Transcriptome-based analysis of key signaling pathways affecting the formation of primordial germ cell in chickens

Bone morphogenetic protein 4(BMP4)can induce the formation of chicken primordial germ cells(PGCs)in vitro;however,its regulatory mechanism in poultry remains unknown.This study aimed to use RNA-seq to analyze PGCs in chicken embryos and iPGCs induced by BMP4 in vitro,clarify the internal regulatory factors of PGCs,analyze the mechanism of the formation of PGCs,and lay a theoretical foundation for the further optimization of PGCs induction systems.Embryonic stem cells(ESCs),PGCs and iPGCs induced by BMP4 in vitro were collected.The transcriptional maps of the three cell types were studied using RNA-seq.The results showed 6,142 genes differentially expressed between PGCs and iPGCs,of which 2,728 were upregulated in iPGCs and 3,414 were downregulated in iPGCs.Compared to that in ESCs,BMP4 was significantly upregulated in PGCs and iPGCs.KEGG results showed that both the TGF-βand Wnt signaling pathways were activated during the formation of PGCs in vitro and in vivo,and the activation was more significant during iPGCs induced by BMP4.The expression of Nodal,an inhibitory factor of TGF-βsignaling,was significantly decreased in PGCs and iPGCs,but was not expressed in iPGCs,which further supports our conclusion.Additionally,the Lysosome and PI3K-AKT signaling pathways were significantly enriched in PGCs and iPGCs,respectively.Further,transmission electron microscopy(TEM)results showed that the number of autolysosomes was significantly higher after the addition of BMP4,which is consistent with the KEGG results.Furthermore,the number of PGCs was significantly reduced after ATG14 was interfered in vivo and in vitro.In conclusion,this study screened out the key signaling pathways during the formation of PGCs,aiming to provide help for enriching the mechanism network regulating PGCs formation in chicken and laying a theoretical foundation for further improving the efficiency of inducing PGCs in vitro.

miR-24-3p promotes proliferation and inhibits apoptosis of porcine granulosa cells by targeting P27

Ovarian follicle development is associated with the physiological functions of granulosa cells(GCs),including proliferation and apoptosis.The level of miR-24-3p in ovarian tissue of high-yielding Yorkshire×Landrace sows was significantly higher than that of low-yielding sows.However,the functions of miR-24-3p on GCs are unclear.In this study,using flow cytometry,5-ethynyl-2′-de-oxyuridine(EdU)staining,and cell count,we showed that miR-24-3p promoted the proliferation of GCs increasing the proportion of cells in the S phase and upregulating the expression of cell cycle genes,moreover,miR-24-3p inhibited GC apoptosis.Mechanistically,on-line prediction,bioinformatics analysis,a luciferase reporter assay,RT-qPCR,and Western blot results showed that the target gene of miR-24-3p in proliferation and apoptosis is cyclin-dependent kinase inhibitor 1B(P27/CDKN1B).Furthermore,the effect of miR-24-3p on GC proliferation and apoptosis was attenuated by P27 overexpression.These findings suggest that miR-24-3p regulates the physiological functions of GCs.

Dynamic transcriptome profiles and novel markers in bovine spermatogenesis revealed by single-cell sequencing

Testicular development is an important biological process in male and requires interaction between the male germ cells and somatic cells.However,the mechanisms of testicular development in livestock,particularly in cattle,are poorly understood.Furthermore,cellular heterogeneity hinders the profiling of different cell types at different developmental stages.In this study,we first performed a single-cell transcriptomic study of the bovine testis development during puberty by using 10×genomics single-cell RNA sequencing(scRNA-seq).By collecting the scRNA-seq data from 11,083 cells from prepubertal and pubertal bovine testes,a high-resolution scRNA-seq atlas was described,identifying 9 somatic and 13 spermatogenic clusters.We also distinguished several stage-specific marker genes for bovine germ cells and somatic cells,such as GRAF2 and MORC1 for SSC(spermatogonial stem cells),HJURP and TCF19 for differentiating spermatogonia,ARSE for immature Sertoli,CLEC12B for mature Sertoli,LOC112441470 for Leydig.In conclusion,we have examined the transcription levels and constructed the single-cell developmental maps of germ cells and somatic cells during testicular development in Angus cattle.The datasets provided new insights into spermatogenesis and testicular somatic cell development in cattle.

Transcriptome analysis reveals the genetic basis of crest cushion formation in duck

The Chinese crested duck is a unique duck breed having a bulbous feather shape on its duck head.However,the mechanisms involved in its formation and development are unclear.In the present study,RNA sequencing analysis was performed on the crested tissues of 6 Chinese crested ducks and the scalp tissues of 6 cherry valley ducks(CVs)from 2 developmental stages.This study identified 261 differentially expressed genes(DEGs),122 upregulated and 139 downregulated,in the E28 stage and 361 DEGs,154 upregulated and 207 downregulated in the D42 stage between CC and CV ducks.The subsequent results of weighted gene co-expression network analysis(WGCNA)revealed that the turquoise and cyan modules were associated with the crest trait in the D42 stage,meanwhile,the green,brown,and pink modules were associated with the crest trait in the E28 stage.Venn analysis of the DEGs and WGCNA showed that 145 and 45 genes are associated between the D42 and E28 stages,respectively.The expression of WNT16,BMP2,SLC35F2,SLC6A15,APOBEC2,ABHD6,TNNC2,MYL1,and TNNI2 were verified by real-time quantitative PCR.This study provides an approach to reveal the molecular mechanisms underlying the crested trait development.

Transcriptomic and epigenomic landscapes of muscle growth during the postnatal period of broilers

Background Broilers stand out as one of the fastest-growing livestock globally,making a substantial contribution to animal meat production.However,the molecular and epigenetic mechanisms underlying the rapid growth and development of broiler chickens are still unclear.This study aims to explore muscle development patterns and regulatory networks during the postnatal rapid growth phase of fast-growing broilers.We measured the growth performance of Cornish(CC)and White Plymouth Rock(RR)over a 42-d period.Pectoral muscle samples from both CC and RR were randomly collected at day 21 after hatching(D21)and D42 for RNA-seq and ATAC-seq library construction.Results The consistent increase in body weight and pectoral muscle weight across both breeds was observed as they matured,with CC outpacing RR in terms of weight at each stage of development.Differential expression analysis identified 398 and 1,129 genes in the two dimensions of breeds and ages,respectively.A total of 75,149 ATAC-seq peaks were annotated in promoter,exon,intron and intergenic regions,with a higher number of peaks in the promoter and intronic regions.The age-biased genes and breed-biased genes of RNA-seq were combined with the ATAC-seq data for subsequent analysis.The results spotlighted the upregulation of ACTC1 and FDPS at D21,which were primarily associated with muscle structure development by gene cluster enrichment.Additionally,a noteworthy upregulation of MUSTN1,FOS and TGFB3 was spotted in broiler chickens at D42,which were involved in cell differentiation and muscle regeneration after injury,suggesting a regulatory role of muscle growth and repair.Conclusions This work provided a regulatory network of postnatal broiler chickens and revealed ACTC1 and MUSTN1 as the key responsible for muscle development and regeneration.Our findings highlight that rapid growth in broiler chickens triggers ongoing muscle damage and subsequent regeneration.These findings provide a foundation for future research to investigate the functional aspects of muscle development.

Spatially resolved transcriptomic profiling of placental development in dairy cow

The placenta plays a crucial role in successful mammalian reproduction.Ruminant animals possess a semi-invasive placenta characterized by a highly vascularized structure formed by maternal endometrial caruncles and fetal placental cotyledons,essential for full-term fetal development.The cow placenta harbors at least two trophoblast cell populations:uninucleate(UNC)and binucleate(BNC)cells.However,the limited capacity to elucidate the transcriptomic dynamics of the placental natural environment has resulted in a poor understanding of both the molecular and cellular interactions between trophoblast cells and niches,and the molecular mechanisms governing trophoblast differentiation and functionalization.To fill this knowledge gap,we employed Stereo-seq to map spatial gene expression patterns at near single-cell resolution in the cow placenta at 90 and 130 days of gestation,attaining high-resolution,spatially resolved gene expression profiles.Based on clustering and cell marker gene expression analyses,key transcription factors,including YBX1 and NPAS2,were shown to regulate the heterogeneity of trophoblast cell subpopulations.Cell communication and trajectory analysis provided a framework for understanding cell-cell interactions and the differentiation of trophoblasts into BNCs in the placental microenvironment.Differential analysis of cell trajectories identified a set of genes involved in regulation of trophoblast differentiation.Additionally,spatial modules and co-variant genes that help shape specific tissue structures were identified.Together,these findings provide foundational insights into important biological pathways critical to the placental development and function in cows.

Insights into the interplay between gut microbiota and lipid metabolism in the obesity management of canines and felines

Obesity is a prevalent chronic disease that has significant negative impacts on humans and our companion animals,including dogs and cats.Obesity occurs with multiple comorbidities,such as diabetes,hypertension,heart disease and osteoarthritis in dogs and cats.A direct link between lipid metabolism dysregulation and obesity-associated diseases has been implicated.However,the understanding of such pathophysiology in companion animals is lim-ited.This review aims to address the role of lipid metabolism in various metabolic disorders associated with obesity,emphasizing the involvement of the gut microbiota.Furthermore,we also discuss the management of obesity,including approaches like nutritional interventions,thus providing novel insights into obesity prevention and treatment for canines and felines.

Identification of key genes affecting porcine fat deposition based on coexpression network analysis of weighted genes

Background:Fat deposition is an important economic consideration in pig production.The amount of fat deposition in pigs seriously affects production efficiency,quality,and reproductive performance,while also affecting consumers’choice of pork.Weighted gene co-expression network analysis(WGCNA)is effective in pig genetic studies.Therefore,this study aimed to identify modules that co-express genes associated with fat deposition in pigs(Songliao black and Landrace breeds)with extreme levels of backfat(high and low)and to identify the core genes in each of these modules.Results:We used RNA sequences generated in different pig tissues to construct a gene expression matrix consisting of 12,862 genes from 36 samples.Eleven co-expression modules were identified using WGCNA and the number of genes in these modules ranged from 39 to 3,363.Four co-expression modules were significantly correlated with backfat thickness.A total of 16 genes(RAD9A,IGF2R,SCAP,TCAP,SMYD1,PFKM,DGAT1,GPS2,IGF1,MAPK8,FABP,FABP5,LEPR,UCP3,APOF,and FASN)were associated with fat deposition.Conclusions:RAD9A,TCAP,SMYD1,PFKM,GPS2,and APOF were the key genes in the four modules based on the degree of gene connectivity.Combining these results with those from differential gene analysis,SMYD1 and PFKM were proposed as strong candidate genes for body size traits.This study explored the key genes that regulate porcine fat deposition and lays the foundation for further research into the molecular regulatory mechanisms underlying porcine fat deposition.

Stimulation Study of Gene Pyramiding inAnimals by Marker-Assisted Selection

This gene pyramiding strategy is based on the idea of efficiently pyramiding genes of interest by crosses and selection to obtain a population with favorable alleles from different breeds or lines, which is called an ideal population. We investigate impacts of some factors on the pyramiding efficiencies by simulation. These factors include selection strategies （the breeding value selection, the molecular scores selection and the index selection）, proportion selected （2, 10 and 20%）, recombination rates between adjacent target genes （0.1, 0.3 and 0.5） and different mating types （the random mating and the positive assortative mating avoiding sib mating）. The results show that： （1） The more recombination rate and the lower proportion male selected, the better pyramiding efficiency; （2） the ideal population is obtained via various selection strategies, while different selection strategies are suitable for different breeding objectives. From the perspective of pyramiding target genes merely, the molecular scores selection is the best one, for the purpose of pyramiding target genes and recovering genetic background of the target trait, the index selection is the best one, while from the saving cost point of view, the breeding value selection is the best one; （3） the positive assortative mating is more efficient for gene pyramiding compared with the random mating in the terms of the number of generations of intercross for getting the ideal population.

CLOCK inhibits the proliferation of porcine ovarian granulosa cells by targeting ASB9

Background Clock circadian regulator(CLOCK)is a core factor of the mammalian biological clock system in regulat-ing female fertility and ovarian physiology.However,CLOCK’s specific function and molecular mechanism in porcine granulosa cells(GCs)remain unclear.In this study,we focused on CLOCK’s effects on GC proliferation.Results CLOCK significantly inhibited cell proliferation in porcine GCs.CLOCK decreased the expression of cell cycle-related genes,including CCNB1,CCNE1,and CDK4 at the mRNA and protein levels.CDKN1A levels were upregulated by CLOCK.ASB9 is a newly-identified target of CLOCK that inhibits GC proliferation;CLOCK binds to the E-box element in the ASB9 promoter.Conclusions These findings suggest that CLOCK inhibits the proliferation of porcine ovarian GCs by increasing ASB9 level.

Genetic gain and inbreeding from simulation of different genomic mating schemes for pig improvement

Background Genomic selection involves choosing as parents those elite individuals with the higher genomic estimated breeding values(GEBV)to accelerate the speed of genetic improvement in domestic animals.But after multi-generation selection,the rate of inbreeding and the occurrence of homozygous harmful alleles might increase,which would reduce performance and genetic diversity.To mitigate the above problems,we can utilize genomic mating(GM)based upon optimal mate allocation to construct the best genotypic combinations in the next generation.In this study,we used stochastic simulation to investigate the impact of various factors on the efficiencies of GM to optimize pairing combinations after genomic selection of candidates in a pig population.These factors included:the algorithm used to derive inbreeding coefficients;the trait heritability(0.1,0.3 or 0.5);the kind of GM scheme(focused average GEBV or inbreeding);the approach for computing the genomic relationship matrix(by SNP or runs of homozygosity(ROH)).The outcomes were compared to three traditional mating schemes(random,positive assortative or negative assortative matings).In addition,the performance of the GM approach was tested on real datasets obtained from a Large White pig breeding population.Results Genomic mating outperforms other approaches in limiting the inbreeding accumulation for the same expected genetic gain.The use of ROH-based genealogical relatedness in GM achieved faster genetic gains than using relatedness based on individual SNPs.The GROH-based GM schemes with the maximum genetic gain resulted in 0.9%-2.6%higher rates of genetic gainΔG,and 13%-83.3%lowerΔF than positive assortative mating regardless of heritability.The rates of inbreeding were always the fastest with positive assortative mating.Results from a purebred Large White pig population,confirmed that GM with ROH-based GRM was more efficient than traditional mating schemes.Conclusion Compared with traditional mating schemes,genomic mating can not only achieve sustainable genetic progress but also effectively control the rates of inbreeding accumulation in the population.Our findings demonstrated that breeders should consider using genomic mating for genetic improvement of pigs.

Antimicrobial peptides act on the rumen the performance of castrated bulls

Background Many countries have already banned the use of antibiotics in animal husbandry,making it extremely difficult to maintain animal health in livestock breeding.In the livestock industry,there is an urgent need to develop alternatives to antibiotics which will not lead to drug resistance on prolonged use.In this study,eighteen castrated bulls were randomly divided into two groups.The control group(CK)was fed the basal diet,while the antimicrobial peptide group(AP)was fed the basal diet supplemented with 8 g of antimicrobial peptides in the basal diet for the experimental period of 270 d.They were then slaughtered to measure production performance,and the ruminal contents were isolated for metagenomic and metabolome sequencing analysis.Result The results showed that antimicrobial peptides could improve the daily weight,carcass weight,and net meat weight of the experimental animals.Additionally,the rumen papillae diameter and the micropapillary density in the AP were significantly greater than those in the CK.Furthermore,the determination of digestive enzymes and fermentation parameters showed that the contents of protease,xylanase,andβ-glucoside in the AP were greater than those in the CK.However,lipase content in the CK was greater than that in the AP.Moreover,the content of acetate,propionate,butyrate,and valerate was found to be greater in AP than those in CK.The metagenomic analysis annotated 1993 differential microorganisms at the species level.The KEGG enrichment of these microorganisms revealed that the enrichment of drug resistance-related pathways was dramatically decreased in the AP,whereas the enrichment of immune-related pathways was significantly increased.There was also a significant reduction in the types of viruses in the AP.187 probiotics with significant differences were found,135 of which were higher in AP than in CK.It was also found that the antimicrobial mechanism of the antimicrobial peptides was quite specific.Seven low-abundance microorganisms(Acinetobactersp.Ac1271,Aequorivita soesokkakensis,Bacillus lacisalsi,Haloferax larsenii,Lysinibacillussp.3DF0063,Parabacteroidessp.217,Streptomycessp.So13.3)were found to regulate growth performance of the bull negatively.Metabolome analysis identified 45 differentially differential metabolites that significantly different between the CK and the AP groups.Seven upregulated metabolites(4-pyridoxic acid,Ala-Phe,3-ureidopropionate,hippuric acid,terephthalic acid,L-alanine,uridine 5-monophosphate)improve the growth performance of the experimental animals.To detect the interactions between the rumen microbiome and metabolism,we associated the rumen microbiome with the metabolome and found that negative regulation between the above 7 microorganisms and 7 metabolites.Conclusions This study shows that antimicrobial peptides can improve the growth performance of animals while resisting viruses and harmful bacteria and are expected to become healthy alternatives to antibiotics.We demonstrated a new antimicrobial peptides pharmacological model.We demonstrated low-abundance microorganisms may play a role by regulating the content of metabolites.

Genomic analysis of indigenous goats in Southwest Asia reveals evidence of ancient adaptive introgression related to desert climate

Understanding how evolutionary pressures related to climate change have shaped the current genetic background of domestic animals is a fundamental pursuit of biology. Here, we generated wholegenome sequencing data from native goat populations in Iraq and Pakistan. Combined with previously published data on modern, ancient(Late Neolithic to Medieval periods), and wild Capra species worldwide, we explored the genetic population structure, ancestry components, and signatures of natural positive selection in native goat populations in Southwest Asia(SWA). Results revealed that the genetic structure of SWA goats was deeply influenced by gene flow from the eastern Mediterranean during the Chalcolithic period, which may reflect adaptation to gradual warming and aridity in the region. Furthermore, comparative genomic analysis revealed adaptive introgression of the KITLG locus from the Nubian ibex(C. nubiana) into African and SWA goats. The frequency of the selected allele at this locus was significantly higher among goat populations located near northeastern Africa. These results provide new insights into the genetic composition and history of goat populations in the SWA region.

Micro RNA transcriptome of skeletal muscle during yak development reveals that miR-652 regulates myoblasts differentiation and survival by targeting ISL1

The growth and development of skeletal muscle also determine the meat production of yak, ultimately affecting the economic benefits. Hence, improving growth performance is a top priority in the yak industry. Skeletal muscle development is a complex process involving the regulation of several genes, including microRNAs(miRNAs). However,the transcription of miRNAs in yak skeletal muscle during prenatal to postnatal stages is unknown. We used small RNA sequencing(small RNA-Seq) to determine the global miRNAs of longissimus dorsi muscle from yak(the samples were collected from three fetuses and three adults). Totally 264 differently expressed miRNAs(|log2(fold change)|>1and P-value≤0.05) were detected between the two groups. Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) analysis showed that differently expressed miRNAs-targeted genes participated in pathways associated with muscle development, such as MAPK, PI3K-Akt, and Hippo signaling pathways, etc. MiR-652, which was up-regulated in the fetal group, was transfected into C2C12 myoblasts to examine its role. miR-652 promoted(P≤0.05)proliferation and differentiation, but inhibited(P≤0.001) apoptosis at early period. Furthermore, miR-652 reduced(P≤0.001) the proportion of C2C12 myoblasts in the G1 phase while increasing(P≤0.01) the proportion of cells in the S and G2 phases. Dual-luciferase reporter assays indicated that ISL1 served as a target of miR-652. In general, these findings expand our understanding of yak skeletal muscle miRNAs, and suggested that miR-652 probably regulated myogenesis by regulating ISL1.

Identification of biomarkers associated with the feed efficiency by metabolomics profiling:results from the broiler lines divergent for high or low abdominal fat content

Background:Improving feed efficiency(FE)is one of the main objectives in broiler breeding.It is difficult to directly measure FE traits,and breeders hence have been trying to identify biomarkers for the indirect selection and improvement of FE traits.Metabolome is the"bridge"between genome and phenome.The metabolites may potentially account for more of the phenotypic variation and can suitably serve as biomarkers for selecting FE traits.This study aimed to identify plasma metabolite markers for selecting high-FE broilers.A total of 441 birds from Northeast Agricultural University broiler lines divergently selected for abdominal fat content were used to analyze plasma metabolome and estimate the genetic parameters of differentially expressed metabolites.Results:The results identified 124 differentially expressed plasma metabolites(P<0.05)between the lean line(highFE birds)and the fat line(low-FE birds).Among these differentially expressed plasma metabolites,44 were found to have higher positive or negative genetic correlations with FE traits(|r_g|≥0.30).Of these 44 metabolites,14 were found to display moderate to high heritability estimates(h~2≥0.20).However,among the 14 metabolites,4 metabolites whose physiological functions have not been reported were excluded.Ultimately,10 metabolites were suggested to serve as the potential biomarkers for breeding the high-FE broilers.Based on the physiological functions of these metabolites,reducing inflammatory and improving immunity were proposed to improve FE and increase production efficiency.Conclusions:According to the pipeline for the selection of the metabolite markers established in this study,it was suggested that 10 metabolites including 7-ketocholesterol,dimethyl sulfone,epsilon-(gamma-glutamyl)-lysine,gamma-glutamyltyrosine,2-oxoadipic acid,L-homoarginine,testosterone,adenosine 5'-monophosphate,adrenic acid,and calcitriol could be used as the potential biomarkers for breeding the"food-saving broilers".

Signatures of positive selection for local adaptation of African native cattle populations:A review

Cattle are central to the lives and diverse cultures of African people.It has played a crucial role in providing valuable protein for billions of households and sources of income and employment for producers and other actors in the livestock value chains.The long-term natural selection of African cattle typically signals signatures in the genome,contributes to high genetic differentiations across breeds.This has enabled them to develop unique adaptive traits to cope with inadequate feed supply,high temperatures,high internal and external parasites,and diseases.However,these unique cattle genetic resources are threatened by indiscriminate cross-breeding,breed replacements with exotic cosmopolitan breeds,and climate change pressures.Although there are no functional genomics studies,recent advancements in genotyping and sequencing technologies have identified and annotated limited functional genes and causal variants associated with unique adaptive and economical traits of African cattle populations.These genome-wide variants serve as candidates for breed improvement and support conservation efforts for endangered cattle breeds against future climate changes.Therefore,this review plans to collate comprehensive information on the identified selection footprints to support genomic studies in African cattle to confirm the validity of the results and provide a framework for further genetic association and QTL fine mapping studies.

Genome‑wide DNA methylation and transcriptome analyses reveal the key gene for wool type variation in sheep

Background Wool fibers are valuable materials for textile industry.Typical wool fibers are divided into medullated and non-medullated types,with the former generated from primary wool follicles and the latter by either primary or secondary wool follicles.The medullated wool is a common wool type in the ancestors of fine wool sheep before breeding.The fine wool sheep have a non-medullated coat.However,the critical period determining the type of wool follicles is the embryonic stage,which limits the phenotypic observation and variant contrast,making both selection and studies of wool type variation fairly difficult.Results During the breeding of a modern fine(MF)wool sheep population with multiple-ovulation and embryo transfer technique,we serendipitously discovered lambs with ancestral-like coarse(ALC)wool.Whole-genome rese-quencing confirmed ALC wool lambs as a variant type from the MF wool population.We mapped the significantly associated methylation locus on chromosome 4 by using whole genome bisulfite sequencing signals,and in turn identified the SOSTDC1 gene as exons hypermethylated in ALC wool lambs compare to their half/full sibling MF wool lambs.Transcriptome sequencing found that SOSTDC1 was expressed dozens of times more in ALC wool lamb skin than that of MF and was at the top of all differentially expressed genes.An analogy with the transcriptome of coarse/fine wool breeds revealed that differentially expressed genes and enriched pathways at postnatal lamb stage in ALC/MF were highly similar to those at the embryonic stage in the former.Further experiments validated that the SOSTDC1 gene was specifically highly expressed in the nucleus of the dermal papilla of primary wool follicles.Conclusion In this study,we conducted genome-wide differential methylation site association analysis on differen-tial wool type trait,and located the only CpG locus that strongly associated with primary wool follicle development.Combined with transcriptome analysis,SOSTDC1 was identified as the only gene at this locus that was specifically overexpressed in the primary wool follicle stem cells of ALC wool lamb skin.The discovery of this key gene and its epigenetic regulation contributes to understanding the domestication and breeding of fine wool sheep.

Analysis and Identification of miRNA Expression in the Skeletal Muscle of Sichuan White Rabbits

[Objectives]miRNAs play an important role in the proliferation and differentiation of different myoblasts.This study was conducted to elucidate the complex genetic mechanisms that affect the meat production performance of Sichuan white rabbits and reveal the regulatory role of miRNAs in their muscle growth and meat quality formation.[Methods]Three constructed skeletal muscle libraries of Sichuan white rabbits aged six months were sequenced by the solexa technology to identify known miRNAs,predict new miRNAs and construct an expression profile of muscle miRNAs.[Results]A total of 511 known miRNAs and 42 miRNAs were detected in 34089472 pure sequences,and the proportion of miRNAs with a length of 22 nt was the highest.The number of known miRNA sequences accounted for 71.38%of pure sequences,which was much higher than the proportion of other types of RNAs.The proportion of sequences from exons was 0.38%,indicating a low degree of mRNA degradation in the samples.Base U had the highest proportion at the first position,and the bases with the highest proportions at positions 8 and 10 were U and A,respectively.Muscle-specific miRNAs(miR-1,miR-133,and miR-206)ranked in the top 10 in terms of expression level.The number and expression levels of new miRNAs were lower than those of known miRNAs.The length distribution,base bias at different positions and expression profile characteristics of miRNAs might be related to the biological function of miRNAs in regulating muscle proliferation and differentiation and the action mechanisms with target genes.[Conclusions]The identification and expression of miRNAs in muscle tissues of Sichuan white rabbits will help to understand the complex molecular mechanisms of meat production performance and provide a theoretical basis for the functional research of miRNAs in meat rabbits.

Single‑cell sequencing reveals the reproductive variations between primiparous and multiparous Hu ewes

Background In the modern sheep production systems,the reproductive performance of ewes determines the economic profitability of farming.Revealing the genetic mechanisms underlying differences in the litter size is important for the selection and breeding of highly prolific ewes.Hu sheep,a high-quality Chinese sheep breed,is known for its high fecundity and is often used as a model to study prolificacy traits.In the current study,animals were divided into two groups according to their delivery rates in three consecutive lambing seasons(namely,the high and low reproductive groups with≥3 lambs and one lamb per season,n=3,respectively).The ewes were slaughtered within 12 h of estrus,and unilateral ovarian tissues were collected and analyzed by 10×Genomics single-cell RNA sequencing.Results A total of 5 types of somatic cells were identified and corresponding expression profiles were mapped in the ovaries of each group.Noticeably,the differences in the ovary somatic cell expression profiles between the high and low reproductive groups were mainly clustered in the granulosa cells.Furthermore,four granulosa cell subtypes were identified.GeneSwitches analysis revealed that the abundance of JPH1 expression and the reduction of LOC101112291 expression could lead to different evolutionary directions of the granulosa cells.Additionally,the expression levels of FTH1 and FTL in mural granulosa cells of the highly reproductive group were significantly higher.These genes inhibit necroptosis and ferroptosis of mural granulosa cells,which helps prevent follicular atresia.Conclusions This study provides insights into the molecular mechanisms underlying the high fecundity of Hu sheep.The differences in gene expression profiles,particularly in the granulosa cells,suggest that these cells play a critical role in female prolificacy.The findings also highlight the importance of genes such as JPH1,LOC101112291,FTH1,and FTL in regulating granulosa cell function and follicular development.

Hepatic steatosis is associated with dysregulated cholesterol metabolism and altered protein acetylation dynamics in chickens

Background Hepatic steatosis is a prevalent manifestation of fatty liver, that has detrimental effect on the health and productivity of laying hens, resulting in economic losses to the poultry industry. Here, we aimed to systematically investigate the genetic regulatory mechanisms of hepatic steatosis in laying hens.Methods Ninety individuals with the most prominent characteristics were selected from 686 laying hens according to the accumulation of lipid droplets in the liver, and were graded into three groups, including the control, mild hepatic steatosis and severe hepatic steatosis groups. A combination of transcriptome, proteome, acetylome and lipidome analyses, along with bioinformatics analysis were used to screen the key biological processes, modifications and lipids associated with hepatic steatosis.Results The rationality of the hepatic steatosis grouping was verified through liver biochemical assays and RNA-seq. Hepatic steatosis was characterized by increased lipid deposition and multiple metabolic abnormalities. Integration of proteome and acetylome revealed that differentially expressed proteins(DEPs) interacted with differentially acetylated proteins(DAPs) and were involved in maintaining the metabolic balance in the liver. Acetylation alterations mainly occurred in the progression from mild to severe hepatic steatosis, i.e., the enzymes in the fatty acid oxidation and bile acid synthesis pathways were significantly less acetylated in severe hepatic steatosis group than that in mild group(P < 0.05). Lipidomics detected a variety of sphingolipids(SPs) and glycerophospholipids(GPs) were negatively correlated with hepatic steatosis(r ≤-0.5, P < 0.05). Furthermore, the severity of hepatic steatosis was associated with a decrease in cholesterol and bile acid synthesis and an increase in exogenous cholesterol transport.Conclusions In addition to acquiring a global and thorough picture of hepatic steatosis in laying hens, we were able to reveal the role of acetylation in hepatic steatosis and depict the changes in hepatic cholesterol metabolism. The findings provides a wealth of information to facilitate a deeper understanding of the pathophysiology of fatty liver and contributes to the development of therapeutic strategies.