Kim Yeonjung, Clark David J
Laboratory of Cellular and Developmental Biology, National Institute of Diabetes and Digestive and Kidney Diseases, Building 50, Room 3148, National Institutes of Health, Bethesda, MD 20892-8028, USA.
Proc Natl Acad Sci U S A. 2002 Nov 26;99(24):15381-6. doi: 10.1073/pnas.242536699. Epub 2002 Nov 13.
Current models for the role of the SWISNF chromatin remodeling complex in gene regulation are focused on promoters, where the most obvious changes in chromatin structure occur. Here we present evidence that the SWISNF complex is involved in the remodeling of the chromatin structure of an entire gene in vivo. We compared the native chromatin structures of a small yeast plasmid containing the HIS3 gene purified from uninduced and induced cells. Relative to uninduced chromatin, induced chromatin displayed a large reduction in negative supercoiling, a large reduction in sedimentation rate, and increased accessibility to restriction enzymes with sites located both near and far from the HIS3 promoter. These observations indicate that the entire plasmid was remodeled as a result of induction. Loss of supercoiling required the presence of the SWISNF remodeling complex and the activator Gcn4p in vivo. The TATA boxes were not required, suggesting that remodeling was not the result of transcription. The induction-dependent loss of negative supercoiling was not apparent in cells, indicating that the supercoils were lost preferentially from induced chromatin during purification. Thus, induced HIS3 chromatin has a highly labile structure that is revealed as a result of purification. It is concluded that induction of HIS3 creates a domain of labile chromatin structure that extends far beyond the promoter to include the entire gene. We propose that the SWISNF complex is recruited to the HIS3 promoter by Gcn4p and then directs remodeling of a chromatin domain, with important implications for transcription.
目前关于SWISNF染色质重塑复合体在基因调控中作用的模型主要集中在启动子上，染色质结构在启动子处会发生最明显的变化。在此，我们提供证据表明SWISNF复合体在体内参与了整个基因染色质结构的重塑。我们比较了从未诱导细胞和诱导细胞中纯化得到的含有HIS3基因的小酵母质粒的天然染色质结构。相对于未诱导的染色质，诱导后的染色质负超螺旋大幅减少，沉降速率大幅降低，并且对位于HIS3启动子附近和远处的限制性内切酶的可及性增加。这些观察结果表明整个质粒因诱导而发生了重塑。体内超螺旋的丧失需要SWISNF重塑复合体和激活剂Gcn4p的存在。TATA框并非必需，这表明重塑不是转录的结果。负超螺旋的诱导依赖性丧失在细胞中并不明显，这表明在纯化过程中超螺旋优先从诱导的染色质中丢失。因此，诱导后的HIS3染色质具有高度不稳定的结构，这种结构在纯化过程中得以显现。可以得出结论，HIS3的诱导产生了一个不稳定染色质结构域，该结构域远远超出启动子，涵盖了整个基因。我们提出，SWISNF复合体被Gcn4p招募到HIS3启动子，然后指导染色质结构域的重塑，这对转录具有重要意义。
