Regulation of Translation of Sea Urchin Egg mRNPs

这项建议的主要目的是阐明在海胆卵受精时控制储存的母体mRNAs激活的机制。这些RNA以翻译抑制的信使核糖核蛋白复合体(MRNP)的形式存储。细胞内钙或pH水平的变化或蛋白激酶C的激活以某种方式改变受精时卵子mRNPs的结构，导致它们的翻译激活。PI建议详细分析控制储存的母体mRNPs的抑制和去抑制的因素。参与翻译抑制的mPNP蛋白将通过比较纯化的非活性和活性的聚A(+)mRNPs的蛋白质模式来鉴定。此外，将从体外激活的鸡蛋mRNPs中分离出mRNP结合蛋白。针对这些mRNP掩蔽因子的抗体将用于监测受精时mRNP激活时mRNP蛋白的变化。蛋白激酶C的激活或细胞内钙离子的变化会导致鸡蛋mRNPs的暴露。这些变化可能会激活一连串的蛋白质磷酸化反应。PI将研究在活体和海胆体外翻译系统中磷酸化在激活卵子mRNPs中的潜在作用。

The broad objective of this proposal is to elucidate the mechanisms which control the activation of stored maternal mRNAs at fertilization in sea urchin eggs. These RNAs are stored as translationally repressed messenger ribonucleoprotein complexes (mRNPs). A change in intracellular Ca or pH levels or the activation of protein kinase C in some manner alters the structure of the egg mRNPs at fertilization, resulting in their translational activation. The PI proposes to analyze in detail the factors that control the repression and derepression of stored maternal mRNPs. mPNP proteins involved in translation repression will be identified by by comparing the protein patterns of purified inactive and active poly A (+) mRNPs. In addition, mRNP binding proteins will be isolated from in vitro activated egg mRNPs. Antibodies against these mRNP masking factors will be used to monitor changes in mRNP proteins upon mRNP activation at fertilization. Protein kinase C activation or intracellular calcium changes result in the unmasking of egg mRNPs. These changes may activate a cascade of protein phosphorylation reactions. The PI will examine the potential role of phosphorylation in the activation of egg mRNPs both in vivo and in a sea urchin in vitro translation system.