Integrative analysis of hypothalamic transcriptome and genetic association study reveals key genes involved in the regulation of egg production in indigenous chickens

Indigenous chicken products are increasingly favored by consumers due to their unique meat and egg quality. However, the relatively poor egg-laying performance largely impacts the economic benefits and hinders sustainable development of the local chicken industry. Thus, excavating key genes and effective molecular markers associated with egg-laying performance is necessary to improve egg production via genetic selection in indigenous breeds. In the present study, comparative hypothalamic transcriptome between pre-laying (15 weeks old) and peak-laying (30 weeks old) Lushi blue-shelled-egg (LBS) chicken was performed. A total of 518 differentially expressed genes (DEGs) were identified. Among the DEGs, 64 genes were enriched in 10 Gene Ontology (GO) terms associated with reproductive regulation via GO analysis and considered as potential candidate genes regulating egg-laying performance. Of the 64 genes, 16 showed high connectivity (degree≥12) by protein–protein interaction (PPI) network analysis and were considered as potential core candidate genes (PCCGs). To further look for key candidate genes from the PCCGs, firstly, the expression patterns of the 16 genes were examined in the hypothalamus of two indigenous breeds (LBS and Gushi (GS) chickens) between the pre-laying and peak-laying stages using quantitative real-time PCR (qRT-PCR). Eleven out of the 16 genes showed significantly differential expression (P<0.05) with the same changing trends in the two breeds. Then, correlations between the expression levels of the above 11 genes and egg numbers and reproductive hormone concentrations in serum were investigated in high-yielding and low-yielding GS chickens. Of the 11 genes, eight showed significant correlations (P<0.05) between their expression levels and egg numbers, and between expression levels and reproductive hormone concentration in serum. Furthermore, an association study on single nucleotide polymorphisms (SNPs) identified in these eight genes and egg production traits was carried out in 640 GS hens, and a significant association (P<0.05) between the SNPs and egg numbers was confirmed. In conclusion, the eight genes, including CNR1, AP2M1, NRXN1, ANXA5, PENK, SLC1A2, SNAP25 and TRH, were demonstrated as key genes regulating egg production in indigenous chickens, and the SNPs sites within the genes might be served as markers to provide a guide for indigenous chicken breeding. These findings provide a novel insight for further understanding the regulatory mechanisms of egg-laying performance and developing molecular markers to improve egg production of indigenous breeds.