Palmiter R D, Lee D C
J Biol Chem. 1980 Oct 25;255(20):9693-8.
Estrogen and progesterone markedly stimulate transcription of ovalbumin and conalbumin (transferrin) genes in chick oviduct as measured by hybridization of labeled RNA synthesized in isolated nuclei to immobilized plasmid DNA containing these gene sequences. Using this direct assay for specific gene transcription, we explored the basis of previous reports indicating that steroid hormones also cause changes in oviduct chromatin structure that can be detected by Escherichia coli RNA polymerase. We observed no effect of these hormones on the ability of E. coli RNA polymerase to transcribe specifically the conalbumin and ovalbumin genes 8 1/2 h after hormone administration when transcription of these genes by endogenous RNA polymerase was elevated 5- and 30-fold, respectively. Furthermore, we were unable to detect any significant effect of either of these hormones on the total number of E. coli RNA polymerase binding sites in oviduct nuclei or chromatin. In contrast, after several days of hormone administration, we detected an apparent preferential ovalbumin RNA synthesis by E. coli RNA polymerase and this effect could be transferred to unstimulated nuclei by a 0.35 M salt extract of active nuclei. However, further experiments revealed that this preferential ovalbumin RNA synthesis is an artifact produced by transcription from contaminating ovalbumin mRNA. We conclude that E. coli RNA polymerase does not recognize steroid hormone-induced changes in oviduct chromatin.
通过将在分离的细胞核中合成的标记RNA与含有这些基因序列的固定化质粒DNA杂交来测量,雌激素和孕酮能显著刺激鸡输卵管中卵清蛋白和伴清蛋白(转铁蛋白)基因的转录。利用这种对特定基因转录的直接检测方法,我们探究了先前报道的依据,这些报道表明类固醇激素也会引起输卵管染色质结构的变化,这种变化可用大肠杆菌RNA聚合酶检测到。当内源性RNA聚合酶对这些基因的转录分别提高了5倍和30倍时,我们观察到在激素给药8个半小时后,这些激素对大肠杆菌RNA聚合酶特异性转录伴清蛋白和卵清蛋白基因的能力没有影响。此外,我们无法检测到这些激素中的任何一种对输卵管细胞核或染色质中大肠杆菌RNA聚合酶结合位点总数有任何显著影响。相反,在给药几天后,我们检测到大肠杆菌RNA聚合酶明显优先合成卵清蛋白RNA,并且这种效应可以通过活性细胞核的0.35M盐提取物转移到未受刺激的细胞核中。然而,进一步的实验表明,这种优先合成卵清蛋白RNA是由污染的卵清蛋白mRNA转录产生的假象。我们得出结论,大肠杆菌RNA聚合酶不能识别类固醇激素诱导的输卵管染色质变化。