Daar I O, Maquat L E
Department of Human Genetics, Roswell Park Memorial Institute, Buffalo, New York 14263.
Mol Cell Biol. 1988 Feb;8(2):802-13. doi: 10.1128/mcb.8.2.802-813.1988.
We characterized an anemia-inducing mutation in the human gene for triosephosphate isomerase (TPI) that resulted in the production of prematurely terminated protein and mRNA with a reduced cytoplasmic half-life. The mutation converted a CGA arginine codon to a TGA nonsense codon and generated a protein of 188 amino acids, instead of the usual 248 amino acids. To determine how mRNA primary structure and translation influence mRNA stability, in vitro-mutagenized TPI alleles were introduced into cultured L cells and analyzed for their effect on TPI RNA metabolism. Results indicated that mRNA stability is decreased by all nonsense and frameshift mutations. To determine the relative contribution of the changes in mRNA structure and translation to the altered half-life, the effects of individual mutations were compared with the effects of second-site reversions that restored translation termination to normal. All mutations that resulted in premature translation termination reduced the mRNA half-life solely or mainly by altering the length of the mRNA that was translated. The only mutation that altered translation termination and that reduced the mRNA half-life mainly by affecting the mRNA structure was an insertion that shifted termination to a position downstream of the normal stop codon.
我们鉴定了人类磷酸丙糖异构酶（TPI）基因中的一种导致贫血的突变，该突变导致产生过早终止的蛋白质和细胞质半衰期缩短的mRNA。该突变将CGA精氨酸密码子转换为TGA无义密码子，产生了一个188个氨基酸的蛋白质，而不是通常的248个氨基酸。为了确定mRNA一级结构和翻译如何影响mRNA稳定性，将体外诱变的TPI等位基因导入培养的L细胞，并分析其对TPI RNA代谢的影响。结果表明，所有无义突变和移码突变均会降低mRNA稳定性。为了确定mRNA结构和翻译变化对改变的半衰期的相对贡献，将单个突变的影响与恢复翻译终止正常的第二位点回复突变的影响进行了比较。所有导致过早翻译终止的突变均仅或主要通过改变翻译的mRNA长度来降低mRNA半衰期。唯一改变翻译终止并主要通过影响mRNA结构而降低mRNA半衰期的突变是一个插入突变，该突变将终止位点转移到正常终止密码子下游的位置。
