Brown S A, Imbalzano A N, Kingston R E
Department of Genetics, Harvard Medical School, Boston, Massachusetts 02114, USA.
Genes Dev. 1996 Jun 15;10(12):1479-90. doi: 10.1101/gad.10.12.1479.
Promoter-proximal pausing during transcriptional elongation is an important way of regulating many diverse genes, including human c-myc and c-fos, some HIV genes, and the Drosophila heat shock loci. To characterize the mechanisms that regulate pausing, we have established an in vitro system using the human hsp7O gene. We demonstrate that nucleosome formation increases by >100-fold the duration of a transcriptional pause on the human hsp7O gene in vitro at the same location as pausing is observed in vivo. Readthrough of this pause is increased by an activator that contains the human heat shock factor 1 (HSF1) transcriptional activation domains. Maximal effect of the activator requires that the system be supplemented with fractions that have hSWI/SNF activity, which has been shown previously to alter nucleosome structure. No significant readthrough is observed in the absence of activator, and neither the activator nor the hSWI/SNF fraction affected elongation on naked DNA; therefore, these results suggest that an activator can cause increased readthrough of promoter-proximal pausing by decreasing the inhibitory effect of nucleosomes on transcriptional elongation.
转录延伸过程中的启动子近端暂停是调控许多不同基因的重要方式，这些基因包括人类的c-myc和c-fos、一些HIV基因以及果蝇的热休克基因座。为了阐明调控暂停的机制，我们利用人类hsp70基因建立了一个体外系统。我们证明，在体外，核小体的形成使人类hsp70基因转录暂停的持续时间增加了100多倍，且暂停位置与体内观察到的相同。含有人类热休克因子1（HSF1）转录激活域的激活剂可增加这种暂停的通读。激活剂的最大效应要求该系统补充具有hSWI/SNF活性的组分，此前已证明该活性可改变核小体结构。在没有激活剂的情况下未观察到明显的通读，激活剂和hSWI/SNF组分均不影响裸DNA上的延伸；因此，这些结果表明，激活剂可通过降低核小体对转录延伸的抑制作用来增加启动子近端暂停的通读。
