Multi-Stage transcriptomic dissection identifies key molecular determinants of chicken follicular development

Follicular development in the chicken ovary is a critical biological process that influences egg-laying performance and directly impacts poultry production efficiency. However, elucidating the dynamic regulatory networks involving key genes and transcription factors (TFs) during follicular development remains a major challenge. In this study, we performed a comprehensive transcriptomic analysis across six follicular stages - small white follicles, large white follicles, small yellow follicles, and three hierarchical follicles - in high and low egg-laying hens. We identified a set of essential genes and pathways closely associated with egg-laying performance, including steroid hormone biosynthesis (e.g., CYP19A1, HSD3B1) and steroid biosynthesis (e.g., CEL, DHCR24, FDFT1), which play central roles in lipid metabolism, amino acid biosynthesis, and signal transduction. Pearson correlation analysis further enabled the construction of stage-specific TF regulatory networks, highlighting pivotal TFs such as FOXO1 and HOXA10 in follicular development. Additionally, two key genes - FABP3 and ENSGALG00000054605 were identified as significantly associated with reproductive traits, showing negative correlations with egg-laying rate and age at first egg, respectively. Their differential expression trends between high and low egg-laying hens suggest potential roles in regulating laying performance and sexual maturity. Importantly, based on transcriptome predictions, we conducted functional validation of BMPR1B, a core receptor in the TGF-beta signaling pathway identified as a major regulatory hub during follicle hierarchy formation. In vitro experiments demonstrated that BMPR1B promotes granulosa-cell proliferation, confirming its essential role in follicle maturation. Collectively, this study provides new insights into the molecular regulation of chicken follicular development by integrating multi-stage transcriptomics with functional validation. The findings offer valuable molecular markers - including BMPR1B, FABP3, and ENSGALG00000054605 and potential targets for genetic improvement in poultry breeding, while also contributing to broader understanding of vertebrate reproductive biology.