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The very long chain fatty acid elongase (ELOVL) plays an important role in the synthesis of long-chain polyunsaturated fatty acids (LCPUFA). Previous studies suggest that chicken could be an alternate source of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). In this study, we detected that ELOVL5, which plays a key role in the biosynthesis of omega-3 (n-3) and omega-6 (n-6) polyunsaturated fatty acids (PUFA), was highly expressed in the liver of laying hens and increased rapidly after sexual maturity. Bioinformatic analysis revealed ELOVL fatty acid elongase 5 (ELOVL5) gene as a putative target of miR-218-5p, miR-19a-3p, miR-19b-3p, miR-30a-5p, miR-30b-5p, and miR-30e-5p. We demonstrated estrogen downregulated microRNA (miRNA), and that ELOVL5 is a direct target of miR-218-5p, which was located in intron 14 of the Slit guidance ligand 2 (SLIT2) gene and co-expressed with the host gene. Overall, estrogen enhanced hepatic synthesis of LCPUFA by functioning as a negative regulator of miRNA thereby augmenting the expression of these miRNA target genes, especially ELOVL5, which plays a key role in the biosynthesis of n-3 and n-6 LCPUFA. This study provides a novel model for the use of estrogen in the poultry industry as an inducer of ELOVL5 expression to enhance hepatic n-3 and n-6 LCPUFA synthesis at the post-transcriptional level. [ABSTRACT FROM AUTHOR]

Simple Summary: In this study, we analyzed the microRNA (miRNA) sequencing libraries of the hen ovary at four different developmental stages (15, 20, 30, and 68 W) and found 209 differently expressed (DE) miRNAs in the six comparisons of the four stages, which might be involved in ovarian growth and functions. In particular, five key miRNAs (gga-miR-2954, gga-miR-6634-5p, gga-miR-449b-5p, gga-miR-449c-3p, and gga-miR449c-5p) targeting 23 predicted genes were obtained in the interaction network and functional enrichment analysis through a combination of DE miRNAs and differentially expressed genes (DEGs), which might be involved in cell differentiation and proliferation, steroid hormone biosynthesis, and angiogenesis in ovarian development. Our findings can contribute to a better understanding of the role of functional miRNAs in ovarian development and the ovulatory cycle during the four developmental stages of hens. It is well-known that multiple functional miRNAs are found in mammals' ovaries, which are linked not only to ovarian development, but also to maturation and apoptosis. However, there is still a lack of knowledge regarding the role of miRNAs in the hen ovary. In the present study, we analyzed the miRNA sequencing libraries of ovaries at the four different developmental stages of hens (15, 20, 30, and 68 W) and a total of 677 known miRNAs and 61 novel miRNAs were identified. In total, 209 of them were differently expressed miRNAs (DE miRNAs) obtained from comparisons of the four stages, including 84 upregulated and 125 downregulated DE miRNAs. Furthermore, the five key DE miRNAs gga-miR-2954, gga-miR-6634-5p, gga-miR-449b-5p, gga-miR-449c-3p, and gga-miR449c-5p were screened using an analysis of the miRNA-mRNA interaction network and functional enrichment annotated in seven significantly enriched pathways, such as endocytosis, lysine degradation, the biosynthesis of amino acids, and the MAPK signaling pathway, which may primarily participate in cell differentiation and proliferation, steroid hormone biosynthesis, and angiogenesis by targeting the related genes. For instance, gga-miR-449 family members were predicted to target 15 genes, including TGFB1, TPM1, TPM3, and CAMKB2, which were reported to regulate follicular growth, selection, and the ovulatory cycle. Taken together, our results illustrate the ovarian miRNA profiles of the four classic developmental stages of hens and highlight the significant role of miRNAs in ovarian development and functions. However, in-depth research needs to be carried out to validate the potential functional miRNAs found in this study. [ABSTRACT FROM AUTHOR]

The fatty acid-binding protein (FABP) gene family, which encodes a group of fatty acid-trafficking molecules that affect cellular functions, has been studied extensively in mammals. However, little is known about the gene structure, expression profile, and regulatory mechanism of the gene family in chickens. In the present study, bioinformatics-based methods were used to identify the family members and investigate their evolutionary history and features of gene structure. Real-time PCR combined with in vivo and in vitro experiments were used to examine the spatiotemporal expression pattern, and explore the regulatory mechanism of FABP genes. The results show that nine members of the FABP gene family, which branched into two clusters and shared a conserved FATTYACIDBP domain, exist in the genome of chickens. Of these, seven FABP genes, including FABP1, FABP3-7, and FABP10 were abundantly expressed in the liver of hens. The expression levels of FABP1, FABP3, and FABP10 were significantly increased, FABP5 and FABP7 were significantly decreased, and FABP4 and FABP6 remained unchanged in hens at the peak laying stage in comparison to those at the pre-laying stage. Transcription of FABP1 and FABP3 were activated by estrogen via estrogen receptor (ER) α, whilst FABP10 was activated by estrogen via ERβ. Meanwhile, the expression of FABP1 was regulated by peroxisome proliferator activated receptor (PPAR) isoforms, of which tested PPARα and PPARβ agonists significantly inhibited the expression of FABP1, while tested PPARγ agonists significantly increased the expression of FABP1, but downregulated it when the concentration of the PPARγ agonist reached 100 nM. The expression of FABP3 was upregulated via tested PPARβ and PPARγ agonists, and the expression of FABP7 was selectively promoted via PPARγ. The expression of FABP10 was activated by all of the three tested PPAR agonists, but the expression of FABP4-6 was not affected by any of the PPAR agonists. In conclusion, members of the FABP gene family in chickens shared similar functional domains, gene structures, and evolutionary histories with mammalian species, but exhibited varying expression profiles and regulatory mechanisms. The results provide a valuable resource for better understanding the biological functions of individual FABP genes in chickens. [ABSTRACT FROM AUTHOR]

Accumulating evidence has shown that miR-34a serves as a posttranscriptional regulatory molecule of lipid metabolism in mammals. However, little studies about miR-34a on lipid metabolism in poultry have been reported until now. To gain insight into the biological functions and action mechanisms of miR-34a on hepatic lipid metabolism in poultry, we firstly investigated the expression pattern of miR-34a-5p, a member of miR-34a family, in liver of chicken, and determined its function in hepatocyte lipid metabolism by miR-34a-5p overexpression and inhibition, respectively. We then validated the interaction between miR-34a-5p and its target using dual-luciferase reporter assay, and explored the action mechanism of miR-34a-5p on its target by qPCR and Western blotting. Additionally, we looked into the function of the target gene on hepatocyte lipid metabolism by gain- and loss-of-function experiments. Our results indicated that miR-34a-5p showed a significantly higher expression level in livers in peak-laying hens than that in pre-laying hens. miR-34a-5p could increase the intracellular levels of triglycerides and total cholesterol in hepatocyte. Furthermore, miR-34a-5p functioned by inhibiting the translation of its target gene, long-chain acyl-CoA synthetase 1 (ACSL1), which negatively regulates hepatocyte lipid content. In conclusion, miR-34a-5p could increase intracellular lipid content by reducing the protein level, without influencing mRNA stability of the ACSL1 gene in chickens. [ABSTRACT FROM AUTHOR]