Sandler P, Weisblum B
Department of Genetics, University of Wisconsin, Madison 53706.
J Mol Biol. 1988 Oct 20;203(4):905-15. doi: 10.1016/0022-2836(88)90116-7.
Erythromycin-induced stabilization of ermA mRNA was studied in Staphylococcus aureus, its original host background, and in Bacillus subtilis, subcloned on plasmid vectors. By RNA blot analysis it was shown that 40 nM-erythromycin specifically increased the chemical half-life of ermA mRNA from 2.5 to 17.5 minutes whereas the half-life of cat-86 mRNA was not increased by erythromycin. While expression of ermA has been shown to be induced by erythromycin at the level of translation, our studies with three ermA constitutive mutants demonstrated that mRNA stabilization in growing cells occurred independently of induced gene expression, suggesting that the stabilized mRNA was not functional for protein synthesis. Studies of ermA/lacZ fusions demonstrated that the 5' end of the mRNA was sufficient to confer stabilization. Translation of specific amino acid codons in a leader peptide located at the extreme 5' end of the mRNA was required for the erythromycin-induced stabilization as a frameshift mutation introduced into the leader peptide determinant abolished stabilization. By S1 mapping, no differences were detected in the length of the 5' or 3' end of ermA mRNA with the addition of erythromycin, indicating that the stabilized transcript was not processed at its ends.
在金黄色葡萄球菌（其原始宿主背景）以及在枯草芽孢杆菌（亚克隆于质粒载体上）中研究了红霉素诱导的ermA mRNA稳定性。通过RNA印迹分析表明，40 nM的红霉素特异性地将ermA mRNA的化学半衰期从2.5分钟提高到17.5分钟，而cat-86 mRNA的半衰期未被红霉素提高。虽然已表明ermA的表达在翻译水平上由红霉素诱导，但我们对三个ermA组成型突变体的研究表明，生长细胞中的mRNA稳定性独立于诱导的基因表达而发生，这表明稳定的mRNA对蛋白质合成无功能。对ermA/lacZ融合体的研究表明，mRNA的5'端足以赋予稳定性。位于mRNA最5'端的前导肽中特定氨基酸密码子的翻译是红霉素诱导稳定性所必需的，因为引入前导肽决定簇的移码突变消除了稳定性。通过S1作图，添加红霉素后ermA mRNA的5'或3'端长度未检测到差异，表明稳定的转录本在其末端未被加工。
