Proliferation, apoptosis, and steroid hormone secretion by granulosa cells (GCs) and theca cells (TCs) are essential for maintaining the fate of chicken follicles. Our previous study showed that the Wnt inhibitor factor 1 (WIF1) plays a role in follicle selection. However, the significance of WIF1 in GC- and TC-associated follicular development was not explicitly investigated. This study found that WIF1 expression was strongly downregulated during follicle selection (p < 0.05) and was significantly higher in GCs than in TCs (p < 0.05). WIF1 inhibits proliferation and promotes apoptosis in GCs. Additionally, it promotes progesterone secretion in prehierarchal GCs (pre-GCs, 1.16 ± 0.05 ng/mg vs. 1.58 ng/mg ± 0.12, p < 0.05) and hierarchal GCs (hie-GCs, 395.00 ng/mg ± 34.73 vs. 527.77 ng/mg ± 27.19, p < 0.05) with the participation of the follicle-stimulating hormone (FSH). WIF1 affected canonical Wnt pathways and phosphorylated β-catenin expression in GCs. Furthermore, 604 upregulated differentially expressed genes (DEGs) and 360 downregulated DEGs in WIF1-overexpressed GCs were found through RNA-seq analysis (criteria: |${\log }_2\left(\mathrm{F}\mathrm{o}\mathrm{l}\mathrm{d}\mathrm{C}\mathrm{h}\mathrm{a}\mathrm{n}\mathrm{g}\mathrm{e}\right)$| > 1 and p_adj < 0.05). Cytokine–cytokine receptor interaction and the steroid hormone biosynthesis pathway were identified. In addition, the transcript of estrogen receptor 2 (ESR2) increased significantly (${\log }_2\left(\mathrm{F}\mathrm{o}\mathrm{l}\mathrm{d}\mathrm{C}\mathrm{h}\mathrm{a}\mathrm{n}\mathrm{g}\mathrm{e}\right)$ = 1.27, p_adj < 0.05). Furthermore, we found that WIF1 regulated progesterone synthesis by upregulating ESR2 expression in GCs. Additionally, WIF1 suppressed proliferation and promoted apoptosis in TCs. Taken together, these results reveal that WIF1 stimulates follicle development by promoting GC differentiation and progesterone synthesis, which provides an insight into the molecular mechanism of follicle selection and egg-laying performance in poultry.

Background: As a ubiquitous reversible epigenetic RNA modification, N6-methyladenosine (m6A) plays crucial regulatory roles in multiple biological pathways. However, its functional mechanisms in sex determination and differentiation during gonadal development of chicken embryos are not clear. Therefore, we established a transcriptome-wide m6A map in the female and male chicken left gonads of embryonic day 7 (E7) by methylated RNA immunoprecipitation sequencing (MeRIP-seq) to offer insight into the landscape of m6A methylation and investigate the post-transcriptional modification underlying gonadal differentiation. Results: The chicken embryonic gonadal transcriptome was extensively methylated. We found 15,191 and 16,111 m6A peaks in the female and male left gonads, respectively, which were mainly enriched in the coding sequence (CDS) and stop codon. Among these m6A peaks, we identified that 1013 and 751 were hypermethylated in females and males, respectively. These differential peaks covered 281 and 327 genes, such as BMP2, SMAD2, SOX9 and CYP19A1, which were primarily associated with development, morphogenesis and sex differentiation by functional enrichment. Further analysis revealed that the m6A methylation level was positively correlated with gene expression abundance. Furthermore, we found that YTHDC2 could regulate the expression of sex-related genes, especially HEMGN and SOX9, in male mesonephros/gonad mingle cells, which was verified by in vitro experiments, suggesting a regulatory role of m6A methylation in chicken gonad differentiation. Conclusions: This work provided a comprehensive m6A methylation profile of chicken embryonic gonads and revealed YTHDC2 as a key regulator responsible for sex differentiation. Our results contribute to a better understanding of epigenetic factors involved in chicken sex determination and differentiation and to promoting the future development of sex manipulation in poultry industry.

This study evaluated the effects of three polysaccharides, xanthan gum (XG), kappa-carrageenan (CA), and guar gum (GG), on the foaming and emulsifying properties of egg white liquid (EWL) and explored the intermolecular interactions and aggregation states in the initial polysaccharide–EWL complex. The results showed that the addition of XG and GG significantly improved the foaming stability of EWL on the one hand, from 66% to 78% and 69%, respectively (p < 0.05). On the other hand, the addition of XG and GG significantly improved the foam uniformity and density, and the average foam area decreased from 0.127 to 0.052 and 0.022 mm², respectively (p < 0.05). The addition of XG and CA significantly improved the emulsification activity index (from 13.32 to 14.58 and 14.36 m²/mg, respectively, p < 0.05) and the emulsion stability index (from 50.89 to 53.62 and 52.18 min, respectively, p < 0.05), as well as the interfacial protein adsorption at the oil–water interface; it also reduced the creaming index. However, GG negatively affected these indicators. Furthermore, the electrostatic and hydrophobic interactions among molecules in EWL due to XG and the electrostatic, hydrogen bonding, and hydrophobic interactions among molecules in EWL due to CA ultimately led to the irregular aggregation of egg white proteins. Hydrophobic interactions and disulfide bonds between molecules in EWL–containing GG formed filamentous aggregations of egg white proteins. This work reveals that molecules in the polysaccharide–egg white complexes aggregate by interaction forces, which in turn have different effects on the foaming and emulsifying properties of egg white proteins.

Background: Gene expression variation is a key underlying factor influencing phenotypic variation, and can occur via cis- or trans-regulation. To understand the role of cis- and trans-regulatory variation on population divergence in chicken, we developed reciprocal crosses of two chicken breeds, White Leghorn and Cornish Game, which exhibit major differences in body size and reproductive traits, and used them to determine the degree of cis versus trans variation in the brain, liver, and muscle tissue of male and female 1-day-old specimens. Results: We provided an overview of how transcriptomes are regulated in hybrid progenies of two contrasting breeds based on allele specific expression analysis. Compared with cis-regulatory divergence, trans-acting genes were more extensive in the chicken genome. In addition, considerable compensatory cis- and trans-regulatory changes exist in the chicken genome. Most importantly, stronger purifying selection was observed on genes regulated by trans-variations than in genes regulated by the cis elements. Conclusions: We present a pipeline to explore allele-specific expression in hybrid progenies of inbred lines without a specific reference genome. Our research is the first study to describe the regulatory divergence between two contrasting breeds. The results suggest that artificial selection associated with domestication in chicken could have acted more on trans-regulatory divergence than on cis-regulatory divergence.

Background: Duration of fertility (DF) is an important economic trait in poultry production because it has a strong effect on chick output. Various criteria or traits to assess DF on individual hens have been reported but they are affected by many nongenetic factors. Thus, a reliable definition and associated genetic parameters are needed. Because egg production is also vital in chicken breeding, knowledge of the relationship between DF and laying performance is needed for designing selection programs. Methods: We used five traits that consider both fertility and embryonic livability to delineate DF. Phenotypic and genetic analyses were completed for 2094 hens, with measurements of DF at 35 and 60 weeks of age and hatching egg production at 400 days of age (HEP400). The selection differentials for DF and HEP400 were evaluated. Results: DF is largely independent of the number of oviposited eggs in the peak laying period but both egg production and DF naturally decline with age. The heritability of the five DF traits ranged from 0.11 to 0.13 at 35 weeks of age and increased slightly in the later laying period, ranging from 0.14 to 0.17 (except for efficient duration, time between insemination and the first unhatched egg). Estimates of the genetic correlation for a given trait measured at the two ages were moderate (0.37–0.44), except for efficient duration. However, number of viable embryos depends strongly on egg production. Estimates of genetic correlations of fertility duration day (FDD) at both ages with HEP were weak. Selection for FDD improved DF but without a significant change in laying performance. Selection for increased HEP400 did not contribute to DF improvement. Conclusions: Although estimates of heritability of the five traits related to DF were low, selection to improve DF based on any one of them is possible. Among these, FDD is an effective selection criterion when the eggs are collected for approximately two weeks after insemination. The best selection procedure for DF improvement would involve multiple measurements at various ages. FDD is independent of laying performance and can be incorporated into a breeding program with egg production to improve reproductive efficiency.

Studies on trimethylamine (TMA) in egg yolk have focused on how it impacts the flavor of eggs, but there has been little focus on its other functions. We designed an in vitro antibacterial test of TMA according to TMA concentrations that covered the TMA contents typically found in egg yolk. The change in TMA content in yolk was analyzed at different storage temperatures and for different storage durations. The known antibacterial components of eggs, including the cuticle quality of the eggshell and the lysozyme activity and content in egg white, were also assessed. The total bacterial count (TBC) of different parts of eggs were detected. The results showed that the inhibitory effect of TMA on Escherichia coli (E. coli) growth increased with increasing TMA concentration, and the yolk TMA content significantly increased with storage duration (p < 0.05). The cuticle quality and lysozyme content and activity significantly decreased with storage time and increasing temperature, accompanied by a significant increase in the TBC on the eggshell surface and in the egg white (p < 0.05). This work reveals a new role for trace TMA in yolks because it reduces the risk of bacterial colonization, especially when the antibacterial function of eggs is gradually weakened during storage.