ATAC-seq and RNA-seq analysis unravel the mechanism of sex differentiation and infertility in sex reversal chicken

Abstract

Background Sex determination and differentiation are complex and delicate processes. In female chickens, the process of sex differentiation is sensitive and prone to be affected by the administration of aromatase inhibitors, which result in chicken sex reversal and infertility. However, the molecular mechanisms underlying sex differentiation and infertility in chicken sex reversal remain unclear. Therefore, we established a sex-reversed chicken flock by injecting an aromatase inhibitor, fadrozole, and constructed relatively high-resolution profiles of the gene expression and chromatin accessibility of embryonic gonads. Results We revealed that fadrozole affected the transcriptional activities of several genes, such as DMRT1, SOX9, FOXL2, and CYP19A1, related to sex determination and differentiation, and the expression of a set of gonadal development-related genes, such as FGFR3 and TOX3, by regulating nearby open chromatin regions in sex-reversed chicken embryos. After sexual maturity, the sex-reversed chickens were confirmed to be infertile, and the possible causes of this infertility were further investigated. We found that the structure of the gonads and sperm were greatly deformed, and we identified several promising genes related to spermatogenesis and infertility, such as SPEF2, DNAI1, and TACR3, through RNA-seq. Conclusions This study provides clear insights into the exploration of potential molecular basis underlying sex differentiation and infertility in sex-reversed chickens and lays a foundation for further research into the sex development of birds.

Overview of Avian Sex Reversal

“…An in-depth investigation has confirmed that the variation in chromatin accessibility corresponds to the alteration of gene expression [163]. For instance, compared to females, the transcriptional activity of DMRT1 in males is upregulated, which is accompanied by increased chromatin accessibility [163]. Based on these findings, we conclude that epigenetic regulation plays a key role in controlling the activation and maintenance of masculine developmental pathways in birds.…”

“…Notably, epigenetic factors are likely to contribute to maintaining gonadal fate. The chromatin accessibility of DMRT1 in embryonic female-to-male sex-reversed chickens cannot be increased to the normal male level, corresponding to its transcriptional activity [163]. In addition, the pattern of DNA methylation around CYP19A1 in estrogen-mediated sex-reversed chicken gonads fails to be induced to form the same as that in control females, suggesting that epigenetic elements are involved in the establishment of avian gonadal CASI [177,178].…”

“…Similarly, studies have revealed striking differences in the landscape of genomic chromatin accessibility between female and male embryonic left gonads [163]. An in-depth investigation has confirmed that the variation in chromatin accessibility corresponds to the alteration of gene expression [163]. For instance, compared to females, the transcriptional activity of DMRT1 in males is upregulated, which is accompanied by increased chromatin accessibility [163].…”

“…Experiments have shown that the m6A-recognized protein YTH Domain Containing 2 (YTHDC2) can regulate the expression of genes related to male sex differentiation, such as SOX9 and HEMGN [162]. Similarly, studies have revealed striking differences in the landscape of genomic chromatin accessibility between female and male embryonic left gonads [163]. An in-depth investigation has confirmed that the variation in chromatin accessibility corresponds to the alteration of gene expression [163].…”

Transcriptomic and epigenomic insights into pectoral muscle fiber formation at the late embryonic development in pure chicken lines

“…Combining this technique with RNA-seq greatly enhances our understanding of gene regulation. Researches using both omics technologies to investigate mechanisms such as sex differentiation ( Li et al, 2022b ; Zhang et al, 2023a ) and the fate specification of feathers/scales ( Lai et al, 2018 ) has been conducted, while studies on muscle development during the embryonic period remain scarce.…”