Gilmour D S, Dietz T J, Elgin S C
Department of Biology, Washington University, St. Louis, Missouri 63130.
Mol Cell Biol. 1988 Aug;8(8):3204-14. doi: 10.1128/mcb.8.8.3204-3214.1988.
We monitored protein-DNA interactions that occur on the hsp26, hsp70, histone H3, and histone H4 promoters in nuclear extracts derived from frozen Drosophila melanogaster embryos. All four of these promoters were found to be transcribed in vitro at comparable levels by extracts from both heat-shocked and non-heat-shocked embryos. Factors were detected in both types of extracts that block exonuclease digestion from a downstream site at ca. +35 and -20 base pairs from the start of transcription of all four of these promoters. In addition, factors in extracts from heat-shocked embryos blocked exonuclease digestion at sites flanking the heat shock consensus sequences of hsp26 and hsp70. Competition experiments indicated that common factors cause the +35 and -20 barriers on all four promoters in both extracts. The formation of the barriers at +35 and -20 required a TATA box but did not appear to require specific sequences downstream of +7. We suggest that the factors responsible for the +35 and -20 barriers are components whose association with the promoter precedes transcriptional activation.
我们监测了从冷冻的黑腹果蝇胚胎中提取的核提取物中,在hsp26、hsp70、组蛋白H3和组蛋白H4启动子上发生的蛋白质 - DNA相互作用。发现热激胚胎和非热激胚胎的提取物在体外均能以相当的水平转录这四个启动子。在这两种提取物中均检测到能阻止核酸外切酶从下游位点(约在所有这四个启动子转录起始点下游 +35和 -20碱基对处)进行消化的因子。此外,热激胚胎提取物中的因子在hsp26和hsp70热激共有序列侧翼的位点处阻止了核酸外切酶的消化。竞争实验表明,共同因子导致了两种提取物中所有四个启动子上的 +35和 -20屏障。在 +35和 -20处形成屏障需要TATA盒,但似乎不需要 +7下游的特定序列。我们认为,负责 +35和 -20屏障的因子是在转录激活之前与启动子结合的成分。