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Background The growth and development of skeletal muscle are regulated by protein-coding genes and non-coding RNA. Circular RNA (circRNA) is a type of non-coding RNA involved in a variety of biological processes, especially in post-transcriptional regulation. To better understand the regulatory mechanism of circRNAs during the development of muscle in chicken, we performed RNA-seq with linear RNA depletion for chicken breast muscle in 12 (E 12) and17 (E 17) day embryos, and 1 (D 1), 14 (D 14), 56 (D 56), and 98 (D 98) days post-hatch. Results We identified 5755 differentially expressed (DE)-circRNAs during muscle development. We profiled the expression of DE-circRNAs and mRNAs (identified in our previous study) at up to six time points during chicken muscle development and uncovered a significant profile (profile 16) for circRNA upregulation during aging in muscle tissues. To investigate competing endogenous RNA (ceRNA) regulation in muscle and identify muscle-related circRNAs, we constructed a circRNA-miRNA-mRNA regulatory network using the circRNAs and mRNAs from profile 16 and miRNAs identified in our previous study, which included 361 miRNAs, 68 circRNAs, 599 mRNAs, and 31,063 interacting pairs. Functional annotation showed that upregulated circRNAs might contribute to glycolysis/gluconeogenesis, biosynthesis of amino acids, pyruvate metabolism, carbon metabolism, glycogen and sucrose metabolism through the ceRNA network, and thus affected postnatal muscle development by regulating muscle protein deposition. Of them, circRNA225 and circRNA226 from the same host gene might be key circRNAs that could regulate muscle development by interacting with seven common miRNAs and 207 mRNAs. Our experiments also demonstrated that there were interactions among circRNA225, gga-miR-1306-5p, and heat shock protein alpha 8 (HSPA8). Conclusions Our results suggest that adequate supply of nutrients such as energy and protein after hatching may be a key factor in ensuring chicken yield, and provide several candidate circRNAs for future studies concerning ceRNA regulation during chicken muscle development.

To investigate the change in zinc homeostasis of broilers under heat stress, 512 broiler chickens were raised to the age of 28 days. The broilers were then assigned to heat stress and normal temperature (36.0°C vs. 26.0°C) groups for 7 days. The results showed that oxidative stress induced by high temperature had a negative effect on the growth performance of broilers. Heat stress altered zinc homeostasis and led to a redistribution of zinc in broilers, which was reflected in increased zinc concentrations in the jejunum, liver, and tibia. Upregulation of the expression of the zinc exporter ZnT1 and importers ZIP8 and ZIP14 in the jejunum indicated that more zinc was absorbed and transported from the jejunum into the blood, while the liver increased its capacity to hold zinc through upregulation of metallothionein (MT) expression, which was achieved by reducing ZnT1 expression and upregulating the expression of the importer ZIP3. The pathway was mediated by zinc transporters, but the capacity of MT to chelate and release zinc ions also played a crucial role. The mechanism of alterations in zinc homeostasis under heat stress was revealed by the changes in zinc transporters and MT levels in the intestine and liver. Heat stress also altered cecal microbial diversity and reduced the relative abundances of Bilophila and Dialister. In conclusion, broilers altered systemic zinc homeostasis through the regulation of zinc transporters and MT in the liver and jejunum to resist oxidative stress induced by high temperature.

Dominant white, Dun, and Smoky are alleles at the Dominant white locus, which is one of the major loci affecting plumage color in the domestic chicken. Both Dominant white and Dun inhibit the expression of black eumelanin. Smoky arose in a White Leghorn homozygous for Dominant white and partially restores pigmentation. PMEL17 encodes a melanocyte-specific protein and was identified as a positional candidate gene due to its role in the development of eumelanosomes. Linkage analysis of PMEL17 and Dominant white using a red jungle fowl/White Leghorn intercross revealed no recombination between these loci. Sequence analysis showed that the Dominant white allele was exclusively associated with a 9-bp insertion in exon 10, leading to an insertion of three amino acids in the PMEL17 transmembrane region. Similarly, a deletion of five amino acids in the transmembrane region occurs in the protein encoded by Dun. The Smoky allele shared the 9-bp insertion in exon 10 with Dominant white, as expected from its origin, but also had a deletion of 12 nucleotides in exon 6, eliminating four amino acids from the mature protein. These mutations are, together with the recessive silver mutation in the mouse, the only PMEL17 mutations with phenotypic effects that have been described so far in any species.

In birds, males are the homogametic sex (ZZ) and females the heterogametic sex (ZW). Primary sex determination is thought to depend on a sex chromosome gene dosage mechanism, and the most likely sex determinant is the Z chromosome gene Doublesex and Mab-3–Related Transcription factor 1 (DMRT1). To clarify this issue, we used a CRISPR-Cas9–based monoallelic targeting approach and sterile surrogate hosts to generate birds with targeted mutations in the DMRT1 gene. The resulting chromosomally male (ZZ) chicken with a single functional copy of DMRT1 developed ovaries in place of testes, demonstrating the avian sex-determining mechanism is based on DMRT1 dosage. These ZZ ovaries expressed typical female markers and showed clear evidence of follicular development. However, these ZZ adult birds with an ovary in place of testes were indistinguishable in appearance to wild-type adult males, supporting the concept of cell-autonomous sex identity (CASI) in birds. In experiments where estrogen synthesis was blocked in control ZW embryos, the resulting gonads developed as testes. In contrast, if estrogen synthesis was blocked in ZW embryos that lacked DMRT1, the gonads invariably adopted an ovarian fate. Our analysis shows that DMRT1 is the key sex determination switch in birds and that it is essential for testis development, but that production of estrogen is also a key factor in primary sex determination in chickens, and that this production is linked to DMRT1 expression.

Background Chicken intramuscular fat (IMF) content is closely related to meat quality and performance, such as tenderness and flavor. Abdominal fat (AF) in chickens is one of the main waste products at slaughter. Excessive AF reduces feed efficiency and carcass quality. Results To analyze the differential deposition of IMF and AF in chickens, gene expression profiles in the breast muscle (BM) and AF tissues of 18 animals were analyzed by differential expression analysis and weighted co-expression network analysis. The results showed that IMF deposition in BM was associated with pyruvate and citric acid metabolism through GAPDH , LDHA , GPX1 , GBE1 , and other genes. In contrast, AF deposition was related to acetyl CoA and glycerol metabolism through FABP1 , ELOVL6 , SCD , ADIPOQ , and other genes. Carbohydrate metabolism plays an essential role in IMF deposition, and fatty acid and glycerol metabolism regulate AF deposition. Conclusion This study elucidated the molecular mechanism governing IMF and AF deposition through crucial genes and signaling pathways and provided a theoretical basis for producing high-quality broilers.

This study was conducted to characterize metabolic features of the breast muscle (pectoralis major) in chickens affected with the Wooden Breast myopathy. Live birds from two purebred chicken lines and one crossbred commercial broiler population were clinically examined by manual palpation of the breast muscle (pectoralis major) at 47-48 days of age. Metabolite abundance was determined by gas chromatography/mass spectrometry (GC/MS) and liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) using breast muscle tissue samples from 16 affected and 16 unaffected chickens. Muscle glycogen content was also quantified in breast muscle tissue samples from affected and unaffected chickens. In total, levels of 140 biochemicals were significantly different (FDR<0.1 and fold-change A/U>1.3 or <0.77) between affected and unaffected chickens. Glycogen content measurements were considerably lower (1.7-fold) in samples taken from Wooden Breast affected birds when compared with samples from unaffected birds. Affected tissues exhibited biomarkers related to increased oxidative stress, elevated protein levels, muscle degradation, and altered glucose utilization. Affected muscle also showed elevated levels of hypoxanthine, xanthine, and urate molecules, the generation of which can contribute to altered redox homeostasis. In conclusion, our findings show that Wooden Breast affected tissues possess a unique metabolic signature. This unique profile may identify candidate biomarkers for diagnostic utilization and provide mechanistic insight into altered biochemical processes contributing to tissue hardening associated with the Wooden Breast myopathy in commercial chickens.

Humans and mammalian species are unable to synthesize significant amounts of polyunsaturated fatty acids (PUFA), which therefore must be introduced with the diet. In birds, lipogenesis takes place primarily in the liver, whereas adipose tissue serves as the storage site for triacylglycerols (TG, composed by 80–85% esterified fatty acids). However, both the nature (unsaturation level, n-3, or n-6 series) and the allocation (such as constituents of complexed lipids) of PUFA are very important to evaluate their function in lipid metabolism. The objective of the present investigation was to study the liver lipid metabolism, with particular attention to non-esterified fatty acids (NEFA), TG, phospholipids (PL), FADS2 gene expression, and Δ6-desaturase activity of three chicken genotypes, Leghorn (Leg), Ross 308 (Ross), and their crossbreed (LxR), by LC/MS analysis. The concentration of single fatty acids in muscle was quantified by GC-FID. The results showed that the Ross has a lipid metabolism related mainly to storage and structural roles, exhibiting higher levels of TG, phosphatidylethanolamine (PE) and phosphatidylcholine (PC) that are largely unsaturated. Meanwhile Leg showed a relevant amount of n-3 NEFA characterized by a higher phosphatidylserine (PS) unsaturation level, FADS2 gene expression and enzyme activity. The LxR seem to have a moderate trend: n-6 and n-3 NEFA showed intermediate values compared with that of the Ross and Leg and the TG trend was similar to that of the Ross, while PE and PC were largely unsaturated (mainly 6 and 7 UNS most of the metabolic energy for storage fatty acids in their tissues (TG) whereas, the Leg birds were characterized by different lipid metabolism showing in their liver a higher content of n-3 NEFA and higher unsaturation level in PS. Furthers details are needed to better attribute the lipid energy to the different metabolic portion.

The gut microbiota has an important role in animal health and performance, but its contribution is difficult to determine, in particular given the effects of host genetic factors. Here, whole-genome sequencing of the hosts and 16S rRNA gene sequencing of the microbiota were performed to separate the effects between host genetics and the microbiota in the duodenum, jejunum, ileum, caecum and faeces on fat deposition in 206 yellow broilers reared under identical conditions. Despite the notable spatial variation in the diversity, composition and potential function of the gut microbiota, host genetics exerted limited effects on the gut microbial community. The duodenal and caecal microbiota made greater contributions to fat deposition and could separately account for 24% and 21% of the variance in the abdominal fat mass after correcting for host genetic effects. We further identified two caecal microbial taxa, Methanobrevibacter and Mucispirillum schaedleri , which were significantly correlated with fat deposition. Chickens with a lower Methanobrevibacter abundance had significantly lower abdominal fat content than those with a higher abundance of Methanobrevibacter (35.51 vs. 55.59 g), and the body weights of these chickens did not notably differ. Chickens with a higher M. schaedleri abundance exhibited lower abdominal fat accumulation (39.88 vs. 55.06 g) and body weight (2.23 vs. 2.41 kg) than those with a lower abundance of this species. These findings may aid the development of strategies for altering the gut microbiota to control fat deposition during broiler production.

Folliculogenesis is a complex biological process involving a central oocyte and its surrounding somatic cells. Three-dimensional chromatin architecture is an important transcription regulator; however, little is known about its dynamics and role in transcriptional regulation of granulosa cells during chicken folliculogenesis. We investigate the transcriptomic dynamics of chicken granulosa cells over ten follicular stages and assess the chromatin architecture dynamics and how it influences gene expression in granulosa cells at three key stages: the prehierarchical small white follicles, the first largest preovulatory follicles, and the postovulatory follicles. Our results demonstrate the consistency between the global reprogramming of chromatin architecture and the transcriptomic divergence during folliculogenesis, providing ample evidence for compartmentalization rearrangement, variable organization of topologically associating domains, and rewiring of the long-range interaction between promoter and enhancers. These results provide key insights into avian reproductive biology and provide a foundational dataset for the future in-depth functional characterization of granulosa cells. The domestic chicken Gallus gallus domesticus is a classic model for the study of folliculogenesis. Here the authors integrate multi-omics analyses characterizing the dynamic transcriptome and chromatin architecture in granulosa cells during chicken folliculogenesis.

Besides its typical role as an amino acid in protein synthesis, methionine is an important intermediate in methylation reactions. In addition, it can also be converted to cysteine and hence plays a role in the defence against oxidative stress. The present study was conducted to investigate further the role of dl-methionine (DLM) and its hydroxy analogue, dl-2-hydroxy-4-methylthiobutanoic acid (DL-HMTBA), on zootechnical performance and oxidative status of broiler chickens. Male broiler chickens were reared on two diets differing in crude protein (CP) content (low-protein, 18·3 % v. high-protein, 23·2 % CP) and were supplemented either with 0·25 % DLM or 0·25 % DL-HMTBA. Reducing the dietary protein content resulted in an impaired body weight gain (P < 0·0001). However, supplementation of DL-HMTBA to the low-protein diet partially alleviated these negative effects (P = 0·0003). This latter phenomenon could be explained by the fact that chickens fed DL-HMTBA-supplemented diets displayed a better antioxidant status as reflected in lower lipid peroxidation probably as a consequence of their higher hepatic concentrations of total and reduced glutathione compared with their DLM counterparts. On the other hand, within the high protein levels, uric acid might be an important antioxidant to explain the lower lipid peroxidation of high-protein DL-HMTBA-supplemented chickens. Hepatic methionine sulfoxide reductase-A gene expression was not significantly affected by the dietary treatments. In conclusion, the present study indicates that there are interactions between dietary protein content and supplementation of methionine analogues with respect to broiler performance and antioxidant status, also suggesting a causal link between these traits.