A requirement for protein phosphorylation in regulating the meiotic and mitotic cell cycles in echinoderms☆

摘要

Populationsofhormone-stimulatedstarfishoocytesandfertilizedseaurchineggsundergosynchronousmeioticandmitoticdivisions.Wehavestudiedtherequirementforproteinphosphorylationduringtheseeventsbytestingtheeffectsof6-dimethylaminopurine(6-DMAP)upontheincorporationof[32P]orthophosphate.Itwasfoundthat6-DMAPblockedmeiosisreinitiationandearlycleavageandsimultaneouslyinhibitedproteinphosphorylation,withoutchangingtherateof[35S]methionineincorporationorpatternofproteinsynthesis.Theprotein,cyclin(54kDainstarfishand57kDainseaurchin),continuestobesynthesizedinthepresenceof6-DMAP.Thisproteinisdestroyedatfirstandsecondcellcycleswhen6-DMAPisadded30minfollowingfertilizationbutnotwhenthisdrugispresentbeforefertilization.Thus,cyclinbreakdowndoesnotdependonthecompletionofthenucleareventsofM-phase,anditstimeofbreakdownissetatanearlystepbetweenfertilizationandfirstcleavage.Usingtubulinimmunostaining,wefoundthat6-DMAPdidnotaffectthecorticalmicrotubulesandrestingfemalecentriolesofprophase-arrestedstarfishoocytes,whereasitinducedaprecociousdisappearanceofspindlefiberswhenappliedtohormone-stimulatedoocytes.Whileanearlyadditionof6-DMAPprecludednuclearbreakdownandspindleformationinbothsystems,alatetreatmentalwaysallowedchromosomeseparationandcentrioleseparation.Undertheseconditionspericentriolartubulinpersistedandcouldorganizenewspindlesaftertheinhibitorwasremoved.Itissuggestedthat(1)theassemblyofcorticalandcentriolar-associatedmicrotubulesisnotcontrolledbythesamefactorsasspindle-associatedtubulin;(2)specificproteinswhicharerequiredforthecelltoenterthefollowingM-phasecanbecomeoperativeonlyviaaprocessdependinguponproteinphosphorylation;(3)microtubule-associatedkinasesmayplayanimportantroleinMPFfunctionandspindledynamics.