A basic helix-loop-helix transcription factor,figα,is one of the earliest marker genes of oocyte differentiation in vertebrates.In the present study,we made figαknockout medaka by CRISPR/Cas9,expecting aborted progr...A basic helix-loop-helix transcription factor,figα,is one of the earliest marker genes of oocyte differentiation in vertebrates.In the present study,we made figαknockout medaka by CRISPR/Cas9,expecting aborted progress of oogenesis,to see if differentiation of somatic ovarian tissues is affected.Figαknockout male gonads differentiated normally into testes with functional sperm.The females,on the other hand,were sterile;there are oocytes only up to pachytene.No growing oocytes in diplotene were found.The phenotype was already apparent at 10 days after hatching,when diplotene oocytes start to develop in the control.Furthermore,several putative target genes of figαwere not expressed in the mutant female gonads.Previous studies showed that medaka lacking germ cells have morphologically abnormal gonads and female to male sex reversal occurred.Figαknockout female gonads differentiated morphologically into ovaries,suggesting that in medaka,figαknockout prevents oogenesis progress into diplotene stage,and ovarian differentiation do not need developed oocytes beyond pachytene stage.This ovarian phenotype reminded us of"pachytene checkpoint",which blocks meiotic progress into diplotene when chromosomal abnormalities are present.Therefore,we further analyzed phenotype of oocytes in the mutant ovaries.The number of oocytes at each meiotic stage suggested that oogenesis normally proceeded up to pachytene stage.The number of apoptotic oocytes in the knockout ovaries was small.Finally,we did not detect any abnormal pairings or unrepaired double-strand breaks by immunostaining.Taken together,figαknockout likely stop oocyte growth at or around pachytene via unknown mechanisms other than"pachytene checkpoint"and arrested oocytes remained in the ovary.This is in contrast to figαknockout mice where all oocytes underwent apoptosis quickly after birth.展开更多
基金supported in part by a Grant-in-Aid for Scientific Research from JSPS(15K07127 to AK and MK).
文摘A basic helix-loop-helix transcription factor,figα,is one of the earliest marker genes of oocyte differentiation in vertebrates.In the present study,we made figαknockout medaka by CRISPR/Cas9,expecting aborted progress of oogenesis,to see if differentiation of somatic ovarian tissues is affected.Figαknockout male gonads differentiated normally into testes with functional sperm.The females,on the other hand,were sterile;there are oocytes only up to pachytene.No growing oocytes in diplotene were found.The phenotype was already apparent at 10 days after hatching,when diplotene oocytes start to develop in the control.Furthermore,several putative target genes of figαwere not expressed in the mutant female gonads.Previous studies showed that medaka lacking germ cells have morphologically abnormal gonads and female to male sex reversal occurred.Figαknockout female gonads differentiated morphologically into ovaries,suggesting that in medaka,figαknockout prevents oogenesis progress into diplotene stage,and ovarian differentiation do not need developed oocytes beyond pachytene stage.This ovarian phenotype reminded us of"pachytene checkpoint",which blocks meiotic progress into diplotene when chromosomal abnormalities are present.Therefore,we further analyzed phenotype of oocytes in the mutant ovaries.The number of oocytes at each meiotic stage suggested that oogenesis normally proceeded up to pachytene stage.The number of apoptotic oocytes in the knockout ovaries was small.Finally,we did not detect any abnormal pairings or unrepaired double-strand breaks by immunostaining.Taken together,figαknockout likely stop oocyte growth at or around pachytene via unknown mechanisms other than"pachytene checkpoint"and arrested oocytes remained in the ovary.This is in contrast to figαknockout mice where all oocytes underwent apoptosis quickly after birth.
