Epigenetic mechanism of Gtl2-miRNAs causes the primitive sheep characteristics found in purebred Merino sheep.

BACKGROUND: It is not uncommon for some individuals to retain certain primitive characteristics even after domestication or long-term intensive selection. Wild ancestors or original varieties of animals typically possess strong adaptability to environmental preservation, a trait that is often lacking in highly artificially selected populations. In the case of the Merino population, a world-renowned fine wool sheep breed, a phenotype with primitive coarse wool characteristic has re-emerged. It is currently unclear whether this characteristic is detrimental to the production of fine wool or whether it is linked to the adaptability of sheep. The underlying genetic/epigenetic mechanisms behind this trait are also poorly understood. RESULTS: This study identified lambs with an ancestral-like coarse (ALC) wool type that emerged during the purebred breeding of Merino fine wool sheep. The presence of this primitive sheep characteristic resulted in better environmental adaptability in lambs, as well as improved fine wool yield in adulthood. Reciprocal cross experiments revealed that the ALC phenotype exhibited maternal genetic characteristics. Transcriptomic SNP analysis indicated that the ALC phenotype was localized to the imprinted Gtl2-miRNAs locus, and a significant correlation was found between the ALC wool type and a newly identified short Interstitial Telomeric Sequences (s-ITSs) at this locus. We further confirmed that a novel 38-nt small RNA transcribed from these s-ITSs, in combination with the previously reported 22-nt small RNAs cluster from the Gtl2-miRNAs locus, synergistically inhibited PI3K/AKT/Metabolic/Oxidative stress and subsequent apoptotic pathways in wool follicle stem cells, resulting in the ALC wool type. The necessity of Gtl2-miRNAs in controlling primary hair follicle morphogenesis, as well as the wool follicle type for ALC wool lambs, was verified using intergenic differentially methylated region-knockout mice. CONCLUSION: The ALC wool type of Merino sheep, which does not reduce wool quality but increases yield and adaptability, is regulated by epigenetic mechanisms in the imprinted Gtl2-miRNAs region on sheep chromosome 18, with the maternally expressed imprinted gene responsible for the ALC phenotype. This study highlights the significance of epigenetic regulation during embryonic and juvenile stages and emphasizes the advantages of early adaptation breeding for maternal parents in enhancing the overall performance of their offspring.

The Immune Barrier of Porcine Uterine Mucosa Differs Dramatically at Proliferative and Secretory Phases and Could Be Positively Modulated by Colonizing Microbiota.

Endometrial immune response is highly associated with the homeostatic balance of the uterus and embryo development; however, the underlying molecular regulatory mechanisms are not fully elucidated. Herein, the porcine endometrium showed significant variation in mucosal immunity in proliferative and secretory phases by single-cell RNA sequencing. The loose arrangement and high motility of the uterine epithelium in the proliferative phase gave opportunities for epithelial cells and dendritic cells to cross talk with colonizing microbial community, guiding lymphocyte migration into the mucosal and glandular epithelium. The migrating lymphocytes were primarily NK and CD8+ T cells, which were robustly modulated by the chemokine signaling. In the secretory phase, the significantly strengthened mechanical mucosal barrier and increased immunoglobulin A alleviated the migration of lymphocytes into the epithelium when the neuro-modulation, mineral uptake, and amino acid metabolism were strongly upregulated. The noticeably increased intraepithelial lymphocytes were positively modulated by the bacteria in the uterine cavity. Our findings illustrated that significant mucosal immunity variation in the endometrium in the proliferative and secretory phases was closely related to intraepithelial lymphocyte migration, which could be modulated by the colonizing bacteria after cross talk with epithelial cells with higher expressions of chemokine.

Genome-wide profiling of RNA editing sites in sheep.

BACKGROUND: The widely observed RNA-DNA differences (RDDs) have been found to be due to nucleotide alteration by RNA editing. Canonical RNA editing (i.e., A-to-I and C-to-U editing) mediated by the adenosine deaminases acting on RNA (ADAR) family and apolipoprotein B mRNA editing catalytic polypeptide-like (APOBEC) family during the transcriptional process is considered common and essential for the development of an individual. To date, an increasing number of RNA editing sites have been reported in human, rodents, and some farm animals; however, genome-wide detection of RNA editing events in sheep has not been reported. The aim of this study was to identify RNA editing events in sheep by comparing the RNA-seq and DNA-seq data from three biological replicates of the kidney and spleen tissues. RESULTS: A total of 607 and 994 common edited sites within the three biological replicates were identified in the ovine kidney and spleen, respectively. Many of the RDDs were specific to an individual. The RNA editing-related genes identified in the present study might be evolved for specific biological functions in sheep, such as structural constituent of the cytoskeleton and microtubule-based processes. Furthermore, the edited sites found in the ovine BLCAP and NEIL1 genes are in line with those in previous reports on the porcine and human homologs, suggesting the existence of evolutionarily conserved RNA editing sites and they may play an important role in the structure and function of genes. CONCLUSIONS: Our study is the first to investigate RNA editing events in sheep. We screened out 607 and 994 RNA editing sites in three biological replicates of the ovine kidney and spleen and annotated 164 and 247 genes in the kidney and spleen, respectively. The gene function and conservation analysis of these RNA editing-related genes suggest that RNA editing is associated with important gene function in sheep. The putative functionally important RNA editing sites reported in the present study will help future studies on the relationship between these edited sites and the genetic traits in sheep.

Genome-wide association study and a post replication analysis revealed a promising genomic region and candidate genes for chicken eggshell blueness.

The eggshell blueness is an interesting object for chicken genetic studies and blue-shelled chicken industry, especially after the discovery of the causative mutation of chicken blue eggshell. In the present study, genome wide association study (GWAS) was conducted in Chinese Dongxiang blue-shelled chicken underlying four traits of blue eggshell pigments: quantity of biliverdin (QB), quantity of protoporphyrin (QP), quantity of total pigment (QT), and color density trait (CD). A total of 139 individuals were randomly collected for GWAS. We detected two SNPs in genome-wise significance and 35 in suggestive significance, 24 out of the 37 SNP were located either within intron/exon or near 15 genes in a range of ~1.17 Mb on GGA21. For further confirmation of the identified SNP loci by GWAS, the follow-up replication studies were performed in two populations. A total of 146 individuals of the second generation derived from the former GWAS population, as well as 280 individuals from an alternative independent population were employed for genotyping by MALDI-TOF MS in a genotype-phenotype association study. Eighteen SNPs evenly distributed on the GGA21 significant region were successfully genotyped in the two populations, of which 4 and 6 SNP loci were shown significantly associated with QB, QT and QP in the two repeat populations, respectively. Further, the SNPs were narrowed down to a region of ~ 653.819 Kb on GGA21 that harbors five candidate genes: AJAP1, TNFRSF9, C1ORF174, CAMTA1, and CEP104. Shell gland of chickens laying dark and light blue eggshell was chosen for detection of mRNA expression of the five candidate genes. The results showed differential expression levels of these genes in the two groups. The specific function of these genes has not yet been defined clearly in chickens and further in-depth studies are needed to explore the new functional role in chicken eggshell blueness.

Molecular cloning and characterization of the endothelin 3 gene in black bone sheep.

BACKGROUND: Black bone sheep was first discovered in Yunnan province of China in 1970, with unique black pigmentation on the body and internal organs. Endothelin 3 (EDN3) has been known as a key gene causing hyperpigmentation in black bone chicken, the Silky fowl. METHODS: In this study, EDN3 was employed as a candidate gene for regulating black color pigmentation. First, EDN3 was cloned from sheep to obtain the full-length cDNA by using the rapid amplification of cDNA ends (RACE). Genomic EDN3 was screened and a total of thirty predicted single nucleotide polymorphisms (SNPs) were genotyped for allele and genotype frequency analysis in a case-control study involving two black bone sheep populations. Genomic copy number analysis of EDN3 in sheep was conducted to measure the variation in copy number. EDN3 expression levels were observed among the groups in adult liver, lymph node, and kidney tissues, as well as embryo kidney samples. Also, among the tissues of black bone and non-black bone sheep. RESULTS: The size of the full-length cDNA was 1,578 bp, which included 426 bp of 5'-untranslated region (5'-UTR), an open reading frame (ORF) of 639 bp encoding a protein of 212 amino acids, and a 3'-UTR of 513 bp. Genotype and allele frequencies of all the discovered SNPs were found insignificantly different in black bone and non-black bone sheep (P > 0.05). Genomic copy number analysis of EDN3 in sheep revealed no significant difference between the two sheep groups. No significant variations were found in the adult liver and kidney embryo samples. However, the expression in lymph node and kidney tissue was significantly higher in black bone sheep than that in non-black bone sheep (P < 0.05). Significant variations in the EDN3 expression levels were observed among the tissues of non-black bone sheep. CONCLUSIONS: The findings of the present study indicate that unlike in Silky chickens, EDN3 is not responsible for hyperpigmentation but may play a key functional role in immune and excretory systems of black bone sheep.