vsx1(visual system homeobox-1),a homeobox gene originally identified from an adult goldfish retinal cDNA library,has been shown to regulate retina progenitor proliferation,differentiation and functional maintenance of bipolar cells in vertebrates.However,in all the examined vertebrate species,vsx1 transcripts can be also detected at the early developmental stage,suggesting that it may play an important role in regulating early embryogenesis as well.Here,we investigated the function of vsx1 in early embryogenesis of goldfish(Carassius auratus) with both overexpression and gene knockdown approaches.It was found that vsx1 overexpression specifically blocked dorsal midline structure for-mation and vsx1 knockdown led to disorganized dorsal midline structure.Whole-mount in situ hy-bridization revealed that the midline expression of ntl,a key regulatory gene for chordamesoderm,was repressed by vsx1 overexpression but enhanced in the vsx1 knockdown.Furthermore,VSX1 protein could bind ntl promoter directly and was sufficient to inhibit ntl promoter-driven reporter gene green fluorescence protein transcription.Together,these results suggested that vsx1 may act to repress ec-topic expression of ntl in neural progenitor cells to ensure neural tube development in a spatially coordinated pattern during early embryogenesis.
LIU ZhengHuaCHEN JinHuiMA ShanShanTONG YingLUO Chen
In mammals,genomic imprinting confers developmental asymmetry and complementation on the parental genomes and makes both parental genomes essential for complete development.Genomic imprinting is,therefore,the first regulatory step of genome-wide gene expression of embryogenesis and thought to be the epigenetic foundation of bisexual reproduction.However,how the genomic imprinting is originated,established and maintained during vertebrate evolution remains unknown.Because no endogenous imprinting gene has been identified in non-mammalian vertebrates,genomic imprinting is thought to be a unique evolutionary event of mammals.Here,in order to study the evolutionary origin of genomic imprinting in vertebrates,we examined whether parent-specific methylation occurred in the teleost homologue of mammalian imprinting gene during gametogenesis.Bisulfate sequencing analy-sis showed that,as mammalian Igf2 CpG island,goldfish Igf2 CpG island was a parental differentially methylated region(DMR) that was hypermethylated in sperm but unmethylated in eggs.Unlike mam-malian imprinting gene DMR,however,the parent-specific methylation pattern of goldfish Igf2 DMR was not maintained during embryogenesis,suggesting that the parent-specific methylation of goldfish Igf2 DMR might be a primitive genomic imprinting in the early period of vertebrate evolution.These results indicate that the evolutionary foundation of genomic imprinting exists in lower vertebrates and genomic imprinting should not be considered as a unique evolutionary event of mammals.
β-catenin gene is essential for the formation of normal dorsal axial structure in vertebrates. Recent studies have provided evidence that β-catenin has a certain role in restricting the amount of organizer-induced neurectoderm formation in zebrafish and Xenopus laevis. To further understand how β-catenin represses the induction of neurectoderm formation and whether the inhibition of neural progenitor fates is essential for the normal organizer formation, it was investigated whether β-catenin was involved in repressing the early neural regulatory gene Visual system homeobox-1 (vsx1) expression during embryogenesis. It was observed that functional inhibition of endogenous β-catenin by morpholino oligonucleotides (MO) resulted in extensive and ubiquitous vsx1 expression, β-catenin could repress vsx1 promoter-driven GFP expression and transcriptional factor gene boz, the direct target gene of β-catenin, could directly repress vsx1 transcription by binding to vsx1 proximal promoter. These observations indicate that β-catenin can repress vsx1 ectopic expression cell-autonomously while it activates chordamesodermal genes expression in the precursory cells of chordamesoderm, suggesting both activation of chordamesodermal genes and repression of neural regulatory genes vsx1 in precursory chordamesoderm by β-catenin are essential for normal notochord formation.
