Germ cell-specific DNA methylation and genome diploidization in primitive vertebrates

Abstract

Genomic imprinting in mammals causes certain genes to be expressed according to their parental origin and thereby prevents parthenogenesis. However, the evolutionary origin and significance of genomic imprinting remains unclear. Here, we study the methylation status of ntl, a developmentally decisive gene, in egg, sperm and early embryos at various developmental stages in bisexual diploid fish and unisexual polyploid fish. Bisulfite sequencing analysis revealed that maternal-specific methylation occurs at ntl promoter during gametogenesis in bisexual diploid fish, where the epigenetic asymmetry of the parental alleles is maintained during cleavage, but methylation does not occur at ntl promoter in unisexual polyploid fish. Knocking down Kaiso, a methyl-CpG dependent transcription repressor, greatly increased the expression level of the methylated maternal allele during early embryogenesis and rescued the failure of anterior notochord formation in the gynogenetic haploid of bisexual fish. This indicated that the methylated maternal ntl allele is silent during early embryogenesis and this early silencing is dependent on the methylated-CpG binding protein Kaiso. Using single sequence polymorphisms in distinguishing paternally and maternally derived transcripts in the diploid fish, we demonstrated that the unmethylated paternal allele begins transcription at the 2-cell stage and maintains transcriptional activity during cleavage. These results suggest that genomic imprinting originates from primitive vertebrates in association with genome diploidization and bisexual reproduction during vertebrate genome evolution and has a clear effect in preventing parthenogenesis.