Audet R G, Goodchild J, Richter J D
Dev Biol. 1987 May;121(1):58-68. doi: 10.1016/0012-1606(87)90138-2.
The injection of heterologous mRNA into fully grown Xenopus oocytes results not only in the synthesis of the heterologous protein but also in a reciprocal decrease in the synthesis of endogenous proteins. This indicates that injected and endogenous mRNAs compete for some component which is rate-limiting for translation in oocytes. We have attempted to identify this rate-limiting translational component. We find that heterologous and homologous polysomes compete with endogenous mRNAs as effectively as naked mRNA, indicating that polysomes do not contain detectable levels of the rate-limiting factor. In addition, we have used micrococcal nuclease digestion and a mRNA-specific oligonucleotide to destroy the mRNA component of polysomes. The remaining polysome factors, when injected into oocytes, failed to stimulate translation. When several eukaryotic translation initiation factors were injected into oocytes, initiation factor 4A consistently increased general oocyte protein synthesis by about twofold. It is possible that the availability of eIF-4A in oocytes is a key factor in limiting the overall rate of protein synthesis.
将异源mRNA注射到完全成熟的非洲爪蟾卵母细胞中,不仅会导致异源蛋白质的合成,还会使内源性蛋白质的合成相应减少。这表明注射的mRNA和内源性mRNA竞争某种对卵母细胞翻译起限速作用的成分。我们试图鉴定这种限速翻译成分。我们发现异源和同源多核糖体与内源性mRNA的竞争效果与裸mRNA一样有效,这表明多核糖体不含可检测水平的限速因子。此外,我们使用微球菌核酸酶消化和mRNA特异性寡核苷酸来破坏多核糖体的mRNA成分。将剩余的多核糖体因子注射到卵母细胞中时,未能刺激翻译。当将几种真核翻译起始因子注射到卵母细胞中时,起始因子4A始终使卵母细胞总体蛋白质合成增加约两倍。卵母细胞中eIF-4A的可用性可能是限制蛋白质合成总体速率的关键因素。