The InDel variants of sheep IGF2BP1 gene are associated with growth traits.

Insulin-like growth factor 2 mRNA binding protein 1 (IGF2BP1) plays positive roles in the growth, proliferation of cells and early embryos development by binding mRNA targets. Recently, it had been shown that some polymorphic loci within IGF2BP1 gene were associated with growth traits in animals, especially in goats. Therefore, it has been hypothesized that some variants within IGF2BP1 gene may be also involved in growth traits of sheep. Nine insertion/deletion (InDel) mutations within IGF2BP1 were identified and three loci were polymorphic. Meanwhile, the association analyses between three InDels and growth traits were carried out in 745 sheep. The results showed that all InDels included 5 bp InDel in downstream region, 9 bp InDel in intron 4 and 15 bp InDel in intron 2 within IGF2BP1 were significantly associated with growth traits (p<.05). Furthermore, at 5 and 9 bp InDel loci, the individuals of heterozygous genotype (ID) had superior growing performance especially at body weight (BW). In all, three InDels were crucial variants correlated with growth traits and could be applied in marker-assisted selection (MAS) in sheep.

Copy number variations of the KAT6A gene are associated with body measurements of Chinese sheep breeds.

Copy number variation (CNV) is one kind of genomic structure variations and presents as gains and losses of genomic fragments. More recently, we have made an atlas of CNV maps for livestock. In the future, it is a primary focus to determine the phenotypic effects of candidate CNVs. Lysine Acetyltransferase 6 A (KAT6A) is a protein coding gene and plays a critical role in many cellular processes. However, the effects of KAT6A CNVs on sheep body measurements remains unknown. In this study, we performed quantitative polymerase chain reaction (qPCR) to detect the presences and distributions of three CNV regions within KAT6A gene in 672 sheep from four Chinese breeds. Association analysis indicated that the three CNVs of KAT6A gene were significantly associated with body measurement(s) in Small-tailed Han sheep (STH) and Hu sheep (HU) (p < 0.05), while no effects on Large-tailed Han sheep (LTH) were observed (p > 0.05) were observed. Additionally, only one CNV was significantly associated with body measurement (body length) in Chaka sheep (CK) (p < 0.05). Our study provided evidence that the CNV(s) of KAT6A gene could be used as candidate marker(s) for molecular breedings of STH, HU, and CK breeds.

Whole-genome resequencing of Dorper and Hu sheep to reveal selection signatures associated with important traits.

Dorper and Hu sheep exhibit different characteristics in terms of reproduction, growth, and meat quality. Comparison of the genomes of two breeds help to reveal important genomic information. In this study, whole genome resequencing of 30 individuals (Dorper, DB and Hu sheep, HY) identified 15,108,125 single nucleotide polymorphisms (SNPs). Population differentiation (Fst) and cross population extended haplotype homozygosity (XP-EHH) were performed for selective signal analysis. In total, 106 and 515 overlapped genes were present in both the Fst results and XP-EHH results in HY vs DB and in DB vs HY, respectively. In HY vs DB, 106 genes were enriched in 12 GO terms and 83 KEGG pathways, such as ATP binding (GO:0005524) and PI3K-Akt signaling pathway (oas04151). In DB vs HY, 515 genes were enriched in 109 GO terms and 215 KEGG pathways, such as skeletal muscle cell differentiation (GO:0035914) and MAPK signaling pathway (oas04010). According to the annotation results, we identified a series of candidate genes associated with reproduction (UNC5C, BMPR1B, and GLIS1), meat quality (MECOM, MEF2C, and MYF6), and immunity (GMDS, GALK1, and ITGB4). Our investigation has uncovered genomic information for important traits in sheep and provided a basis for subsequent studies of related traits.

The identification and validation of target genes of IGFBP3 protein in sheep skeletal muscle cells.

This study aimed to identify the target genes of IGFBP3（insulin growth factor binding protein）protein and to investigate its target genes effects on the proliferation and differentiation of Hu sheep skeletal muscle cells. IGFBP3 was an RNA-binding protein that regulates mRNA stability. Previous studies have reported that IGFBP3 promotes the proliferation of Hu sheep skeletal muscle cells and inhibits differentiation, but the downstream genes that bind to it have not been reported yet. We predicted the target genes of IGFBP3 through RNAct and sequencing data, and verified by qPCR and RIP（RNA Immunoprecipitation）experiments, and demonstrated GNAI2（G protein subunit alpha i2）as one of the target gene of IGFBP3. After interference with siRNA, we carried out qPCR, CCK8, EdU, and immunofluorescence experiments, and found that GNAI2 can promote the proliferation and inhibit differentiation of Hu sheep skeletal muscle cells. This study revealed the effects of GNAI2 and provided one of the regulatory mechanisms of IGFBP3 protein underlying sheep muscle development.

Effects of YAP1 on proliferation and differentiation of Hu sheep skeletal muscle satellite cells in vitro.

This study aimed to understand the expression level of YAP1 in the skeletal muscle of Hu sheep and to reveal the regulatory mechanism of YAP1 on Hu sheep skeletal muscle satellite cells (SMSCs). Previous research by our group has found that YAP1 may affect the growth and development of Hu sheep skeletal muscle. In the present study, we found the expression of YAP1 in the skeletal muscle is higher than in other tissues of Hu sheep. Then, we detected the effect of YAP1 on proliferation and differentiation in Hu sheep SMSCs. According to the results of qPCR, CCK-8, EDU, and Western blot, compared to the group of negative control, overexpression of YAP1 promoted the proliferation and inhibited the differentiation of SMSCs according to the results of qPCR, CCK-8, EDU, Western blot, while the interference of YAP1 was on the contrary. Overall, our study suggests that YAP1 is an important functional molecule in the growth and development of skeletal muscle by regulating the proliferation and differentiation of SMSCs. These findings are of great use for understanding the roles of YAP1 in the skeletal muscle of Hu sheep.

SOX18 Promotes the Proliferation of Dermal Papilla Cells via the Wnt/ß-Catenin Signaling Pathway.

SRY-box transcription factor 18 (SOX18) is known to play a crucial role in the growth and development of hair follicles (HF) in both humans and mice. However, the specific effect of SOX18 on sheep hair follicles remains largely unknown. In our previous study, we observed that SOX18 was specifically expressed within dermal papilla cells (DPCs) in ovine hair follicles, leading us to investigate its potential role in the growth of hair follicles in sheep. In the present study, we aimed to examine the effect of SOX18 in DPCs and preliminarily study its regulatory mechanism through RNA-seq. We initially found that the overexpression of SOX18 promoted the proliferation of DPCs compared to the negative control group, while the interference of SOX18 had the opposite effect. To gain further insight into the regulatory mechanism of SOX18, we conducted RNA-seq analysis after knocking down SOX18 in Hu sheep DPCs. The result showed that the Wnt/ß-Catenin signaling pathway was involved in the growth process of DPC after SOX18 knockdown. Subsequently, we investigated the effect of SOX18 on the Wnt/ß-Catenin signaling pathway in DPCs using TOP/FOP-flash, qRT-PCR, and Western blot (WB) analysis. Our data demonstrated that SOX18 could activate the Wnt/ß-Catenin signaling pathway in DPCs. Additionally, we observed that SOX18 could rescue the proliferation of DPCs after inhibiting the Wnt/ß-Catenin signaling pathway. These findings underscore the essential role of SOX18 as a functional molecule governing the proliferation of DPCs. Additionally, these findings also greatly enhance our understanding of the role of SOX18 in the proliferation of DPCs and the growth of wool in Hu sheep.

Preliminary study of melatonin in the proliferation and apoptosis of Hu sheep dermal papilla cells in vitro.

Previous studies have shown that melatonin has a certain regulatory effect on the growth of sheep wool. However, the mechanism of melatonin action remains unknown. In the present study, we aimed to understand the role of exogenous melatonin in the dermal papilla cells of Hu sheep. To confirm the optimal melatonin treatment regimen for Hu sheep dermal papilla cells, we detected the cell viability by exposing them to different concentrations of melatonin and different treatment times. The results showed that cell viability was best when dermal papilla cells were treated with 1000 pg/ml of melatonin for 48 h. According to the results of qPCR, CCK-8, EDU, Western blot, and Flow cytometry analysis, we found that 1000 pg/ml melatonin promoted the proliferation and inhibited the apoptosis of dermal papilla cells compared with the exogenous melatonin blank group (control group). Furthermore, we also found that 1000 pg/ml of melatonin promoted the cell cycle progress of dermal papilla cells according to the results of qPCR and Flow cytometry analysis. Overall, our findings showed that melatonin plays an important role in the dermal papilla cells of Hu sheep.

Novel copy number variation of the BAG4 gene is associated with growth traits in three Chinese sheep populations.

Copy number variation (CNV) as an important source of genetic phenotypic and variation is related to complex phenotypic traits. The aim of this study was to investigate the potential associations of BAG4 (Bcl-2-associated athanogene 4) copy numbers variations with sheep growth traits in three Chinese sheep breeds (CKS, STHS, and HS). BAG4 is located within the stature and udder attachment quantitative trait loci (QTL) in sheep. Expression profiling revealed that the BAG4 gene was widely expressed in the tissues of sheep. The distribution of BAG4 gene copy number showed that the loss of copy number was more dominant in CKS and HS which was different from that in STHS. Statistical analysis revealed that the BAG4 CNV was significantly associated with body height in CKS (p < 0.05), with body slanting length in HS (p < 0.05), and with body height and hip cross height in STHS (p < 0.05). The χ2 values showed significant differences in the BAG4 CNV distribution frequency between varieties. In conclusion, the results establish the association between BAG4 CNV and sheep traits and suggest that BAG4 CNV may be a promising marker for the molecular breeding of Chinese sheep.

Transcriptional regulation of the bovine FGFR1 gene facilitates myoblast proliferation under hypomethylation of the promoter.

DNA methylation, which can affect the expression level of genes, is one of the most vital epigenetic modifications in mammals. Fibroblast growth factor receptor 1 (FGFR1) plays an important role in muscle development; however, DNA methylation of the FGFR1 promoter has not been studied to date in cattle. Our study focused on methylation of the FGFR1 promoter and its effect on bovine myoblast proliferation and differentiation. We identified the FGFR1 core promoter by using luciferase reporter assays; we then studied FGFR1 expression by reverse transcription quantitative polymerase chain reaction, and the methylation pattern in the FGFR1 core promoter by bisulfite sequencing polymerase chain reaction in bovine muscle tissue at three different developmental stages. We used RNAi strategy to investigate the function of FGFR1 in myoblast proliferation and differentiation. Results showed that the FGFR1 core promoters were located at the R2 (-509 to ~-202 bp) and R4 (-1295 to ~-794 bp) regions upstream of the FGFR1 gene. FGFR1 expression level was negatively associated with the degree of methylation of the FGFR1 core promoter during the developmental process. In addition, we found that FGFR1 can promote myoblast proliferation, but had no effect on myoblast differentiation. In conclusion, our results suggest that FGFR1 can promote myoblast proliferation and its transcription can be regulated by the methylation level of the core promoter. Our findings provide a mechanistic basis for the improvement of animal breeding.

Population structure, genetic diversity, and selective signature of Chaka sheep revealed by whole genome sequencing.

BACKGROUND: Chaka sheep, named after Chaka Salt Lake, are adapted to a harsh, highly saline environment. They are known for their high-grade meat quality and are a valuable genetic resource in China. Furthermore, the Chaka sheep breed has been designated a geographical symbol of agricultural products by the Chinese Ministry of Agriculture. RESULTS: The genomes of 10 Chaka sheep were sequenced using next-generation sequencing, and compared to that of additional Chinese sheep breeds (Mongolian: Bayinbuluke and Tan; Tibetan: Oula sheep) to explore its population structure, genetic diversity and positive selection signatures. Principle component analysis and a neighbor-joining tree indicated that Chaka sheep significantly diverged from Bayinbuluke, Tan, and Oula sheep. Moreover, they were found to have descended from unique ancestors (K = 2 and K = 3) according to the structure analysis. The Chaka sheep genome demonstrated comparable genetic diversity from the other three breeds, as indicated by observed heterozygosity (Ho), expected heterozygosity (He), runs of homozygosity (ROH), linkage disequilibrium (LD) decay. The enrichment analysis revealed that in contrast to Mongolian or Tibetan lineage groups, the genes annotated by specific missense mutations of Chaka sheep were enriched with muscle structure development (GO:0061061) factors including insulin-like growth factor 1 (IGF1), growth differentiation factor 3 (GDF3), histone deacetylase 9 (HDAC9), transforming growth factor beta receptor 2 (TGFBR2), and calpain 3 (CAPN3), among others. A genome-wide scan using Fst and XP-CLR revealed a list of muscle-related genes, including neurofibromin 1 (NF1) and myomesin 1 (MYOM1), under potential selection in Chaka sheep compared with other breeds. CONCLUSIONS: The comprehensive genome-wide characterization provided the fundamental footprints for breeding and management of the Chaka sheep and confirmed that they harbor unique genetic resources.

Integrative analysis of APOL3 gene CNV for adult cattle stature.

Copy number variations (CNVs) have been identified as another important structural variation of genome. In recent years, a large amount of CNVRs have been identified in humans and animals. However, association and dosage effects studies of CNVs are very limited. Apolipoprotein L3 (APOL3) gene plays a central role in modulating gene transcription and is located within a CNVR that encompasses quantitative trait locis (QTLs) for economic traits like meat quality. Herein, we analyzed the CNV polymorphism of APOL3 in 421 individuals from five distinct cattle breeds, and then correlated their genotypes with growth traits. Association analysis revealed that the APOL3 CNV was significantly associated with hip height and cannon circumference of Xianan (XN) cattle (P < .01), and visibly associated with body slanting length and hucklebone width of Pinan (PN) cattle (P < .05). Overall, the data provide evidence for the functional role of APOL3 CNV and a basis for future applications in cattle breeding.

MiR-208b regulates cell cycle and promotes skeletal muscle cell proliferation by targeting CDKN1A.

Skeletal muscle is the most abundant tissue in the body. The development of skeletal muscle cell is complex and affected by many factors. A sea of microRNAs (miRNAs) have been identified as critical regulators of myogenesis. MiR-208b, a muscle-specific miRNA, was reported to have a connection with fiber type determination. However, whether miR-208b has effect on proliferation of muscle cell was under ascertained. In our study, cyclin-dependent kinase inhibitor 1A (CDKN1A), which participates in cell cycle regulation, was predicted and then validated as one target gene of miR-208b. We found that overexpression of miR-208b increased the expression of cyclin D1, cyclin E1, and cyclin-dependent kinase 2 at the levels of messenger RNA and protein in cattle primary myoblasts in vivo and in vitro. Flow cytometry showed that forced expression of miR-208b increased the percentage of cells at the S phase and decreased the percentage of cells at the G0/G1 phase. These results indicated that miR-208b participates in the cell cycle regulation of cattle primary myoblast cells. 5-Ethynyl-20-deoxyuridine and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays showed that overexpression of miR-208b promoted the proliferation of cattle primary myoblasts. Therefore, we conclude that miR-208b participates in the cell cycle and proliferation regulation of cattle primary skeletal muscle cell through the posttranscriptional downregulation of CDKN1A.

Self-Assembled Microgels Arrays for Electrostatic Concentration and Surface-Enhanced Raman Spectroscopy Detection of Charged Pesticides in Seawater.

Development of flexible surface-enhanced Raman spectroscopy (SERS) substrate with controllable "hot spots" has spurred increasing interest because of its unique structure and plasmonic properties. Here, charged poly(vinyl alcohol) microgels containing silver nanoparticles are developed by using microfluidic emulsification to produce a "smart" SERS sensor with charge screening and signals amplification. Importantly, this charged microgel enables the selective concentration of counter-charged molecules and induces the formation of assembled arrays at an immiscible liquid-liquid interface because of the electrostatic interaction. The SERS-active microgels arrays possess controllable structures and facilitate on-site determination of charged pesticides with an enhancement factor of 5.0 × 105. Such nanostructures present the ease of assembly, stability, and reproducibility which allow multiplex detection of analytes at aqueous and organic phases without any pretreatment of the complex matrix samples. The interfacial sensing platform for on-site SERS analysis of charged pesticides will open vast possibilities for a wide range of in-field applications.

Integrating CNVs into meta-QTL identified GBP4 as positional candidate for adult cattle stature.

Copy number variation (CNV) of DNA sequences, functionally significant but yet fully ascertained, is believed to confer considerable increments in unexplained heritability of quantitative traits. Identification of phenotype-associated CNVs (paCNVs) therefore is a pressing need in CNV studies to speed up their exploitation in cattle breeding programs. Here, we provided a new avenue to achieve this goal that is to project the published CNV data onto meta-quantitative trait loci (meta-QTL) map which connects causal genes with phenotypes. Any CNVs overlapping meta-QTL therefore will be potential paCNVs. This study reported potential paCNVs in Bos taurus autosome 3 (BTA3). Notably, overview indexes and CNVs both highlighted a narrower region (BTA3 54,500,000-55,000,000 bp, named BTA3_INQTL_6) within one constructed meta-QTL. Then, we ascertained guanylate-binding protein 4 (GBP4) among the nine positional candidate genes was significantly associated with adult cattle stature, including body weight (BW, P < 0.05) and withers height (WHT, P < 0.05), fitting GBP4 CNV either with three levels or with six levels in the model. Although higher copy number downregulated the mRNA levels of GBP2 (P < 0.05) and GBP4 (P < 0.05) in 1-Mb window (54.0-55.0 Mb) in muscle and adipose, additional analyses will be needed to clarify the causality behind the ascertained association.

Correlation between ZBED6 Gene Upstream CpG Island methylation and mRNA expression in cattle.

DNA methylation is essential for the regulation of gene expression and important roles in muscle development. To assess the extent of epigenetic modifications and gene expression on the differentially methylated region (DMR) in ZBED6, we simultaneously examined DNA methylation and expression in six tissues from two different developmental stages (fetal bovine and adult bovine). The DNA methylation pattern was compared using bisulfite sequencing polymerase chain reaction (BSP) and combined bisulfite restriction analysis (COBRA). The result of quantitative real-time PCR (qPCR) analysis showed that ZBED6 has a broad tissue distribution and is highly expressed in adult bovine (P < 0.05 or P < 0.01). The DNA methylation level was significantly different in liver, lung and spleen between the two cattle groups (P < 0.05 or P < 0.01). The adult bovine group exhibited a significantly higher mRNA level and lower DNA methylation level than the fetal bovine group in liver, lung, and spleen. No significant association was detected between DNA methylation level and muscle, heart, and kidney at two different stages. In this study, the statistical analyses indicated that DNA methylation patterns are associated with mRNA level in some tissues, these results may be a useful parameter to investigate muscle developmental in cattle and as a model for studies in other species, potentially contributing to an improvement of growth performance selection in beef cattle breeding program.

A Study of the Resistance of Hu Sheep Lambs to Escherichia coli F17 Based on Whole Genome Sequencing.

This study aims to analyze the whole genome sequencing of E. coli F17 in antagonistic and susceptible Hu sheep lambs. The objective is to investigate the critical mutation loci in sheep and understand the genetic mechanism of sheep resistance to E. coli F17 at the genome level. Antagonist and susceptible venous blood samples were collected from Hu sheep lambs for whole genome sequencing and whole genome association analysis. A total of 466 genes with significant SNPs (p < 1.0 × 10-3) were found. GO and KEGG enrichment analysis and protein interaction network analysis were performed on these genes, and preliminary investigations showed that SNPs on CTNNB1, CDH8, APOD, HCLS1, Tet2, MTSS1 and YAP1 genes may be associated with the antagonism and susceptibility of Hu sheep lambs to E. coli F17. There are still some shortcomings that have not been explored via in vivo and in vitro functional experiments of the candidate genes, which will be our next research work. This study provides genetic loci and candidate genes for resistance of Hu sheep lambs to E. coli F17 infection, and provides a genetic basis for breeding disease-resistant sheep.

FecB Was Associated with Litter Size and Follows Mendel's Laws of Inheritance When It Transited to Next Generation in Suhu Meat Sheep Breeding Population.

In order to investigate the effect of FecB on litter size and growth and development traits of Suhu meat sheep and the inheritance patterns of FecB between parents and offspring in the population. In this experiment, 2241 sheep from the Suhu meat sheep population were tested for FecB using capillary electrophoresis. We combined the lambing records of 473 ewes, the growth trait records of 881 sheep at both the birth and weaning (2-month-old) stages, and the complete genealogical records of 643 lambs to analysis the distribution of FecB in the Suhu meat sheep breeding population, its effect on litter size of ewes, growth and development of lambs, and the inheritance patterns of FecB. The results showed that there were three genotypes of FecB in the Suhu meat sheep population, namely the AA genotype, AG genotype, and GG genotype. FecB in this population has a moderate polymorphism (0.25 < PIC < 0.5), and deviates from Hardy-Weinberg disequilibrium (p < 0.05). The litter size of GG genotype ewes was significantly higher than that with the AG and AA genotypes (p < 0.01). A Chi-square test showed that the inheritance patterns of FecB follows Mendel's Laws of Inheritance (p > 0.05). An association analysis of different genotypes of FecB with body weight and body size of Suhu meat sheep at birth and weaning revealed that FecB adversely affects the early growth and development of Suhu meat sheep. In summary, FecB can improve the litter size of ewes but it has negative effects on the early growth and survival rate of lambs in sheep. Therefore, FecB test results and feeding management measures should be comprehensively applied to improve the reproductive performance of ewes, the survival rate and production performance of lambs in sheep production, and thus improve the economic benefits of sheep farms.

SP1 and KROX20 Regulate the Proliferation of Dermal Papilla Cells and Target the CUX1 Gene.

Previous studies have demonstrated that CUX1 could contribute to the proliferation of DPCs in vitro, but the upstream transcriptional regulatory mechanisms of CUX1 remain largely unknown. This study aimed to investigate the upstream transcriptional regulators of CUX1 to enhance our comprehension of the mechanism of action of the CUX1 gene in ovine DPCs. Initially, the JASPAR (2024) software was used to predict the upstream target transcription factors for the CUX1 gene. Subsequently, through RT-qPCR and a double luciferase reporter assay, the interaction between SP1, KROX20, and CUX1 was established, respectively. The results indicated that SP1 and KROX20 were two highly reliable upstream transcription regulators for the CUX1 gene. Additionally, we found that SP1 promoted the proliferation of DPCs by overexpressing SP1 in DPCs, and KROX20 inhibited the proliferation of DPCs by overexpressing KROX20 in DPCs. These findings are also consistent with the transcriptional regulation of CUX1 by SP1 and KROX20, respectively. This study suggests that the effect of DPC proliferation in vitro by CUX1 may regulated by the transcription factors SP1 and KROX20.

Cell death-inducing DFFA-like effector c (CIDEC/Fsp27) gene: molecular cloning, sequence characterization, tissue distribution and polymorphisms in Chinese cattles.

Cell death-inducing DFFA-like effector c (CIDEC) protein, also known as fat specific protein 27 (Fsp27), is localized to lipid droplets. CIDEC protein is required for unilocular lipid droplet formation and optimal energy storage in addition to controlling lipid metabolism in adipocytes and hepatocytes. Research found that Ad-36 could induce lipid droplets in the cultured skeletal muscle cells and this process may be mediated by promoting CIDEC expression. The content of intermuscular fat is an important index for evaluation of beef quality, so the CIDEC gene appeared to be a candidate gene for regulation of intermuscular fat, however similar research for the bovine CIDEC gene is lacking. This paper examined the tissue expression profile of CIDEC gene in cattle using real-time RT-PCR to suggest that bovine CIDEC is highly expressed in adipose tissue. In addition, the Bovine CIDEC gene was cloned and inserted into the eukaryotic expression vector pET-28a(+), whereupon recombinant bovine CIDEC protein was induced and identified by Western-blot. A phylogenetic analysis showed that the animo acid sequence of bovine CIDEC was closer to mammalian CIDEC than rasorial CIDEC. We found ten single nucleotide polymorphisms sites (SNPs) in bovine CIDEC gene, of which SNP 2, 3, 4, 6 and 9, and SNP 8 and 10 were in complete linkage disequilibrium, respectively. SNP 1, 2 and 10 were used in further haplotype studies. Eight different haplotypes were identified in 973 cattle, of which haplotype 8 predominated with frequencies ranging from 42.90 to 54.30 %. This research provides a basis for future functional studies of CIDEC in cattle.

CRABP2 Promotes the Proliferation of Dermal Papilla Cells via the Wnt/ß-Catenin Pathway.

In our previous study of Hu sheep hair follicles, we found that CRABP2 was highly expressed in DPCs, which suggested that CRABP2 may influence the number of DPCs. In the present study, we aimed to understand the effect of CRABP2 in Hu sheep dermal papilla cells (DPCs). First, we explored the influence of CRABP2 on the ability of Hu sheep DPCs' proliferation. Based on the results obtained from some experiments, such as CCK-8, EDU, qPCR, and Western blot experiment, we found that the overexpression of CRABP2 facilitated the proliferation of DPCs compared to the negative control group. Then, we also detected the effect of CRABP2 on the Wnt/ß-catenin pathway based on the important function of the Wnt/ß-catenin pathway in hair follicles. The results showed that CRABP2 could activate the Wnt/ß-catenin pathway in DPCs, and it rescues the proliferation of DPCs when the Wnt/ß-catenin pathway was inhibited. In summary, our findings indicate that CRABP2 is a vital functional gene in the proliferation of Hu sheep DPCs. Our study will be of great use for revealing the roles of CRABP2 in the hair follicles of Hu sheep.