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The general metabolic activation of the sea urchin egg at fertilization is dependent on a release of intracellular stores of calcium and the subsequent transient elevation of intracellular Ca2+ (refs 1--3). However, this elevation does not by itself lead to increased macromolecular synthesis and development but initiates steps which result in a long-term elevation of intracellular pH (refs 4--6). Among the developmental processes dependent on the elevation of intracellular pH is the large acceleration in the rate of protein synthesis at fertilization. Weak penetrating bases such as ammonia can be used to mimic the processes resulting in an increase in intracellular pH and so show the corresponding increases in protein synthesis rate. Conversely, it is possible to demonstrate a gradual but complete shut down of protein synthesis if the intracellular pH is reduced to the unfertilized level with penetrating weak acids. However, the rate of protein synthesis in ammonia-activated eggs lags behind that of fertilized controls even though ammonia activation can result in an intracellular pH increase greater than occurs in the fertilized egg. This result has led to the suggestion that factors other than intracellular pH may be regulating protein synthesis following fertilization. To investigate the possibility that the Ca2+ transient may have such a role, we measured the rate of amino acid incorporation in eggs that were activated in various ionic conditions which enabled the effects of Ca2+ and pH changes to be studied separately. Our results, reported here, show that if intracellular pH is elevated, increases in intracellular Ca2+ play an additional part in the activation of protein synthesis at fertilization.

Fertilization of sea urchin eggs results in a large increase in the rate of protein synthesis which is mediated by the translation of stored maternal mRNA. The masked message hypothesis suggests that messenger ribonucleoprotein particles (mRNPs) from unfertilized eggs are translationally inactive and that fertilization results in alterations of the mRNPs such that they become translationaHy active. Previous workers have isolated egg mRNPs by sucrose gradient centrifugation and have assayed their translational activity in heterologous cell-free systems. The conflicting results they obtained are probably due to the sensitivity of mRNPs to artifactual activation and inactivation. Previously, we demonstrated that unfractionated mRNPs in a sea urchin cell-free translation system were translationally inactive. Now, using large-pore gel filtration chromatography, we partially purified egg mRNPs while retaining their translationally repressed state. Polysomal mRNPs from fertilized eggs isolated under the same conditions were translationally active. The changes in the pattern of proteins synthesized by fractionated unfertilized and fertilized mRNPs in vitro were similar to those changes observed in vivo. Treatment of egg mRNPs with buffers containing high salt and EDTA, followed by rechromatography, resulted in the activation of the mRNPs and the release of an inhibitor of translation from the mRNPs. Analysis of the inhibitory fraction on one-dimensional sodium dodecyl sulfate gels indicated that this fraction contains a complex set of proteins, several of which were released from high-salt-EDTA-activated mRNPs and not from inactive low-salt control mRNPs. One of the released proteins may be responsible for the repression of egg mRNPs in vitro and be involved in the unmasking of mRNPs at fertilization.One of the major changes that occurs upon activation of a variety of cell types is an increase in the overall rate of protein synthesis. This increase is largely mediated by the regulation, at the translational level, of mRNAs stored in these quiescent cells. Examples of this type of translational control include the serum stimulation of resting fibroblasts, fertilization of various invertebrate and vertebrate oocytes, and the activation of T lymphocytes (30; for reviews, see references 18 and 26).It appears that many of the basic mechanisms that control the activation of protein synthesis in these various cell types occur in an amplified or exaggerated form after fertilization of sea urchin eggs. The increase in intracellular calcium and pH that follows fertilization results in the mobilization of maternal mRNAs into polysomes (10,14,36). These mRNAs were synthesized during oogenesis and stored in the unfertilized egg as messenger ribonucleoprotein particles (mRNPs). The specific regulatory pathways by which changes in intracellular ion concentrations result in the mobilization of these mRNPs into polysomes are not yet known.The increase in the rate of protein synthesis after fertilization is mainly due to an increase in t...

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