Mukherjee Sonali, Berger Michael F, Jona Ghil, Wang Xun S, Muzzey Dale, Snyder Michael, Young Richard A, Bulyk Martha L
Division of Genetics, Department of Medicine, Harvard Medical School; Boston; Massachusetts 02115, USA.
Nat Genet. 2004 Dec;36(12):1331-9. doi: 10.1038/ng1473. Epub 2004 Nov 14.
We developed a new DNA microarray-based technology, called protein binding microarrays (PBMs), that allows rapid, high-throughput characterization of the in vitro DNA binding-site sequence specificities of transcription factors in a single day. Using PBMs, we identified the DNA binding-site sequence specificities of the yeast transcription factors Abf1, Rap1 and Mig1. Comparison of these proteins' in vitro binding sites with their in vivo binding sites indicates that PBM-derived sequence specificities can accurately reflect in vivo DNA sequence specificities. In addition to previously identified targets, Abf1, Rap1 and Mig1 bound to 107, 90 and 75 putative new target intergenic regions, respectively, many of which were upstream of previously uncharacterized open reading frames. Comparative sequence analysis indicated that many of these newly identified sites are highly conserved across five sequenced sensu stricto yeast species and, therefore, are probably functional in vivo binding sites that may be used in a condition-specific manner. Similar PBM experiments should be useful in identifying new cis regulatory elements and transcriptional regulatory networks in various genomes.
我们开发了一种基于DNA微阵列的新技术，称为蛋白质结合微阵列（PBM），它能够在一天内快速、高通量地表征转录因子体外DNA结合位点的序列特异性。利用PBM，我们确定了酵母转录因子Abf1、Rap1和Mig1的DNA结合位点序列特异性。将这些蛋白质的体外结合位点与其体内结合位点进行比较表明，PBM衍生的序列特异性能够准确反映体内DNA序列特异性。除了先前确定的靶标外，Abf1、Rap1和Mig1分别与107、90和75个假定的新靶标基因间区域结合，其中许多位于先前未表征的开放阅读框上游。比较序列分析表明，这些新确定的位点中有许多在五个测序的狭义酵母物种中高度保守，因此可能是体内功能性结合位点，可能以条件特异性方式使用。类似的PBM实验在鉴定各种基因组中的新顺式调控元件和转录调控网络方面应该是有用的。
