Genome-Wide Methylation Analysis of Chicken Primordial Germ Cells

The field of epigenetics has expanded rapidly in recent years. Epigenetic modifications of the genome appear to be stable in somatic cells, but in germ cells and early embryos, epigenetic reprogramming occurs on a genome-wide scale, which includes de-methylation of DNA and remodeling of histones and their modifications. Imprinted genes are epigenetically regulated so that only one allele is expressed in a parent-of-origin-dependent manner. Although they represent a small subset of the mammalian genome, imprinted genes are essential for normal development. After some investigation there has been no direct data for a similar parent-of-origin mechanism in birds, while phenotypic evidence for sire and dam effects has accumulated. DNA methylation is the most-established epigenetic mark that is critical for the allele-specific expression of imprinted genes. Recent data in the chicken, demonstrating differential promoter methylation patterns, which correlates with gene expression, resulting from environmental manipulation raises new questions on the role of epigenetics in birds. In order to better understand the underlying landscape of DNA methylation in the chicken, genome-wide DNA methylation analysis was conducted on primordial germ cells (PGCs) and partially developed embryo (day 7 of incubation, E7). Genomic libraries were prepared by fragmentation, adaptor ligation, size selection, bisulfite conversion, and were then sequenced using the Illumina next-gen platform. CpG island coverage was determined and specific genome regions with differing methylation status identified. Custom tracks were developed for the UCSC genome browser so that direct comparisons of the genome-wide methylation patterns can be visualized at the nucleotide level. The resulting data shows that many promoter regions are completely methylated in the PGCs as compared to the E7 embryo. Focusing on promoter regions that were hyper-methylated in the E7 embryo as compared to the PGCs, many genes, perhaps PGC specific, were identified including the previously known germ cell markers DAZL dead end homolog 1 (DND 1). A better understanding of the regulation of DNA methylation in the chicken will provide the foundation for further study of its role in cellular differentiation, migration, and maturation.