Modification of ribosomal proteins in sea urchin eggs following fertilization

Abstract

Analysis of the ribosomal proteins in sea urchin eggs by two-dimensional polyacrylamide gel electrophoresis revealed postfertilization changes in the proteins of both the small and the large subunits. Five egg-ribosomal proteins (S7, S16, S19, L19, L31) appeared to undergo rapid changes to the corresponding embryo-specific proteins. These changes were completed within 30 min after fertilization, and identical electrophoretic patterns were observed among the different developmental stages of embryos. One of the five proteins, S7, showed an increase in the phosphorylated form. The remainder showed qualitative shifts to the corresponding embryo-specific proteins ; however, peptide map analyses revealed the existence of common structural units between the corresponding proteins.These modifications were observed in the three species of sea urchin studied (Pseudocentrotu,y depressus, Hemicentrotus pulcherrirnus and Anthocidaris crassispina), except in the case of one protein (L31). Purification of ribosomes by different procedures based on high-salt treatment gave the same results with respect to the egg-specific and embryo-specific proteins.The rapid and large increase in protein synthesis in sea urchin eggs following fertilization is thought to be due to an increase in the availability of maternal mRNA [l -31. Using techniques both in vivo and in vitro, kinetic analyses of a number of components of the translational machinery have been done to find out the mechanism of the 'unmasking' and active utilization of the stored mRNA. However, since numerous parameters are varied under the conditions of cellfree translation systems, there have been conflicting reports concerning the translational activity of ribosomes isolated from eggs and embryos [4-61 and the template activity of masked maternal mRNAs [7,8].Although data on the structural components of ribosomes of sea urchin eggs and embryos are critical for investigating the control mechanism of protein synthesis, very little is known about these subjects. Our previous papers reported the characterization [9] and comparison of species specificity [lo] of the ribosomal proteins of sea urchin eggs. The present study compares the ribosomal proteins of eggs with those of embryos using two-dimensional polyacrylamide gel electrophoresis.To identify the essential variables between egg and embryo ribosomes, we analyzed the ribosomes from (a) unfertilized eggs and embryos at various stages of development, (b) mixtures of unfertilized and fertilized eggs, and (c) three species of sea urchin which have been found to exhibit speciesspecificity in the 60s ribosomal proteins [lo]. We also adopted the fingerprint analysis of tryptic peptides of 'Z51-labeled polypeptides to prove protein identity. EXPERIMENTAL PROCEDURES Eggs and embryosGametes from three species of sea urchin, Pseudocentrotus depressus, Hemicentrotus pulcherrimus and Anthociduris crasThis is the third paper of a series; the first two papers appeared Abbreuiations. SDS, sodium dodecyl sulfate ; mRNPs, messenger el...

Identification and comparative proteomic study of quail and duck egg white protein using 2-dimensional gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry analysis

“…This protein was identified for the first time in avian egg white. It has been reported that egg-ribosomal proteins appeared to undergo rapid changes after fertilization, which may be causally related to the acceleration of protein synthesis following fertilization (Takeshima and Nakano, 1983 ). Another Coturnix coturnix specific protein was identified as Lipocalin Q83 (spot A29), which is a lipocalin family member.…”

Intracellular sites of the synthesis of sweet potato mitochondrial F1ATPase subunits

“…The band of the 6-or 6'-subunit after the SDSpolyacrylamide gel electrophoresis for isolation of the subunit was treated with [125I]sodium iodide and then with trypsin by the method of Takeshima & Nakano (17). The labeled digest was first subjected to electrophoresis on a silica gel thinlayer plate (20×20 cm, Silica Gel 60, Merck), then chromatographed on the plate with n-butyl alcohol/ pyridine/acetic acid/water (32.5:25:5:20) as the developing solvent as described previously (17).…”

“…The labeled digest was first subjected to electrophoresis on a silica gel thinlayer plate (20×20 cm, Silica Gel 60, Merck), then chromatographed on the plate with n-butyl alcohol/ pyridine/acetic acid/water (32.5:25:5:20) as the developing solvent as described previously (17). The plate was exposed to Fuji RXO-G film with an intensifying screen (Grenex, Fuji) at -7 0°C for 2 days.…”

Separate ribosomal pools in sea urchin embryos: ammonia activates a movement between pools

“…(c) The initial activity of monoribosomes from unfertilized eggs is lower, in vitro, than that of ribosomes derived from blástula polyribosomes . Observations on the phosphorylation state of the S6 ribosomal protein, however, cannot yet be correlated with ribosomal activity (Ballinger & Hunt, 1981; Ward et al, 1983;Takeshima & Nakano, 1983;Ballinger et al, 1984).…”