Bunker C A, Kingston R E
Department of Molecular Biology, Massachusetts General Hospital, Boston 02114, USA.
Proc Natl Acad Sci U S A. 1996 Oct 1;93(20):10820-5. doi: 10.1073/pnas.93.20.10820.
DNA binding by transcriptional activators is typically an obligatory step in the activation of gene expression. Activator binding and subsequent steps in transcription are repressed by genomic chromatin. Studies in vitro have suggested that overcoming this repression is an important function of some activation domains. Here we provide quantitative in vivo evidence that the activation domain of GAL4-VP16 can increase the affinity of GAL4 for its binding site on genomic DNA in mammalian cells. Moreover, the VP16 activation domain has a much greater stimulatory effect on expression from a genomic reporter gene than on a transiently transfected reporter gene, where factor binding is more permissive. We found that not all activation domains showed a greater activation potential in a genomic context, suggesting that only some activation domains can function in vivo to alleviate the repressive effects of chromatin. These data demonstrate the importance of activation domains in relieving chromatin-mediated repression in vivo and suggest that one way they function is to increase binding of the activator itself.
转录激活因子与DNA的结合通常是基因表达激活过程中的一个必要步骤。激活因子的结合以及转录的后续步骤会受到基因组染色质的抑制。体外研究表明，克服这种抑制是一些激活结构域的重要功能。在此，我们提供了体内定量证据，表明GAL4-VP16的激活结构域能够增加GAL4在哺乳动物细胞中对其基因组DNA结合位点的亲和力。此外，VP16激活结构域对基因组报告基因表达的刺激作用比对瞬时转染报告基因的刺激作用大得多，在瞬时转染报告基因中因子结合更为宽松。我们发现并非所有激活结构域在基因组环境中都表现出更大的激活潜力，这表明只有一些激活结构域能够在体内发挥作用以减轻染色质的抑制作用。这些数据证明了激活结构域在体内缓解染色质介导的抑制作用中的重要性，并表明它们发挥作用的一种方式是增加激活因子自身的结合。
