Department of Chemistry and Biochemistry, University of Colorado, Boulder, CO 80309, USA.
Proc Natl Acad Sci U S A. 2010 Jun 22;107(25):11283-8. doi: 10.1073/pnas.0914215107. Epub 2010 Jun 4.
The 26-subunit, 1.2 MDa human Mediator complex is essential for expression of perhaps all protein-coding genes. Activator binding triggers major structural shifts within Mediator, suggesting a straightforward means to spatially and temporally regulate Mediator activity. By using mass spectrometry (MudPIT) and other techniques, we have compared the subunit composition of Mediator in three different structural states: bound to the activator SREBP-1a, VP16, or an activator-free state. As expected, consensus Mediator subunits were similarly represented in each sample. However, we identify a set of cofactors that interact specifically with activator-bound but not activator-free Mediator, suggesting activator binding triggers new Mediator-cofactor interactions. Furthermore, MudPIT combined with biochemical assays reveals a nonoverlapping set of coregulatory factors associated with SREBP-Mediator vs. VP16-Mediator. These data define an expanded role for activators in regulating gene expression in humans and suggest that distinct, activator-induced structural shifts regulate Mediator function in gene-specific ways.
人 Mediator 复合物由 26 个亚基组成，分子量为 1.2MDa，对于表达或许所有的蛋白质编码基因都是必需的。激活剂结合触发 Mediator 内的主要结构转变，这表明可以通过空间和时间调节 Mediator 活性的一种直接方法。通过使用质谱（MudPIT）和其他技术，我们比较了 Mediator 在三种不同结构状态下的亚基组成：与激活剂 SREBP-1a、VP16 或无激活剂状态结合。正如预期的那样，在每个样品中都有类似的共识 Mediator 亚基。然而，我们确定了一组与激活剂结合但不与无激活剂 Mediator 相互作用的特定辅助因子，这表明激活剂结合触发了新的 Mediator-辅助因子相互作用。此外，MudPIT 结合生化分析揭示了一组与 SREBP-Mediator 与 VP16-Mediator 相关的非重叠核心调节因子。这些数据定义了激活剂在调节人类基因表达中的扩展作用，并表明不同的、激活剂诱导的结构转变以特定方式调节 Mediator 功能。
