Sea urchin fertilization envelope: Uncoupling of cortical granule exocytosis from envelope assembly and isolation of an envelope intermediate from Strongylocentrotus purpuratus embryos☆

[摘要] Theseaurchinfertilizationenvelope(FE)isanextraembryoniccoatwhichdevelopsfromtheeggvitellineenvelope(VE)andthesecretedparacrystallineproteinfractionofthecorticalgranulesatfertilization.TheFEundergoesfurtherdevelopmentalchangespostinseminationwhicharecharacterizedbychangesinenvelopepermeability,solubilityinreducinganddenaturingsolvents,andmorphology.WehavedevelopedaproceduretouncouplecorticalgranuleexocytosisfromassemblyoftheparacrystallineproteinfractionontotheVEtemplate.EggsuspensionswereinseminatedinnormalseawateranddilutedintoCa2+-andMg2+-freeseawaterat15secpostinsemination.Phase-contrastandelectronmicroscopicobservationsshowedthattheembryosformedanormallyelevated,extremelythinenvelopethroughwhichthecorticalgranuleexudatepermeated.Secretionstudiesshowedthateggswhichweredilutedintodivalention-freeseawaterpostinseminationsecretedasmuchproteinintothesurroundingseawateraseggswhichhadtheirVEsremovedpriortotheexperiment.Wehavetermedtheenvelopeelevatedindivalention-freeseawatertheVE∗andwebelievethatitistheVEstructuralcomponentoftheFEbasedonitsthicknessandmorphology.VE∗swereisolatedbygentlephysicalmeansandthepreparationsappearedtobegreaterthan80%purebasedonradioactivemixingexperimentsandonmalatedehydrogenaseandglucose-6-phosphatedehydrogenasemarkerstudies.VE∗swereatleast80%solublebasedonextractionofradioiodinatedpreparationswithreducinganddenaturingsolvents.Sodiumdodecylsulfate-polyacrylamidegelelectrophoresisofVE∗sshowedeightmajorpolypeptideswhichrangedfrom30,500to270,000inmolecularweight.

[学科分类] 生物科学（综合）