Harrington L A, Andrews B J
Department of Molecular and Medical Genetics, University of Toronto, Ontario, Canada.
Nucleic Acids Res. 1996 Feb 15;24(4):558-65. doi: 10.1093/nar/24.4.558.
In Saccharomyces cerevisiae commitment to cell division occurs late in the G1 phase of the cell cycle at a point called Start and requires the activity of the Cdc28 protein kinase and its associated G1 cyclins. The Swi4,6-dependent cell cycle box binding factor, SBF, is important for maximal expression of the G1 cyclin and HO endonuclease genes at Start. The cell cycle regulation of these genes is modulated through an upstream regulatory element termed the SCB (SwI4,6-dependent cell cycle box, CACGAAA), which is dependent on both SWI4 and SWI6. Although binding of SWI4 and SWI6 to SCB sequences has been well characterized in vitro, the binding of SBF in vivo has not been examined. We used in vivo dimethyl sulfate footprinting to examine the occupancy of SCB sequences throughout the cell cycle. We found that binding to SCB sequences occurred in the G1 phase of the cell cycle and was greatly reduced in G2. In the absence of either SWI4 or SWI6, SCB sequences were not occupied at any cell cycle stage. These results suggest that the G1-specific expression of SCB-dependent genes is regulated at the level of DNA binding in vivo.
在酿酒酵母中，细胞分裂的启动发生在细胞周期G1期的晚期，在一个称为起始点（Start）的位置，并且需要Cdc28蛋白激酶及其相关的G1细胞周期蛋白的活性。Swi4,6依赖性细胞周期盒结合因子SBF，对于起始点处G1细胞周期蛋白和HO内切核酸酶基因的最大表达很重要。这些基因的细胞周期调控是通过一个称为SCB（Swi4,6依赖性细胞周期盒，CACGAAA）的上游调控元件来调节的，它依赖于SWI4和SWI6。尽管SWI4和SWI6与SCB序列的结合在体外已得到充分表征，但体内SBF的结合尚未被研究。我们使用体内硫酸二甲酯足迹法来检测整个细胞周期中SCB序列的占据情况。我们发现与SCB序列的结合发生在细胞周期的G1期，而在G2期则大大减少。在没有SWI4或SWI6的情况下，SCB序列在任何细胞周期阶段都未被占据。这些结果表明，SCB依赖性基因的G1特异性表达在体内DNA结合水平上受到调控。
