Ratnasabapathy R
Department of Pharmacology, Arnold and Marie Schwartz College of Pharmacy, Long Island University, Brooklyn, NY 11201 USA.
Cell Mol Biol Res. 1995;41(6):583-94.
Estrogen-mediated accumulation of the avian apolipoprotein (apo) II mRNA is in part due to its stabilization. To identify the biochemical activity responsible for this effect, radiolabeled, capped, and polyadenylated apoII mRNA was incubated in vitro in liver cytosolic extracts from roosters who received either estrogen (estrogen-treated extract) or the vehicle (control extract) parenterally. The mRNA was very stable in estrogen-treated extract but was rapidly degraded in control extract. The RNA was degraded predominantly by endonuclease rather than exonuclease activity. The addition of the estrogen-treated extract to the control extract prevented the degradation of the mRNA in trans. This biochemical activity was heat labile and was also destroyed by proteinase K but not by micrococcal nuclease, indicating that estrogen treatment resulted in the expression of a protein in the liver that stabilized the apoII mRNA by inhibiting its nucleolytic degradation. This mRNA stabilization factor was labile around 60 degrees C, whereas the RNase remained stable up to 80 degrees C. Studies on mRNA protein interaction showed that both control and estrogen-treated extracts contain mRNA-binding (mRNP) proteins that bind apoII mRNA. An increased binding to apoII mRNA by a subset of these proteins was observed with estrogen-treated extract as compared with the control extract. This activity, although it afforded complete protection from nucleolytic degradation to apoII and apo A1 mRNAs, appeared to provide less protection to mRNAs encoding chicken serum albumin and vitellogenin, suggesting differential stabilization of mRNAs. These studies indicate that a cytosolic mRNA-stabilization factor, providing apoII mRNA complete protection from nucleolytic degradation, is expressed in the avian liver upon estrogen treatment. This appears to be the first time that a biochemical activity responsible for hormone-mediated stabilization of mRNAs and estrogen induction of mRNA binding by specific mRNPs have been identified and partially characterized in vitro.
雌激素介导的禽类载脂蛋白（apo）II mRNA积累部分归因于其稳定性的提高。为了确定导致这种效应的生化活性，将放射性标记、加帽和聚腺苷酸化的apoII mRNA在体外与经腹腔注射雌激素的公鸡肝脏胞质提取物（雌激素处理提取物）或溶剂（对照提取物）一起孵育。该mRNA在雌激素处理提取物中非常稳定，但在对照提取物中迅速降解。RNA主要通过内切核酸酶而非外切核酸酶活性降解。将雌激素处理提取物添加到对照提取物中可防止mRNA在反式中降解。这种生化活性对热不稳定，也被蛋白酶K破坏，但不被微球菌核酸酶破坏，表明雌激素处理导致肝脏中一种蛋白质的表达，该蛋白质通过抑制其核酸降解来稳定apoII mRNA。这种mRNA稳定因子在60℃左右不稳定，而核糖核酸酶在高达80℃时仍保持稳定。对mRNA-蛋白质相互作用的研究表明，对照提取物和雌激素处理提取物都含有与apoII mRNA结合的mRNA结合（mRNP）蛋白。与对照提取物相比，雌激素处理提取物中观察到这些蛋白质的一个子集与apoII mRNA的结合增加。这种活性虽然能为apoII和apo A1 mRNA提供完全的核酸降解保护，但对编码鸡血清白蛋白和卵黄蛋白原的mRNA的保护作用似乎较小，表明mRNA的稳定存在差异。这些研究表明，一种胞质mRNA稳定因子在雌激素处理后在禽类肝脏中表达，该因子能为apoII mRNA提供完全的核酸降解保护。这似乎是首次在体外鉴定并部分表征了负责激素介导的mRNA稳定和雌激素诱导特定mRNP与mRNA结合的生化活性。
