Lipovitellins derived from two forms of vitellogenin are differentially processed during oocyte maturation in haddock (Melanogrammus aeglefinus).
In the process of cloning vitellogenin (Vtg) cDNAs from haddock (Melanogrammus aeglefinus), two related, but distinct, mRNAs were identified. Full-length cDNA sequences were determined for both Vtg types (Had1 and Had2), and the deduced amino acid sequences were found to be 54% identical to each other and 48-58% identical to other teleost Vtgs. To investigate the expression of the two Vtg mRNAs, proteins from prehydrated oocytes and fertilized eggs were separated on SDS-polyacrylamide gels. Only a single lipovitellin I band was detected in each sample, and the egg lipovitellin I was smaller (97 vs. 110 kDa) than the oocyte protein, indicative of proteolytic processing during oocyte hydration. Mass spectrometric (MALDI-TOFMS and tandem mass spectrometry) analyses of tryptic fragments from the haddock oocyte and egg lipovitellin I revealed that the lipovitellin I from prehydrated oocytes contained tryptic fragments that matched the sequences of both types of Vtg, suggesting that there were two proteins in this band, while the egg lipovitellin I contained tryptic fragments that only matched the Had1 cDNA sequence, indicating that the Had2 lipovitellin had been degraded during hydration. Physiological data from haddock oocytes and eggs demonstrate that, as in other marine fish that spawn pelagic eggs, the free amino acid content increases during oocyte hydration and apparently contributes to hydration by driving the osmotic uptake of water. The correlation of the disappearance of one lipovitellin I with the increase of free amino acids in the oocyte suggests that this protein is a major source of the free amino acids for oocyte hydration.
Reith
M
M
NRC Institute for Marine Biosciences, Halifax, Nova Scotia B3H 3Z1, Canada. michael.reith@nrc.ca
Munholland
J
J
Kelly
J
J
Finn
R N
RN
Fyhn
H J
HJ
D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004527:Egg Proteins / Q000235:genetics / Q000378:metabolism; D015387:Egg Proteins, Dietary / Q000378:metabolism*; D005260:Female; D005399:Fishes / Q000378:metabolism*; D008969:Molecular Sequence Data; D009865:Oocytes / Q000254:growth & development* / Q000378:metabolism*; D011499:Protein Processing, Post-Translational / Q000235:genetics; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014819:Vitellogenins / Q000235:genetics / Q000378:metabolism*
D018076:DNA, Complementary; D004527:Egg Proteins; D015387:Egg Proteins, Dietary; D014819:Vitellogenins; C001549:lipovitellin