Shopland L S, Hirayoshi K, Fernandes M, Lis J T
Section of Biochemistry, Molecular and Cell Biology, Cornell University, Ithaca, New York 14853, USA.
Genes Dev. 1995 Nov 15;9(22):2756-69. doi: 10.1101/gad.9.22.2756.
Chromatin structure can modulate gene expression by limiting transcription factor access to gene promoters. We examined sequence elements of the Drosophila hsp70 promoter for their ability to facilitate the binding of the transcription factor, heat shock factor (HSF), to chromatin. We assayed HSF binding to various transgenic heat shock promoters in situ by measuring amounts of fluorescence at transgenic loci of polytene chromosomes that were stained with an HSF antibody. We found three promoter sequences that influence the access of HSF to its binding sites: the GAGA element, sequences surrounding the transcription start site, and a region in the leader of hsp70 where RNA polymerase II arrests during early elongation. The GAGA element has been shown previously to disrupt nucleosome structure. Because the two other critical regions include sequences that are required for stable binding of TFIID in vitro, we examined the in vivo occupancy of the TATA elements in the transgenic promoters. We found that TATA occupancy correlated with HSF binding for some promoters. However, in all cases HSF accessibility correlated with the presence of paused RNA polymerase II. We propose that a complex promoter architecture is established by multiple interdependent factors, including GAGA factor, TFIID, and RNA polymerase II, and that this structure is critical for HSF binding in vivo.
染色质结构可通过限制转录因子与基因启动子的结合来调节基因表达。我们研究了果蝇hsp70启动子的序列元件促进转录因子热休克因子（HSF）与染色质结合的能力。我们通过测量用HSF抗体染色的多线染色体转基因位点处的荧光量，原位检测HSF与各种转基因热休克启动子的结合。我们发现了三个影响HSF与其结合位点结合的启动子序列：GAGA元件、转录起始位点周围的序列，以及hsp70前导序列中RNA聚合酶II在早期延伸过程中停滞的区域。先前已证明GAGA元件可破坏核小体结构。由于其他两个关键区域包含体外TFIID稳定结合所需的序列，我们研究了转基因启动子中TATA元件的体内占据情况。我们发现，对于某些启动子，TATA占据与HSF结合相关。然而，在所有情况下，HSF的可及性都与暂停的RNA聚合酶II的存在相关。我们提出，由多个相互依赖的因子，包括GAGA因子、TFIID和RNA聚合酶II，建立了一个复杂的启动子结构，并且这种结构对于体内HSF的结合至关重要。
