Yolk protein expression in the green crab, Carcinus maenas

Mature oocytes from adult female green crabs (Carcinus maenas) were homogenized and an abundant protein peak (∼ 315 kDa) was purified by gel filtration chromatography. The protein peak was resolved into two bands (74 and 106 kDa) on SDS-PAGE. The bands were digested with trypsin and the tryptic peptides were sequenced by liquid chromatography-mass spectroscopy. Two peptides from the 74 kDa band were homologous to regions in other crustacean yolk proteins. Degenerate primers based on these sequences amplified a 545-bp cDNA fragment. The conceptual translation of this fragment showed a close relationship (∼ 60% identity and ∼ 75% similarity) to vitellogenins (Vgs) found in other crustaceans. Gene-specific primers for Vg showed that its mRNA was present in the ovary and hepatopancreas (HP). Northern blots detected a single 8.6 kb transcript in both tissues. Quantitative real-time PCR indicated that ovarian levels of Vg-mRNA were highest at stage II vitellogenesis, rising 2.4-fold from previtellogenic animals and then dropping over 10-fold in stage III animals. Vg-mRNA levels in the HP rose 60-fold between previtellogenesis and stage I vitellogenesis, remaining elevated through stages II and III. These results indicate that the ovary and the HP are both sites of Vg synthesis in the green crab but the HP appears to be a much more important synthetic site.