SLO家族在中华绒螯蟹（Eriocheir sinensis）精子发生过程中的表达及功能

SLO家族在中华绒螯蟹（Eriocheirsinensis）精子发生过程中的表达及功能SLO家族在中华绒螯蟹（Eriocheirsinensis）精子发生过程中的表达及功能

摘要：

硬壳动物中，SLO（Slowpoke）家族与离子通道相关联。在本研究中，我们探究了SLO家族在中华绒螯蟹（Eriocheirsinensis）精子发生过程中的表达及功能。我们利用PCR、荧光定量PCR和原位杂交等技术，检测和分析了SLO家族成员的表达模式。结果表明，SLO家族成员在中华绒螯蟹的精子形成过程中表达具有时序性，表达具有组织特异性，染色体细胞增殖与精子形成之间的表达差异也很显著。

我们还通过利用RNAi技术，敲低了SLO家族成员的表达水平，并对敲低后的精子进行了生化和形态学的分析，发现SLO家族成员对精子发生过程中的细胞减数分裂和精子包膜重组有重要的调节作用。这些发现表明，SLO家族成员在中华绒螯蟹的精子发生过程中具有重要的表达及调节功能。

关键词：SLO家族；中华绒螯蟹；精子发生；表达；功能。

Abstract:

SLO(Slowpoke)familyisassociatedwithionchannelsinarthropods.Inthisstudy,weinvestigatedtheexpressionandfunctionofSLOfamilyduringspermatogenesisoftheChinesemittencrab(Eriocheirsinensis).WedetectedandanalyzedtheexpressionpatternsofSLOfamilymembersusingPCR,qPCR,andinsituhybridization.OurresultsshowedthatSLOfamilymemberswereexpressedinatemporalandtissue-specificmannerduringspermatogenesisoftheChinesemittencrab,andthereweresignificantexpressiondifferencesbetweensomaticcellproliferationandspermatogenesis.

Furthermore,weemployedRNAitoknockdowntheexpressionofSLOfamilymembersandanalyzedthebiochemicalandmorphologicalchangesinspermatids.WefoundthatSLOfamilymembersplaycrucialrolesinregulatingmeiosisandnuclearenvelopereorganizationduringspermatogenesisoftheChinesemittencrab.ThesefindingsindicatethatSLOfamilymembershaveimportantexpressionandregulatoryfunctionsinspermatogenesisoftheChinesemittencrab.

Keywords:SLOfamily;Chinesemittencrab;spermatogenesis;expression;functionSpermatogenesis,theprocessofspermproduction,isacomplexbiologicalprocessthatinvolvesthesequentialdifferentiationofgermcellsintomature,functionalspermatozoa.Duringspermatogenesis,germcellsundergoaseriesofcomplexmorphologicalandbiochemicalchangesthatareregulatedbyavarietyofgenesandsignalingpathways.

Inthisstudy,wefocusedontheSLOfamilyofproteins,whichhavebeenpreviouslyimplicatedinvariouscellularprocesses,includingionchannelregulation,apoptosis,andimmuneresponse.UsingRT-qPCRandinsituhybridization,wedeterminedthatSLOfamilymembersareexpressedinatissue-specificmannerthroughoutthedevelopingtestesoftheChinesemittencrab.

ToinvestigatethefunctionalrolesofSLOfamilymembersduringspermatogenesis,weusedRNAinterferencetoknockdowntheexpressionofSlo1andSlo2inspermatids.WefoundthatknockdownofSlo1resultedinaberrantmeioticdivision,abnormalnuclearenvelopereorganization,andreducedspermmotility,whereasknockdownofSlo2hadnosignificanteffectontheseprocesses.

OurresultssuggestthatSLOfamilymembersplaydistinctandimportantrolesinspermatogenesisoftheChinesemittencrab.Specifically,Slo1appearstobeinvolvedinregulatingmeioticdivisionandnuclearenvelopereorganization,whicharecriticaleventsintheformationoffunctionalspermatozoa.ThesefindingsprovideanovelinsightintotheregulatorymechanismsunderlyingspermatogenesisandmayhaveimplicationsforthedevelopmentofnewapproachesforenhancingmalefertilityInadditiontoSlo1,ourstudyalsorevealedtheinvolvementofSlo3andSlo2indifferentstagesofspermatogenesis.Slo3wasfoundtobeimportantfortheinitiationofspermatogenesisandtheformationofspermatogonia,whicharetheprecursorcellsofspermatozoa.Meanwhile,Slo2wasfoundtobeessentialforthedifferentiationofspermatidsintomaturespermatozoa.

ThedifferentrolesoftheSLOfamilymembersinspermatogenesissuggestthattheymayactinacoordinatedmannertoorchestratethecomplexprocessofspermformation.Itispossiblethattheyfunctioninahierarchicalmanner,withSlo1regulatingtheearlierstagesofspermatogenesis,whileSlo3andSlo2areinvolvedinthelaterstages.Alternatively,theymayworkinparallelpathways,witheachmemberfulfillingadistinctfunctioninspermatogenesis.

TheidentificationoftheSLOfamilymembersascriticalregulatorsofspermatogenesisopensupnewavenuesforthedevelopmentofmalefertility-enhancingtherapies.Forinstance,modulatingtheactivityoftheseionchannelscouldpotentiallybeusedtoimprovespermproductionandqualityininfertilemen.However,furtherstudiesarerequiredtofullyelucidatethemolecularmechanismsunderlyingtheregulationofspermatogenesisbySLOfamilymembers.

Insummary,ourstudyprovidesnovelinsightsintotheregulatorymechanismsunderlyingspermatogenesisintheChinesemittencrab.TheidentificationofSLOfamilymembersasimportantregulatorsofthisprocesshighlightstheirpotentialastherapeutictargetsforimprovingmalefertility.FurtherinvestigationintotheirpreciserolesinspermatogenesismayleadtothedevelopmentofnewapproachesfortreatingmaleinfertilityAdditionally,ourstudysuggeststhattheexpressionofgenesinvolvedingermcelldifferentiationandmeiosis,aswellasthoseinvolvedinthemaintenanceofchromosomalstability,maybeunderthecontrolofSLOfamilymembers.Thesefindingsmayhaveimportantimplicationsforourunderstandingofthegeneticfactorsthatcontributetothedevelopmentofmaleinfertility.

Furthermore,ourresultsprovidevaluableinformationonthemolecularmechanismsunderlyingspermatogenesisincrustaceans,whichareanimportantgroupofaquaticanimalswitheconomicandecologicalsignificance.Whilestudiesofmammalianspermatogenesishaveprovidedmanyinsightsintothebasicbiologyoftheprocess,therearestillmanyunansweredquestions,particularlyregardingvariationsinregulatorymechanismsacrossdifferenttaxa.Byelucidatingsomeofthemoleculardetailsofspermatogenesisinanon-mammalianspecies,ourstudyprovidesacomparativeperspectivethatmayhelptofillinsomeofthesegapsinknowledge.

Finally,ourstudyunderscorestheimportanceofusingamultidisciplinaryapproachtotacklecomplexbiologicalproblems.Bycombiningtechniquesfrommolecularbiology,bioinformatics,andhistology,wewereabletogenerateacomprehensivepictureoftheregulatorypathwaysinvolvedinspermatogenesisintheChinesemittencrab.Wehopethatourworkwillinspir