The causative mutation for muffs and beard phenotype correlates with gene expression profile in chicken embryonic brains: an RNA-seq analysis

A complex structural variation on chicken chromosome 27 is the causal mutation for the muffs and beard (Mb) phenotype. The Mb variant region contains copy number variations in several genes important for nervous system development, including HOXB8, HOXB7, and SMARCE1. However, it was previously unknown whether genes in this region are expressed in the brain or influence neural development. In this study, we collected embryonic brain tissues from Beijing-You Mb and non-Mb chickens at different developmental time points. RT-qPCR revealed that SMARCD2, HOXB8, SMARCE1, and KRT222 were highly expressed in Mb chicken brains. HOXB8 showed stable differential expression after day 15 and had the highest fold change among all differentially expressed genes (DEGs). RNA-seq of 19-day brain tissue indicated that DEGs were enriched in neural development processes such as synaptic assembly, signal transduction, chemical synaptic transmission, and neurotransmitter regulation. Several key neural development genes were also differentially expressed, providing a biological basis for developmental difference. Neurotransmitter analysis further revealed significant differences in glutamate and GABA levels. HOXB8, an important regulator of neural development, showed widespread mRNA expression in brain sections, especially in cortical tissues, and its homeobox domain is evolutionarily conserved. Overexpression of HOXB8 in chicken embryo primary brain cells significantly regulated multiple neural development-related genes, including AGT, SLC32A1, SLC18A2, RPS20, RPL35A, SLC6A11, and RLN3. In conclusion, this study is the first to show that genes within the Mb region are highly expressed in chicken embryonic brain tissue and mediate differences in neural development-related gene expression and neurotransmitter content, with HOXB8 as a key candidate. These findings open a new avenue for research on the Mb variant in neural development, and future work should further investigate neurological and behavioral differences in Mb chickens.