Chan L, Kohler P O, O'Malley B W
J Clin Invest. 1976 Mar;57(3):576-85. doi: 10.1172/JCI108313.
Ovalbumin messenger RNA (mRNAov) purified from hen oviduct was injected into Xenopus laevis oocytes. The oocytes were incubated in culture medium containing [3H]leucine. Analysis of the oocyte cytosol on Sephadex G-15O columns demonstrated a peak of radioactivity which cochromatographed with authentic ovalbumin. Radioactive protein contained in this peak was precipitated by ovalbumin antiserum, coelectrophoresed with ovalbumin on sodium dodecyl sulfate (SDS) and urea gels at pH 8.7, and eluted with the protein at the same pH (4.8) on CM-cellulose chromatography. Injection of increasing amounts of mRNAov was found to elicit a linear response in terms of ovalbumin synthesis. Moreover, there was linear incorporation of radioactivity into microinjected oocytes over a minimum period of 91 h. Less than 1 ng mRNAov was detected in this system. Ovalbumin mRNA activity was present in RNA preparations from chicks treated with estrogen but was undetectable in animals withdrawn from the hormone. This study constitutes an initial demonstration of a steroid hormone-induced alteration in mRNA population as assayed in intact viable heterologous cells.
从母鸡输卵管中纯化的卵清蛋白信使核糖核酸(mRNAov)被注射到非洲爪蟾卵母细胞中。将卵母细胞置于含有[3H]亮氨酸的培养基中培养。在葡聚糖凝胶G - 150柱上对卵母细胞胞质溶胶进行分析,结果显示放射性峰值与纯卵清蛋白共层析。该峰值中所含的放射性蛋白质可被卵清蛋白抗血清沉淀,在pH 8.7的十二烷基硫酸钠(SDS)和尿素凝胶上与卵清蛋白进行共电泳,并在相同pH(4.8)条件下于CM - 纤维素层析中与该蛋白质一同洗脱。研究发现,注射越来越多的mRNAov会引发卵清蛋白合成方面的线性反应。此外,在至少91小时的时间内,放射性物质线性掺入显微注射的卵母细胞中。在该系统中检测到的mRNAov少于1纳克。在用雌激素处理的小鸡的RNA制剂中存在卵清蛋白mRNA活性,但在摘除激素的动物中未检测到。这项研究首次证明了在完整有活力的异源细胞中检测到的甾体激素诱导的mRNA群体变化。