区域中。尽管如此，当检查这些STAT 5结合位点附近的基因表达时，所有基因均在Ba/f3细胞中表达，并且大多数基因受白细胞介素-3调节。此外，通过这种策略鉴定的基因在急性白血病中显示出独特的表达模式，急性白血病是以活化的STAT 5为特征的肿瘤。尽管两种STAT 5同种型都与测试的所有启动子结合，但STAT 5a和STAT 5 b以不同的动力学结合，表明这两种蛋白质的功能之间的至少一些差异是由它们的DNA结合活性介导的。因此，这种转录因子靶标鉴定方法代表了以无偏方式分离转录因子靶标的有效策略，并且它揭示了传统启动子之外的许多新的STAT 5依赖性调控区域。
STAT5a and STAT5b are two highly related transcription factors that control essential cellular functions. Several STAT5 targets are known, although it is likely that most remain uncharacterized. To identify a more complete set of STAT5-regulated genes, we used a modification of the chromatin immunoprecipitation procedure, which does not presuppose any information regarding these targets. Employing Ba/f3 cells in which STAT5 is activated by interleukin-3, we have identified novel STAT5 binding sites that may be regulatory regions for nearby genes. These sites are typically found far from transcription start sites, and most do not contain CpG islands, indicating that they are not in traditional promoter regions. Nonetheless, when the expression of genes near these STAT5 binding sites was examined, all were expressed in Ba/f3 cells, and most were modulated by interleukin-3. Furthermore, genes identified by this strategy show unique expression patterns in acute leukemias, tumors characterized by activated STAT5. Whereas both STAT5 isoforms bound to all promoters tested, STAT5a and STAT5b bound with different kinetics, suggesting that at least some of the differences between the functions of these two proteins are mediated by their DNA binding activity. Therefore, this method of transcription factor target identification represents an effective strategy to isolate transcription factor targets in an unbiased fashion, and it has revealed many novel STAT5-dependent regulatory regions outside of traditional promoters.