IN-VITRO STUDY OF VITELLOGENESIS IN THE BLUE-CRAB (CALLINECTES-SAPIDUS) - SITE AND CONTROL OF VITELLIN SYNTHESIS

To determine the site of vitellin (Vn) synthesis in the blue crab, Cal linectes sapidus, tissues (ovary, hepatopancreas, and gill) from adult female crabs were incubated in vitro with [S-35]methionine. Tissue ex tracts were separated by sodium dodecyl sulfate-polyacrylamide gel ele ctrophoresis and radiolabeled proteins were visualized by autoradiogra phy. Four radiolabeled ovarian proteins (188, 168, 109, and 86 kDa) co migrated with the subunits of Vn. A Western blot analysis confirmed th at these ovarian proteins were the Vn subunits. Proteins in the hepato pancreas and gill were also radiolabeled but were not Vn-immunoreactiv e. In vitro incorporation of [S-35]methionine into ovarian proteins wa s inhibited by an extract of eyestalks; the inhibition was dose-depend ent and specific. Autoradiography of the electrophoretically separated ovarian proteins revealed that the eyestalk extract inhibited incorpo ration of [S-35]methionine into numerous proteins, including the Vn su bunits. These results suggest that the ovary is a site of Vn synthesis and that the eyestalks contain a factor or factors that directly inhi bit the synthesis of Vn and other ovarian proteins. It is anticipated that the suppression of ovarian protein synthesis will be a useful bio assay for blue crab vitellogenesis-inhibiting hormone. (C) 1995 Wiley- Liss, Inc.