215 A comparative analysis of single-cell open chromatins in the embryonic brain between chicken and turkey.

Abstract Single-cell chromatin analysis provides new opportunities to identify homologous cells that regulate gene expression in a similar manner in different species. The aim of this study is to analyze chromatin accessibility of embryonic brain cells in chicken and turkey. Chromatin accessibility was profiled in single nuclei isolated from day-14 embryonic brain of chicken and turkey using the 10X Genomics Chromium ATAC (Assay for Transposase Accessible Chromatin) sequencing (data availability: GEO accession # GSE269838). Cluster analysis of the single-nuclei ATAC-seq data showed differential cluster patterns of embryonic brain cells between the two species based on the variation of chromatin accessibility. Data analysis further identified a total of 60,376 peaks associated with 22,382 genes in the chicken brain. These open chromatins were identified more frequently in the distal regulatory regions (n=43,224) compared to promoter regions (n=17,152). In the turkey brain, the analysis identified a total of 60,375 peaks associated with 14,516 genes. The majority of these open chromatins, similar to chicken brain, were also identified more frequently in the distal regulatory regions (n=56,834) compared to promoter regions (n=7,871). However, there was a 14-fold greater number of intergenic peaks in the turkey brain compared to chicken brain. Motif enrichment analysis of the peak sequences showed that the binding motif of the transcription factor ARID3A (AT-Rich Interaction Domain 3A) was commonly enriched as a distal cis-regulatory element of specific genes such as AGT, ANO4, ANO5, GPR139, GRIK1, GRM3, HAAO, LRRTM3, MGAT4C, RHCG, SLC25A21, TAL2, and TRPC5 in the brain cells of both chicken and turkey. Bulk RNA-seq analysis further showed that these predicted ARID3 target genes were expressed in the embryonic brain of chicken and turkey. The chorioallantoic membrane plays vital functions for embryonic development in chicken and turkey unlike the placental functions that modulate embryo development in mice. The expression of the predicted ARID3 target genes was quantified in the placenta and fetal brain of mouse. This comparative analysis showed that the majority of the ARID3 target genes were expressed in the mouse fetal brain but suppressed in the placenta. However, three genes AGT, HAAO and SLC25A21 were expressed in the placenta but suppressed in the fetal brain. The findings of this study suggest that ARID3A plays a common regulatory role in the brain development in avian and mammalian embryos. While role of ARID3A in placenta development is documented, further research is needed to understand the role of ARID3A in the development of chorioallantoic membrane of avian embryos.