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The interaction between the dermal papilla cells (DPCs) and epidermal hair matrix cells (HMCs) of hair follicles (HFs) is crucial for the growth and development of HFs, but the molecular mechanism is complex and remains unclear. MicroRNAs (miRNAs) are the key signaling molecules for cellular communication. In this study, the DPCs and HMCs of yak were isolated and cultured, and the differentially expressed mRNA and miRNA were characterized to analyze the molecular basis of the interaction between DPCs and HMCs during hair follicle (HF) development in yak. The mRNA differential expression and functional enrichment analysis revealed that there were significant differences between DPCs and HMCs, and they showed the molecular functional characteristics of dermal cells and epidermal cells, respectively. Multiple KEGG pathways related to HF development were enriched in the highly expressed genes in DPCs, while the pathways associated with microbiota and immunity were significantly enriched in the highly expressed genes in HMCs. By combining analysis with our previous 10× genomics single-cell transcriptome data, 39 marker genes of DPCs of yak were identified. A total of 123 relatively specifically expressed miRNAs were screened; among these, the miRNAs associated with HF development such as miR-143, miR-214, miR-125b, miR-31, and miR-200 were presented. In conclusion, the large changes in yak DPCs and HMCs for both mRNA and miRNA expression were revealed, and numerous specifically expressed mRNAs and miRNAs in DPCs or HMCs were identified, which may contribute to the interaction and cellular communication between DPCs and HMCs during HF development in yak. [ABSTRACT FROM AUTHOR]

Background: The hair coat is available for the yak to live in the harsh environment of the plateau. Besides, improving the hair production of yak is necessary for its textile industry development. Hair grows from hair follicles (HFs). The HFs undergo periodic growth after birth and are regulated by the complex gene regulatory network. However, the molecular mechanism of HFs regeneration in the Tianzhu white yak remains unclear. RNA editing is a post-transcriptional mechanism that regulates gene expression and produces new transcripts. Hence, we investigated the influence of the A-to-I RNA editing events on the HFs cycle of the Tianzhu white yak. Results: We finally identified 54,707 adenosine-to-inosine (A-to-I) RNA editing sites (RESs) from RNA sequencing data of the HFs cycle in the Tianzhu white yak. Annotation results showed RESs caused missense amino acid changes in 7 known genes. And 202 A-to-I editing sites altered 23 target genes of 140 microRNAs. A total of 1,722 differential RESs were identified during the HFs cycle of Tianzhu white yak. GO and KEGG enrichment analysis revealed several signaling pathways and GO terms involved skin development, hair growth, and HFs cycle. Such as genes with differential RNA editing levels were significantly enriched in the peroxisome, metabolic pathways, Notch signaling pathway, and PPAR signaling pathway. Besides, the editing sites in HFs development-related genes FAS, APCDD1, WWOX, MPZL3, RUNX1, KANK2, DCN, DSC2, LEPR, HEPHL1, and PTK2B were suggested as the potential RESs involving HFs development. Conclusion: This study investigated the global A-to-I RNA editing events during the HFs cycle of yak skin tissue and expanded the knowledge of A-to-I RNA editing on the HFs cycle. Furthermore, this study revealed that RNA editing-influenced genes may regulate the HFs cycle by participating in the HFs development-related pathways. The findings might provide new insight into the regulation of RNA editing in hair growth. [ABSTRACT FROM AUTHOR]

Background: Mammalian hair play an important role in mammals' ability to adapt to changing climatic environments. The seasonal circulation of yak hair helps them adapt to high altitude but the regulation mechanisms of the proliferation and differentiation of hair follicles (HFs) cells during development are still unknown. Here, using time series data for transcriptome and hormone contents, we systematically analyzed the mechanism regulating the periodic expression of hair development in the yak and reviewed how different combinations of genetic pathways regulate HFs development and cycling. Results: This study used high-throughput RNA sequencing to provide a detailed description of global gene expression in 15 samples from five developmental time points during the yak hair cycle. According to clustering analysis, we found that these 15 samples could be significantly grouped into three phases, which represent different developmental periods in the hair cycle. A total of 2316 genes were identified in these three consecutive developmental periods and their expression patterns could be divided into 9 clusters. In the anagen, genes involved in activating hair follicle growth are highly expressed, such as the WNT pathway, FGF pathway, and some genes related to hair follicle differentiation. In the catagen, genes that inhibit differentiation and promote hair follicle cell apoptosis are highly expressed, such as BMP4, and Wise. In the telogen, genes that inhibit hair follicle activity are highly expressed, such as DKK1 and BMP1. Through co-expression analysis, we revealed a number of modular hub genes highly associated with hormones, such as SLF2, BOP1 and DPP8. They may play unique roles in hormonal regulation of events associated with the hair cycle. Conclusions: Our results revealed the expression pattern and molecular mechanisms of the seasonal hair cycle in the yak. The findings will be valuable in further understanding the alpine adaptation mechanism in the yak, which is important in order to make full use of yak hair resources and promote the economic development of pastoral plateau areas. [ABSTRACT FROM AUTHOR]

Simple Summary: Yaks can grow in extremely harsh natural environmental conditions, such as high altitude, low temperature, and hypoxia, and hair plays an important role. As an appendage of the skin, hair follicles involve the specialization of various types of cells and intercellular communication during morphogenesis. Here, we used single-cell RNA sequencing technology to identify 11 cell types from the scapular skin of yaks, constructed a transcription map of the main cells of hair follicles in the skin, and described the heterogeneity of DP cells and dermal fibroblasts in the hair follicle cycle. Our findings provide a molecular landscape of the fate determination process of dermal cell lineages. Our research provides valuable resources for further exploration of the molecular pathways involved in hair follicles. The morphogenesis of hair follicle structure is accompanied by the differentiation of skin tissue. Mammalian coats are produced by hair follicles. The formation of hair follicles requires signal transmission between the epidermis and dermis. However, knowledge of the transcriptional regulatory mechanism is still lacking. We used single-cell RNA sequencing to obtain 26,573 single cells from the scapular skin of yaks at hair follicle telogen and anagen stages. With the help of known reference marker genes, 11 main cell types were identified. In addition, we further analyzed the DP cell and dermal fibroblast lineages, drew a single-cell map of the DP cell and dermal fibroblast lineages, and elaborated the key genes, signals, and functions involved in cell fate decision making. The results of this study provide a very valuable resource for the analysis of the heterogeneity of DP cells and dermal fibroblasts in the skin and provide a powerful theoretical reference for further exploring the diversity of hair follicle cell types and hair follicle morphogenesis. [ABSTRACT FROM AUTHOR]

Sperm cryodamage caused by cryopreservation limits the use of frozen yak spermatozoa in artificial insemination (AI). However, the proteomic changes involved in the cryodamage of yak spermatozoa have not been investigated to date. Therefore, this study aimed to identify proteins related to freezing tolerance. Tandem mass tag (TMT) were used in combination with liquid chromatography–tandem mass spectrometry (LC-MS/MS) for identifying differentially expressed proteins (DEPs) between high-motility (HM) and low-motility (LM) frozen-thawed yak spermatozoa. A total of 116 DEPs were identified (>1.5-fold, P < 0.05); of which, 104 proteins were upregulated in HM spermatozoa and 12 proteins were upregulated in LM spermatozoa. The results of functional annotation analysis revealed that the DEPs were mainly involved in metabolic processes. A total of 20 DEPs that were abundantly expressed in HM spermatozoa were strongly associated with carbohydrate metabolism. The results of KEGG analysis revealed that the DEPs were enriched in glycolysis/gluconeogenesis, PPAR signaling pathway, and Ras signaling pathway. In addition, many antioxidant enzymes such as superoxide dismutase (SOD1), peroxiredoxin-6 (PRDX6), and Parkinson disease protein 7 (PARK7) were upregulated in HM spermatozoa, suggesting that these enzymes affect the motility of spermatozoa by regulating the levels of reactive oxygen species (ROS) in frozen-thawed spermatozoa. Altogether, the findings of this study elucidate the mechanisms through which cryopreservation affects the movement of yak spermatozoa and offer a novel basis for refining freezing techniques and modifying cryopreservation extender components. • The impact of cryopreservation on yak spermatozoa quality was firstly investigated. • Comparison of the sperm proteomic in high and low motility frozen-thawed yak spermatozoa. • The differentially expressed proteins are mainly related to antioxidant and energy metabolism. • This study may help us understand the mechanism of sperm cryodamage. [ABSTRACT FROM AUTHOR]

Fat deposition is very important to the growth and reproduction of yaks. In this study, the effect of the feeding system on fat deposition in yaks was explored by transcriptomics and lipidomics. The thickness of the subcutaneous fat in yaks under stall (SF) and graze feeding (GF) was evaluated. The transcriptomes and lipidomes of the subcutaneous fat in yaks under different feeding systems were detected by RNA-sequencing (RNA-Seq) and non-targeted lipidomics based on ultrahigh-phase liquid chromatography tandem mass spectrometry (UHPLC-MS), respectively. The differences in lipid metabolism were explored, and the function of differentially expressed genes (DEGs) was evaluated by gene ontology (GO) and Kyoto encyclopedia of genes and genome (KEGG) analysis. Compared with GF yaks, SF yaks possessed stronger fat deposition capacity. The abundance of 12 triglycerides (TGs), 3 phosphatidylethanolamines (PEs), 3 diglycerides (DGs), 2 sphingomyelins (SMs) and 1 phosphatidylcholine (PC) in the subcutaneous fat of SF and GF yaks was significantly different. Under the mediation of the cGMP–PKG signaling pathway, the blood volume of SF and GF yaks may be different, which resulted in the different concentrations of precursors for fat deposition, including non-esterified fatty acid (NEFA), glucose (GLU), TG and cholesterol (CH). The metabolism of C16:0, C16:1, C17:0, C18:0, C18:1, C18:2 and C18:3 in yak subcutaneous fat was mainly realized under the regulation of the INSIG1, ACACA, FASN, ELOVL6 and SCD genes, and TG synthesis was regulated by the AGPAT2 and DGAT2 genes. This study will provide a theoretical basis for yak genetic breeding and healthy feeding. [ABSTRACT FROM AUTHOR]

Testicular development is a tightly regulated process in mammals. Understanding the molecular mechanisms of yak testicular development will benefit the yak breeding industry. However, the roles of different RNAs, such as mRNA, lncRNA, and circRNA in the testicular development of yak, are still largely unclear. In this study, transcriptome analyses were performed on the expression profiles of mRNAs, lncRNAs, and circRNAs in testis tissues of Ashidan yak at different developmental stages, including 6-months-old (M6), 18-months-old (M18), and 30-months-old (M30). A total of 30, 23, and 277 common differentially expressed (DE) mRNAs, lncRNAs, and circRNAs were identified in M6, M18, and M30, respectively. Furthermore, functional enrichment analysis showed that the common DE mRNAs during the entire developmental process were mainly involved in gonadal mesoderm development, cell differentiation, and spermatogenesis processes. Additionally, co-expression network analysis identified the potential lncRNAs related to spermatogenesis, e.g., TCONS_00087394 and TCONS_00012202. Our study provides new information about changes in RNA expression during yak testicular development, which greatly improves our understanding of the molecular mechanisms regulating testicular development in yaks. [ABSTRACT FROM AUTHOR]

Background: Normal testicular development is highly crucial for male reproduction and is a precondition for spermatogenesis that is the production of spermatozoa in the testes. MiRNAs have been implicated in several testicular biological processes, including cell proliferation, spermatogenesis, hormone secretion, metabolism and reproductive regulation. In the present study, we used deep sequencing data to study the functions of miRNAs during testicular development and spermatogenesis, by analyzing the expression patterns of small RNAs in 6-, 18- and 30-month-old yak testis tissues. Results: A total of 737 known and 359 novel miRNAs were obtained from 6-, 18- and 30-month-old yak testes. In all, we obtained 12, 142 and 139 differentially expressed (DE) miRNAs in 30- vs. 18-, 18- vs. 6-, and 30- vs. 6-month-old testes, respectively. Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of all DE miRNA target genes revealed BMP2, TGFB2, GDF6, SMAD6, TGFBR2 and other target genes as participants in different biological processes, including TGF-β, GnRH, Wnt, PI3K–Akt, MAPK signaling pathways and several other reproductive pathways. In addition, quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) was used to detect the expression of seven randomly selected miRNAs in 6-, 18- and 30-month-old testes, and the results were consistent with the sequencing data. Conclusions: The differential expression of miRNAs in yak testes at different development stages was characterized and investigated using deep sequencing technology. We believe that the results will contribute to further understanding the functions of miRNAs in regulating the development of yak testes and improving the reproductive performance of male yaks. [ABSTRACT FROM AUTHOR]

The development of hair follicles in yak shows significant seasonal cycles. In our previous research, transcriptome data including mRNAs and lncRNAs in five stages during the yak hair follicles (HFs) cycle were detected, but their regulation network and the hub genes in different periods are yet to be explored. This study aimed to screen and identify the hub genes during yak HFs cycle by constructing a mRNA-lncRNA co-expression network. A total of 5000 differently expressed mRNA (DEMs) and 729 differently expressed long noncoding RNA (DELs) were used to construct the co-expression network, based on weighted genes co-expression network analysis (WGCNA). Four temporally specific modules were considered to be significantly associated with the HFs cycle of yak. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that the modules are enriched into Wnt, EMC-receptor interaction, PI3K-Akt, focal adhesion pathways, and so on. The hub genes, such as FER, ELMO1, PCOLCE, and HOXC13, were screened in different modules. Five hub genes (WNT5A, HOXC13, DLX3, FOXN1, and OVOL1) and part of key lncRNAs were identified for specific expression in skin tissue. Furthermore, immunofluorescence staining and Western blotting results showed that the expression location and abundance of DLX3 and OVOL1 are changed following the process of the HFs cycle, which further demonstrated that these two hub genes may play important roles in HFs development. [ABSTRACT FROM AUTHOR]

Background: As the direct organ of villus, hair follicles have obvious seasonal cycles. The hair follicle cycle is orchestrated by multiple cell types that together direct cell renewal and differentiation. But the regulation property of hair follicle cells from anagen to catagen in yak is still unknown. Results: In this study, single-cell RNA sequencing was performed on 24,124 single cells of the scapular skin from white yak. Based on tSNE cluster analysis, the cell types of IFE-DC, epidermal cell lines, fibroblasts, keratinocytes, IRS, DS, INFU, and other cells in yak hair follicles during anagen and catagen were successfully identified, and the gene expression profiles were described. The GO enrichment analysis indicated the different cells characteristic genes to be mainly enriched in the epidermal development, epithelial cell differentiation and wound healing pathways. The pseudotime trajectory analysis described the differentiation trajectory of the epidermal lineage and dermal lineage of the hair follicle during anagen and catagen. Moreover, the dynamic changes of the genes like LHX2, KRT25, and KRT71 were found to be highly expressed in HS and IRS, but not in the IFE-DC, INFU, and keratinocyte during differentiation. Conclusions: Our results analyzed the time-varying process of gene expression in the dermal cell lineage and epidermal cell lineage of hair follicles during anagen and catagen during fate differentiation was expounded at the single cell level, revealing the law of fate specialization of different types of cells. In addition, based on the enrichment analysis, the transcriptional regulatory factors involved in the different cell fates were also revealed. These results will help to enhance our understanding of yak hair follicle cycle and promote the development and utilization of yak villus. [ABSTRACT FROM AUTHOR]

Background: Ovarian follicle fluid (FF) as a microenvironment surrounding oocyte plays critical roles in physio-biochemical processes of follicle development and oocyte maturation. It is hypothesized that proteins in yak FF participate in the physio-biochemical pathways. The primary aims of this study were to find differentially expressed proteins (DEPs) between mature and immature FF, and to elucidating functions of the mature and immature FF in yak. Results: The mature and immature FF samples were obtained from three healthy yaks that were nonpregnant, aged from four to five years, and free from any anatomical reproductive disorders. The FF samples were subjected to mass spectrometry with the isobaric tags for relative and absolute quantification (iTRAQ). The FF samples went through correlation analysis, principle component analysis, and expression pattern analysis based on quantification of the identified proteins. Four hundred sixty-three DEPs between mature and immature FF were identified. The DEPs between the mature and immature FF samples underwent gene ontology (GO), Kyoto encyclopedia of genes and genomes (KEGG), and protein-protein interaction (PPI) analysis. The DEPs highly expressed in the mature FF mainly took parts in the complement and coagulation cascades, defense response, acute-phase response, response to other organism pathways to avoid invasion of exogenous microorganisms. The complement activation pathway contains eight DEPs, namely C2, C5, C6, C7, C9, C4BPA, CFH, and MBL2. The three DEPs, CATHL4, CHGA, and PGLYRP1, take parts in defense response pathway to prevent invasion of exogenetic microorganism. The coagulation cascades pathway involves many coagulation factors, such as F7, F13A1, FGA, FGB, FGG, KLKB1, KNG1, MASP1, SERPINA1, and SERPIND1. While the DEPs highly expressed in the immature FF participated in protein translation, peptide biosynthetic process, DNA conformation change, and DNA geometric change pathways to facilitate follicle development. The translation pathway contains many ribosomal proteins, such as RPL3, RPL5, RPS3, RPS6, and other translation factors, such as EIF3J, EIF4G2, ETF1, MOV10, and NARS. The DNA conformation change and DNA geometric change involve nine DEPs, DDX1, G3BP1, HMGB1, HMGB2, HMGB3, MCM3, MCM5, MCM6, and RUVBL2. Furthermore, the expressed levels of the main DEPs, C2 and SERPIND1, were confirmed by western blot. Conclusions: The differential proteomics revealed the up-regulated DEPs in mature FF take parts in immunoreaction to prevent invasion of microorganisms and the up-regulated DEPs in immature FF participate in protein synthesis, which may improve our knowledge of the follicular microenvironment and its biological roles for reproductive processes in yak. The DEPs, C2 and SERPIND1, can be considered as protein markers for mature yak follicle. [ABSTRACT FROM AUTHOR]

Normal testicular development plays a crucial role in male reproduction and is the precondition for spermatogenesis. PIWI-interacting RNAs (piRNAs) are novel noncoding RNAs expressed in animal germ cells that form complexes with PIWI family proteins and are involved in germ cell development, differentiation, and spermatogenesis. However, changes in piRNA expression profiles during early testicular development in yak have not been investigated. In this study, we used small RNA sequencing to evaluate the differences and potential functions of piRNA expression profiles in 6-, 18-, and 30-month-old yak testis tissues. Differential expression analysis found 109, 293, and 336 differentially expressed piRNAs in M30 vs. M18, M18 vs. M6, and M30 vs. M6, respectively, and found 30 common differentially expressed piRNAs in the three groups of M6, M18, and M30. In addition, the functional enrichment analysis of differentially expressed piRNAs target genes indicated that they were related to testicular development and spermatogenesis. Finally, we detected the expression of the PIWI protein family in the yak testis at different developmental stages and found that PIWIL1, PIWIL2, PIWIL3, and PIWIL4 were highly expressed in 18- and 30-month-old yak testis and almost not expressed in 6-month-old yak testis. In conclusion, this study summarizes the changes of piRNA expression patterns during the early development of yak testis and provides new clues for the regulatory role of piRNA in yak testis. [ABSTRACT FROM AUTHOR]

Simple Summary: Single-cell sequencing technology can reflect cell population heterogeneity at the single-cell level, leading to a better understanding of the role of individual cells in the microenvironment. Over the past few years, single-cell sequencing technology has not only made more new discoveries in the study of cellular heterogeneity of other rare cells such as stem cells, but has also become the most powerful research method for embryonic development, organ differentiation, cancer occurrence, and cell mapping. In this review, we outline the use of scRNA-seq in hair follicles. In particular, by focusing on landmark studies and the recent discovery of novel subpopulations of hair follicles, we summarize the phenotypic diversity of hair follicle cells and their links to hair follicle morphogenesis. Enhancing our understanding of the progress of hair follicle research will help to elucidate the regulatory mechanisms that determine the fate of different types of cells in the hair follicle, thereby guiding hair loss treatment and hair-producing economic animal breeding research. Single-cell sequencing technology can fully reflect the heterogeneity of cell populations at the single cell level, making it possible for us to re-recognize various tissues and organs. At present, the sequencing study of hair follicles is transiting from the traditional ordinary transcriptome level to the single cell level, which will provide diverse insights into the function of hair follicle cells. This review focuses on research advances in the hair follicle microenvironment obtained from scRNA-seq studies of major cell types in hair follicle development, with a special emphasis on the discovery of new subpopulations of hair follicles by single-cell techniques. We also discuss the problems and current solutions in scRNA-seq observation and look forward to its prospects. [ABSTRACT FROM AUTHOR]

Background: The hair follicle development process is regulated by sophisticated genes and signaling networks, and the hair grows from the hair follicle. The Tianzhu white yak population exhibits differences in hair length, especially on the forehead and shoulder region. However, the genetic mechanism is still unclear. Isoform sequencing (Iso-seq) technology with advantages in long reads sequencing. Hence, we combined the Iso-seq and RNA-seq methods to investigate the transcript complexity and difference between long-haired yak (LHY) and normal-haired yak (NHY). Results: The hair length measurement result showed a significant difference between LHY and NHY on the forehead and the shoulder (P-value < 0.001). The skin samples from the forehead and the shoulder of LHY and NHY were pooled for isoform sequencing (Iso-seq). We obtained numerous long transcripts, including novel isoforms, long non-coding RNA, alternative splicing events, and alternative polyadenylation events. Combined with RNA-seq data, we performed differential isoforms (DEIs) analysis between LHY and NHY. We found that some hair follicle and skin development-related DEIs, like BMP4, KRT2, IGF2R, and COL1A2 in the forehead skin; BMP1, KRT1, FGF5, COL2A1, and IGFBP5 in the shoulder skin. Enrichment analysis revealed that DEIs in both two comparable groups significantly participated in skin and hair follicle development-related pathways, like ECM-receptor interaction, focal adhesion, and PI3K-Akt signaling pathways. The results indicated that the hair follicle development of Tianzhu white yak may influence the hair length difference. Besides, the protein-protein interaction (PPI) network of DEIs showed COL2A1 and COL3A1 exhibited a high degree of centrality, and these two genes were suggested as potential candidates for the hair length growth of Tianzhu white yak. Conclusions: The results provided a comprehensive analysis of the transcriptome complexity and identified differential transcripts that enhance our understanding of the molecular mechanisms underlying the variation in hair length growth in Tianzhu white yak. [ABSTRACT FROM AUTHOR]

Background: Ovarian follicle fluid (FF) as a microenvironment surrounding oocyte plays critical roles in physio-biochemical processes of follicle development and oocyte maturation. It is hypothesized that proteins in yak FF participate in the physio-biochemical pathways. The primary aims of this study were to find differentially expressed proteins (DEPs) between mature and immature FF, and to elucidating functions of the mature and immature FF in yak. Results: The mature and immature FF samples were obtained from three healthy yaks that were nonpregnant, aged from four to five years, and free from any anatomical reproductive disorders. The FF samples were subjected to mass spectrometry with the isobaric tags for relative and absolute quantification (iTRAQ). The FF samples went through correlation analysis, principle component analysis, and expression pattern analysis based on quantification of the identified proteins. Four hundred sixty-three DEPs between mature and immature FF were identified. The DEPs between the mature and immature FF samples underwent gene ontology (GO), Kyoto encyclopedia of genes and genomes (KEGG), and protein-protein interaction (PPI) analysis. The DEPs highly expressed in the mature FF mainly took parts in the complement and coagulation cascades, defense response, acute-phase response, response to other organism pathways to avoid invasion of exogenous microorganisms. The complement activation pathway contains eight DEPs, namely C2, C5, C6, C7, C9, C4BPA, CFH, and MBL2. The three DEPs, CATHL4, CHGA, and PGLYRP1, take parts in defense response pathway to prevent invasion of exogenetic microorganism. The coagulation cascades pathway involves many coagulation factors, such as F7, F13A1, FGA, FGB, FGG, KLKB1, KNG1, MASP1, SERPINA1, and SERPIND1. While the DEPs highly expressed in the immature FF participated in protein translation, peptide biosynthetic process, DNA conformation change, and DNA geometric change pathways to facilitate follicle development. The translation pathway contains many ribosomal proteins, such as RPL3, RPL5, RPS3, RPS6, and other translation factors, such as EIF3J, EIF4G2, ETF1, MOV10, and NARS. The DNA conformation change and DNA geometric change involve nine DEPs, DDX1, G3BP1, HMGB1, HMGB2, HMGB3, MCM3, MCM5, MCM6, and RUVBL2. Furthermore, the expressed levels of the main DEPs, C2 and SERPIND1, were confirmed by western blot. Conclusions: The differential proteomics revealed the up-regulated DEPs in mature FF take parts in immunoreaction to prevent invasion of microorganisms and the up-regulated DEPs in immature FF participate in protein synthesis, which may improve our knowledge of the follicular microenvironment and its biological roles for reproductive processes in yak. The DEPs, C2 and SERPIND1, can be considered as protein markers for mature yak follicle. [ABSTRACT FROM AUTHOR]

Simple Summary: Yaks are a major economic source for people in the Tibetan Plateau region. Yaks are rich in nutrients, but their low fat content is not conducive to the large-scale promotion of yak meat, so the study of the mechanism of yak lipid deposition is beneficial to the marketing of yak meat. In this paper, the results of transcriptome sequencing analysis of yak liver showed that it could be determined that the expression levels of genes associated with partial lipid deposition were significantly up-regulated during yak fattening. In addition, this study found that the tenderness of yak meat improved during this process. Fattening significantly affects fat deposition in yaks, which may be realized through its effects on lipid metabolic pathways. Therefore, studying the mechanism of lipid deposition in yaks and fattening yaks will improve the quality of yak meat. Yak meat is nutritionally superior to beef cattle but has a low fat content and is slow-growing. The liver plays a crucial role in lipid metabolism, and in order to determine whether different feeding modes affect lipid metabolism in yaks and how it is regulated, we employed RNA sequencing (RNA-seq) technology to analyze the genome-wide differential gene expression in the liver of yaks maintained under different raising systems. A total of 1663 differentially expressed genes (DEGs) were identified (|log2FC| ≥ 0 and p-value ≤ 0.05), including 698 down-regulated and 965 up-regulated genes. According to gene ontology (GO) and KEGG enrichment analyses, these DEGs were significantly enriched in 13 GO terms and 26 pathways (p < 0.05). Some DEGs were enriched in fatty acid degradation, PPAR, PI3K-Akt, and ECM receptor pathways, which are associated with lipid metabolism. A total of 16 genes are well known to be related to lipid metabolism (e.g., APOA1, FABP1, EHHADH, FADS2, SLC27A5, ACADM, CPT1B, ACOX2, HMGCS2, PLIN5, ACAA1, IGF1, FGFR4, ALDH9A1, ECHS1, LAMA2). A total of 11 of the above genes were significantly enriched in the PPAR signaling pathway. The reliability of the transcriptomic data was verified using qRT-PCR. Our findings provide new insights into the mechanisms regulating yak meat quality. It shows that fattening improves the expression of genes that regulate lipid deposition in yaks and enhances meat quality. This finding will contribute to a better understanding of the various factors that determine yak meat quality and help develop strategies to improve yield and quality. [ABSTRACT FROM AUTHOR]

Background: Yaks (Bos grunniens), prized for their ability to thrive in high-altitude environments, are indispensable livestock in the plateau region. Modifying their feeding systems holds significant promise for improving their growth and meat quality. Tenderness, a key determinant of yak meat quality and consumer appeal, is demonstrably influenced by dietary regimen. Indoor feeding regimes have been shown to enhance tenderness by lowering shear stress and optimizing pH values. CircRNAs, well-known modulators of circulatory function, also play a crucial role in skeletal muscle development across various animal species. However, their functional significance in yak skeletal muscle remains largely unexplored. Results: In this study, we identified a total of 5,534 circRNAs within the longissimus dorsi muscle, and we found 51 differentially expressed circRNAs (20 up-regulated and 31 down-regulated) between the two feeding groups. Constructing a comprehensive ceRNA network illuminated intricate regulatory mechanisms, with PGP and circRNA_0617 converging on bta-miR-2285q, mirrored by KLF15/circRNA_0345/bta-miR-20b and CTSF/circRNA_0348/bta-miR-146a. These findings shed light on the potential of circRNAs to influence yak muscle development and meat quality, offering valuable insights for future research. Conclusions: This investigation unraveled a complex interaction network between circRNAs、mRNAs and miRNAs in yak skeletal muscle. We further elucidated the target genes regulated by these target genes within the network, offering valuable insights into the potential regulatory mechanisms governing muscle development and meat quality-related traits in yaks. [ABSTRACT FROM AUTHOR]

PaLi yak is a very important local breed in China; as a year-round grazing animal, it plays a very important role for the economic and native herdsmen. The PaLi yak complete mitochondrial DNA is sequenced in this study, the total length is 16,324 bp, containing 13 protein-coding genes, 22 tRNA genes, 2 rRNA genes and a non-coding control region (D-loop region). The order and composition are similar to most of the other vertebrates. The base contents are: 33.72% A, 25.80% C, 13.21% G and 27.27% T; A + T (60.99%) was higher than G + C (39.01%). The phylogenetic relationships were analyzed using the complete mitogenome sequence, results showed that the genetic relationship between yak and cattle is distinct. These information provides useful data for further study on protection of genetic resources and the taxonomy of Bovinae. [ABSTRACT FROM AUTHOR]

Background and aim: Mitochondrial genome has aseries of characteristics such as simple structure, no recombination, maternalinheritance, stable structure, fast evolution rate, and high copy number. Moreover, it is easy to be sequenced,contains high-resolution phylogenetic information, and exists in a wide rangeof taxa. Therefore, it is widely used in the study of biological phylogeny. Atpresent, phylogenetic studies focus mainly on D-loop region, cytochrome b gene,and protein-coding sequence. Phylogenetic studies using the mitochondrialcomplete sequence are rarely reported in yak. Therefore, the present studyaimed to construct phylogenetic tree using yak mitochondrial complete sequenceand compare the subsequent results with previous findings obtained usingpartial sequences. Results: Complete mitochondrial sequences of five yakpopulations from Qinghai and Xinjiang were obtained. The mitotype diversity ofthe five populations was Xueduo yak (0.992 ± 0.015), Pamir yak (0.990 ± 0.014),Yushu yak (0.963 ± 0.033), Qilian yak (0.948 ± 0.036), and Huanhu yak (0.905 ±0.048), which showed a higher mitotype diversity compared with other breeds fromthe previous reports, including Jiulong yak, Maiwa yak, Zhongdian yak, andTianzhu yak. A total of 78 mitotypes were obtained from 111 individuals. Amongthese, Yushu yak, Huanhu yak, Xueduo yak, and Qilian yak all shared mitotypes,but the Pamir yak did not share mitotypes with these four populations.Phylogenetic analysis showed that yak populations were separable into threedistinct branches. The analysis identified a new phylogenetic branch containingboth wild and domestic yaks. The 155 mitotypes found in 206 individuals weredivided into 3 haplogroups by mitotype clustering. Thehaplogroup was not associated with the geographical distribution of yaks. Theyaks in the same population or the same ecological environment were distributedin different haplogroups. Among the threehaplogroups, haplogroup A and haplogroup B showed a star-shaped distribution ofmitotypes. The central mitotypes were widely distributed and had a highfrequency. Conclusions: Thegenetic diversity of yaks in Qinghai was high. Both domestic and wild yaks clusteredinto three branches. [ABSTRACT FROM AUTHOR]

Simple Summary: The methylation level of testicular tissue in young yaks and adult yaks was determined, and the N6-methyladenosine (m6A) level was significantly increased after sexual maturity. Functional enrichment analysis of differentially methylated mRNAs screened between the two groups after sequencing showed that they mainly regulated cell development, spermatogenesis, and testicular endocrine function. Functional analysis of the differentially expressed genes screened between the two groups showed that they were involved in the biological processes of mitosis, meiosis, and flagellated sperm motility. Finally, several genes related to spermatogenesis were screened for further verification. Studying the mechanism of spermatogenesis is key to exploring the reproductive characteristics of male yaks. Although N6-methyladenosine (m6A) RNA modification has been reported to regulate spermatogenesis and reproductive function in mammals, the molecular mechanism of m6A in yak testis development and spermatogenesis remains largely unknown. Therefore, we collected testicular tissue from juvenile and adult yaks and found that the m6A level significantly increased after sexual maturity in yaks. In MeRIP-seq, 1702 hypermethylated peaks and 724 hypomethylated peaks were identified. The hypermethylated differentially methylated RNAs (DMRs) (CIB2, AK1, FOXJ2, PKDREJ, SLC9A3, and TOPAZ1) mainly regulated spermatogenesis. Functional enrichment analysis showed that DMRs were significantly enriched in the adherens junction, gap junction, and Wnt, PI3K, and mTOR signaling pathways, regulating cell development, spermatogenesis, and testicular endocrine function. The functional analysis of differentially expressed genes showed that they were involved in the biological processes of mitosis, meiosis, and flagellated sperm motility during the sexual maturity of yak testis. We also screened the key regulatory factors of testis development and spermatogenesis by combined analysis, which included BRCA1, CREBBP, STAT3, and SMAD4. This study indexed the m6A characteristics of yak testicles at different developmental stages, providing basic data for further research of m6A modification regulating yak testicular development. [ABSTRACT FROM AUTHOR]

Reference 1 Zheng Q, Ye N, Bao P. Construction of transcriptome atlas of white yak hair follicle during anagen and catagen using single-cell RNA sequencing. 6 Dynamic changes in the gene expression during specialization of hair follicle fibroblasts and DS. a Construction of the pseudotime differentiation trajectory between the fibroblasts and DS; b Gene expression during fibroblast specialization; c Expression of the characteristic genes in different stages of fibroblast; d GO enrichment analysis of fibroblast characteristic genes; e Gene expression during DS specialization; f GO enrichment analysis of the characteristic genes in DS; g Interaction analysis of the characteristic genes in DS; h Immunofluorescence analysis of the hair follicles The original article [[1]] has been corrected. [Extracted from the article]

Simple Summary: Yak are an important source of produce and living materials for plateau herdsmen. However, the population of yak is small and their reproductive capacity is low, so it is very important to improve their reproductive potential. The epididymis is an important place for sperm maturation and storage, but the differential expression of the cauda epididymis before and after sexual maturity in yak has not been characterized. In this study, the key genes regulating epididymal cauda development and sperm maturation were screened by RNA-seq and proteomics. These results ultimately provide a theoretical basis for how to improve the reproductive potential of yak. Epididymis development is the basis of male reproduction and is a crucial site where sperm maturation occurs. In order to further understand the epididymal development of yak and how to regulate sperm maturation, we conducted a multi-omics analysis. We detected 2274 differential genes, 222 differential proteins and 117 co-expression genes in the cauda epididymis of yak before and after sexual maturity by RNA-seq and proteomics techniques, which included TGFBI, COL1A1, COL1A2, COL3A1, COL12A1, SULT2B1, KRT19, and NPC2. These high abundance genes are mainly related to cell growth, differentiation, adhesion and sperm maturation, and are mainly enriched via extracellular matrix receptor interaction, protein differentiation and absorption, and lysosome and estrogen signaling pathways. The abnormal expression of these genes may lead to the retardation of epididymal cauda development and abnormal sperm function in yak. In conclusion, through single and combined analysis, we provided a theoretical basis for the development of the yak epididymal cauda, sperm maturation, and screening of key genes involved in the regulation of male yak reproduction. [ABSTRACT FROM AUTHOR]

The ovary as one of the most dynamic organs produces steroids to orchestrate female secondary sexual characteristics, harbors ovarian reserve for oocytes, releases mature oocytes for fertilization, and maintains pregnancy. Yak (Bos grunniens) is the only bovid animal that can adapt to the harsh climatic conditions on the Qinghai‐Tibetan Plateau (altitudes of over 3000 m above sea level). However, the cellular atlas is composed of oocytes and other somatic cells, and their individual molecular characteristics remain to be elucidated in the yak ovary. Here, single‐cell RNA sequencing (scRNA‐seq) was performed to delineate the molecular signature of various cell types in the yak ovarian cortex. A cellular atlas of yak ovarian cortex was constructed successfully on the basis of the differentially expressed genes (DEGs) from the distinct cell types and their functional enrichment analysis, comprising endothelial cells, nature kill cells, stromal cells, smooth muscle cells, oocytes, macrophages, epithelial cells, and granulosa cells. Meanwhile, the signature genes were determined based on their expression specificity in each cell type. A cell‐to‐cell communication network was built in light of the differentially overexpressed ligand and receptor genes from each cell type. Further, the oocytes were subdivided into four subtypes based on their individual DEGs and the functional enrichment of the DEGs. FST and TOP2A were identified as maker genes for oocytes by immunostaining in the yak ovarian cortex. The cellular atlas reveals the biological characteristics of the ovarian cortex at the cellular molecular level and provides insights into female reproductive biology via cellular communications in the yak. [ABSTRACT FROM AUTHOR]

Simple Summary: Exosomes are multivesicular bodies produced and secreted by living cells as membranous vesicles with a diameter of about 30–150 nm, density of 1.13–1.19 g/mL, and a typical "cup dish" shape. Exosomes carry proteins, miRNA, lncRNA, circRNA, and mRNA and their degradation fragments are involved in intracellular signal transduction important for regulation of cellular activities. Of these, miRNAs play an important role in the regulation of gene expression in important biological processes such as cell division, tissue development, and immune response. In this study, milk quality was determined and exosome miRNAs from yak and Jeryak milk sampled in June and December were characterized by sequencing. Differentially expressed miRNAs (DEMs) and pathways related to lactation performance were identified. In summary, this study provides data that can be used to develop alternative strategies for improving the lactation performance of yaks and Jeryaks. The Jeryak is the hybrid offspring of yaks and Jersey cattle and exhibit improved milk and meat yields. Biomolecules carried within milk exosomes are important for cell growth, development, immune regulation, and various pathophysiological processes. Previous studies showed that miRNAs regulate mammary gland development, lactation, and milk quality. This study explored the relationship between milk exosomes miRNAs and lactation performance. A comparison of the milk content showed that yak milk was of a better quality compared to Jeryak milk (casein, fat, TS, SNF, lactose). Milk collected in December was superior to that collected in June for both yak and Jeryak, except for lactose concentrations. Exosomes were extracted by density gradient centrifugation and miRNA expression profiles in milk exosomes from three yaks and three Jeryaks collected in June and December were detected by small RNA sequencing. In all, 22, 120, 78, and 62 differentially expressed miRNAs (DEMs) were identified in Jun_ JY vs. Jun_ Y (P1: Jeryak in June vs. Yak in June), Jun_ JY vs. Dec_ JY (P2: Jeryak in June vs. Jeryak in December), Dec_ JY vs. Dec_ Y (P3: Jeryak in December vs. Yak in December), and Jun_ Y vs. Dec_ Y (P4: Yak in June vs. Yak in December) groups. These DEMs were enriched in functions and signaling pathways related to lactation performance. In conclusion, these findings are a reference tool to study the molecular basis of lactation performance. [ABSTRACT FROM AUTHOR]

Simple Summary: The SOX6 (sex determining region Y-box 6) gene belongs to one of the transcription factors in the SRY (sex-determining region Y) family, which affects sex determination, embryonic and nervous system development, bone and various organ formation. In the previous study, the whole-genome sequencing was used to detect multiple genes located in the copy number variation region, including SOX6 gene. In this study, we identified the correlation between the growth traits and CNV of SOX6 in 311 Ashidan yaks. The results showed that SOX6-CNV was significantly correlated with the chest girth of the 6-months old yaks (p < 0.05) and 30-months yaks (p < 0.05), and withers height of 6 months yaks (p < 0.05) and 18-months yaks (p < 0.05), suggesting the SOX6-CNV affect growth traits in yaks, and could be new markers for the selection of yak breeding. Copy number variation (CNV) is a fundamental type of structural variation of the genome affecting the economic traits of livestock. The SOX6 gene (sex-determining region Y-box 6), as a transcription factor, has multiple functions with regard to sex determination, embryonic growth, the nervous system development, as well as bone, and various organ formation. This study employed quantitative real-time fluorescence quota PCR (qPCR) for detecting the SOX6-CNV of the 311 Ashidan yaks and analyzed the correlation of the SOX6-CNV with four phenotypes (including body weight, withers height, body length, and chest girth) of the yaks aged 6, 12, 18, and 30 months using ANOVA and multiple comparisons. Furthermore, the SOX6 gene expression was identified in seven different tissues of the yaks. The experiment results demonstrated the expression of SOX6 in each tissue, and the kidney and muscle tissue were found to have higher relative expression levels. Based on the processing by IBM SPSS software, SOX6-CNV was significantly correlated with the chest girth of the 6-months old yaks (p < 0.05) and 30-months yaks (p < 0.05), and withers height of 6 months yaks (p < 0.05) and 18-months yaks (p < 0.05), as well as the normal type of CNV, was chosen for yak breeding. In conclusion, SOX6 might be prominently involved in promoting growth and development of yaks, suggesting that the SOX6 gene can be used in breeding yaks by molecular marker-assisted selection (MAS). The study also offered some important insights into the references and clues for the genetic breeding of yaks. [ABSTRACT FROM AUTHOR]

Cattle-yak is a hybrid F1 generation of cattle and yak, which has a history of more than 3000 years and has shown better production performance and higher economic benefits than those of yaks. However, up to now, there has been no study on the transcriptome-wide m6A methylation profile of bovine skeletal muscle and its potential biological function during muscle development. Here, we observed significant changes in the expression levels of muscle-related marker genes and methylation-related enzymes during the development of cattle-yak, and the overall m6A content in the Longissimus dorsi muscle of 18-month-old cattle-yak decreased significantly. A total of 36,602 peaks, 11,223 genes and 8388 lncRNAs were identified in the two groups, including 2989 differential peaks (427 up-regulated peaks and 2562 down-regulated peaks), 1457 differentially expressed genes (833 up-regulated genes and 624 down-regulated genes) and 857 differentially expressed lncRNAs (293 up-regulated lncRNAs and 564 down-regulated lncRNAs). GO and KEGG analysis revealed that they were significantly enriched in some muscle-related pathways (Wnt signaling pathway and MAPK signaling pathway) and high-altitude adaptation-related pathway (HIF-1 signaling pathway). Moreover, m6A abundance was positively correlated with gene expression levels, while it was negatively correlated with lncRNA expression levels. This indicates that m6A modification played an important role in the Longissimus dorsi muscle development of cattle-yak; however, the regulation mechanism of m6A-modified mRNA and lncRNA may be different. This study was the first report of transcriptome-wide m6A-modified mRNAs and lncRNAs atlas in the Longissimus dorsi muscle development of cattle-yak, one which will provide new perspectives for genetic improvement in bovines. [ABSTRACT FROM AUTHOR]

Simple Summary: Yaks are among the largest high-altitude mammals in the world, and they are ideally adapted to the harsh environmental conditions of the plateau regions. Yaks are thus central to the lives of herdsmen and other local populations in these high-altitude areas. Copy number variations (CNVs) are an important cause of genomic variation in livestock and identifying advantageous CNVs can aid in livestock breeding efforts. In this study, an association between CNVs in the MICALL2 and MOGAT2 genes and Ashidan yak growth traits was confirmed, providing a theoretical foundation for Ashidan yak breeding and meat production efforts. Copy number variations (CNVs) are a result of genomic rearrangement affecting DNA regions over 1 kb in length, and can include inversions, translocations, deletions, and duplications. The molecule interacting with CasL-like protein 2 (MICALL2) gene is primarily associated with mitochondrial protein targeting and exhibits predicted stress fiber colocalization. The monoacylglycerol O-acyltransferase 2 (MOGAT2) gene encodes an enzyme responsible for catalyzing diacylglycerol synthesis from 2-monoacylglycerol and fatty acyl-CoA. For this study, blood samples were obtained from 315 yaks, and the body weight, body length, withers height, and chest girth of these animals were measured at 6, 12, 18, and 30 months of age. Genomic DNA was harvested from the collected blood samples, and CNVs in these samples were detected by qPCR. The resultant data were compared using ANOVAs, revealing significant associations between MICALL2 gene CNVs and body weight at 6 months of age (p < 0.05), body length and chest girth at 30 months of age (p < 0.05), and withers height at 18 months of age (p < 0.01) in Ashidan yaks. Similarly, MOGAT2 CNVs were significantly associated with body weight at 6 and 30 months of age (p < 0.05), and with withers height at 18 months of age (p < 0.01) in these Ashidan yaks. MICALL2 and MOGAT2 gene expression was further analyzed in yak tissue samples, revealing that MICALL2 was most highly expressed in the adipose tissue, whereas MOGAT2 was most highly expressed in the lung. These results thus confirmed the relationship between CNVs in the MICALL2 and MOGAT2 genes and Ashidan yak growth traits, providing a valuable gene locus that can be leveraged for future marker-assisted yak breeding efforts. [ABSTRACT FROM AUTHOR]

Simple Summary: Kecai yaks are an important genetic resource in Gansu Province, China. These yaks, produced through the crossbreeding of Gannan yaks and wild yaks, exhibit excellent production performance and high fecundity. Despite these beneficial characteristics, the genetic characteristics, population structure, and other properties of Kecai yaks have not been effectively characterized to date. Accordingly, this study was developed to explore the genetic diversity and population structure of Kecai yaks through whole-genome resequencing. These analyses ultimately revealed the status of Kecai yaks as an independent group within the overall Chinese yak population. Kecai yaks are regarded as an important genetic resource in China owing to their high fecundity and flavorful meat. However, the genetic characteristics of Kecai yaks have not been effectively characterized to date, and the relationship between Kecai yaks and other yak breeds remains to be fully characterized. In this paper, the resequencing of the Kecai yak genome is performed leading to the identification of 11,491,383 high-quality single nucleotide polymorphisms (SNPs). Through principal component, phylogenetic, and population genetic structure analyses based on these SNPs, Kecai yaks were confirmed to represent an independent population of yaks within China. In this study, marker and functional enrichment analysis of genes related to positive selection in Kecai yak was carried out, and the results show that such selection in Kecai yaks is associated with the adaptation to alpine environments and the deposition of muscle fat. Overall, these results offer a theoretical foundation for the future utilization of Kecai yak genetic resources. [ABSTRACT FROM AUTHOR]

Although significant advancement has been made in the study of adipogenesis, knowledge about how chromatin accessibility regulates yak adipogenesis is lacking. We here described genome-wide dynamic chromatin accessibility in preadipocytes and adipocytes by using the assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq), and thus revealed the unique characteristics of open chromatin during yak adipocyte differentiation. The chromatin accessibility of preadipocytes and adipocytes exhibited a similar genomic distribution, displaying a preferential location within the intergenic region, intron, and promoter. The pathway enrichment analysis identified that genes with differential chromatin accessibility were involved in adipogenic metabolism regulation pathways, such as the peroxisome proliferator activated receptor-γ (PPAR) signaling pathway, wingless-type MMTV integration site (Wnt) signaling pathway, and extracellular matrix-receptor (ECM–receptor) interaction. Integration of ATAC-seq and mRNA-seq revealed that genes with a high expression were associated with high levels of chromatin accessibility, especially within 1 kb upstream and downstream of the transcription start site. In addition, we identified a series of transcription factors (TFs) related to adipogenesis and created the TF regulatory network, providing the possible interactions between TFs during yak adipogenesis. This study is crucial for advancing the understanding of transcriptional regulatory mechanisms of adipogenesis and provides valuable information for understanding the adaptation of plateau species to high-altitude environments by maintaining whole body homeostasis through fat metabolism. [ABSTRACT FROM AUTHOR]

Cattle–yak is a hybrid offspring resulting from the crossbreeding of yak and cattle, and it exhibits substantial heterosis in production performance. However, male sterility in cattle–yak remains a concern. Reports suggest that noncoding RNAs are involved in the regulation of spermatogenesis. Therefore, in this study, we comprehensively compared testicular transcription profiles among cattle, yak, and cattle–yak. Numerous differentially expressed genes (DEGs), differentially expressed circRNAs (DECs), and differentially expressed miRNAs (DEMs) were identified in the intersection of two comparison groups, namely cattle versus cattle–yak and yak versus cattle–yak, with the number of DEGs, DECs, and DEMs being 4968, 360, and 59, respectively. The DEGs in cattle–yaks, cattle, and yaks were mainly associated with spermatogenesis, male gamete generation, and sexual reproduction. Concurrently, GO and KEGG analyses indicated that DEC host genes and DEM source genes were involved in the regulation of spermatogenesis. The construction of a potential competing endogenous RNA network revealed that some differentially expressed noncoding RNAs may be involved in regulating the expression of genes related to testicular spermatogenesis, including miR-423-5p, miR-449b, miR-34b/c, and miR-15b, as well as previously unreported miR-6123 and miR-1306, along with various miRNA–circRNA interaction pairs. This study serves as a valuable reference for further investigations into the mechanisms underlying male sterility in cattle–yaks. • Cattle-yak male sterility. • Cattle versus cattle-yak and yak versus cattle-yak intersection had more DEGs, DECs and DEMs. • ncRNA is involved in regulating the expression of testicular spermatogenesis-related genes. [ABSTRACT FROM AUTHOR]

Alternative splicing is a prevalent phenomenon in testicular tissues. Due to the low assembly accuracy of short-read RNA sequencing technology in analyzing post-transcriptional regulatory events, full-length (FL) transcript sequencing is highly demanded to accurately determine FL splicing variants. In this study, we performed FL transcriptome sequencing of testicular tissues from 0.5, 1.5, 2.5, and 4-year-old yaks and 4-year-old cattle-yaks using Oxford Nanopore Technologies. The obtained sequencing data were predicted to have 47,185 open reading frames (ORFs), including 26,630 complete ORFs, detected 7645 fusion transcripts, 15,355 alternative splicing events, 25,798 simple sequence repeats, 7628 transcription factors, and 35,503 long non-coding RNAs. A total of 40,038 novel transcripts were obtained from the sequencing data, and the proportion was almost close to the number of known transcripts identified. Structural analysis and functional annotation of these novel transcripts resulted in the successful annotation of 9568 transcripts, with the highest and lowest annotation numbers in the Nr and KOG databases, respectively. Weighted gene co-expression network analysis revealed the key regulatory pathways and hub genes at various stages of yak testicular development. Our findings enhance our comprehension of transcriptome complexity, contribute to genome annotation refinement, and provide foundational data for further investigations into male sterility in cattle-yaks. [ABSTRACT FROM AUTHOR]

Background: High spontaneous miscarriage rate in yak, especially during late pregnancy, have caused a great economic loss to herdsmen living in the Qinghai-Tibet plateau. However, the mechanism underlying spontaneous miscarriage is still poorly understood. In the present study, placenta protein markers were identified to elucidate the pathological reasons for yak spontaneous miscarriage through isobaric tags for relative and absolute quantification (iTRAQ) proteomic technology and bioinformatic approaches. Results: Subsequently, a total of 415 differentially expressed proteins (DEPs) were identified between aborted and normal placentas. The up-regulated DEPs in the aborted placentas were significantly associated with "spinocerebellar ataxia", "sphingolipid signalling", "relaxin signalling", "protein export", "protein digestion and absorption" and "aldosterone synthesis and secretion" pathway. While the down-regulated DEPs in the aborted placentas mainly participated in "valine, leucine and isoleucine degradation", "PPAR signalling", "peroxisome", "oxidative phosphorylation", "galactose metabolism", "fatty acid degradation", "cysteine and methionine metabolism" and "citrate cycle" pathway. Conclusions: The results implied that the identified DEPs could be considered as placental protein markers for yak miscarriage during late pregnancy, and biomacromolecule metabolic abnormality and oxidative damage might be responsible for the high spontaneous miscarriage rate in yak. These findings provide an important theoretical basis for deciphering the pathologic mechanism of late spontaneous miscarriage in yak. [ABSTRACT FROM AUTHOR]

Simple Summary: As a domestic animal living in the Qinghai-Tibet Plateau at high altitudes from 2000 to 5000 m, the growth rate of the yak is slow under extremely harsh natural environmental conditions, such as high altitude, low temperatures, and low oxygen. Compared with conventional selection, using molecular markers in breeding to improve yak growth traits is more efficient. Many studies have indicated that CNV mutations can significantly affect the phenotypic traits of livestock and poultry. This study explored the association between the growth traits and CNVs of SOX5 and SOX8 in 326 Ashidan yaks. Our results showed that CNVs and the CNV combination of SOX5 and SOX8 were significantly associated with withers height and chest girth (p < 0.05), suggesting these mutations could be new markers for the selection of yak growth traits. Copy number variation (CNV) is a structural variant with significant impact on genetic diversity. CNV has been widely used in breeding for growth traits, meat production or quality, and coat color. SRY-like box genes (SOXs) are a class of transcription factors that play a regulatory role in cell fate specification and differentiation. SOX5 and SOX8 belong to subgroups D and E of the SOXs, respectively. Previous studies have shown that SOX5 and SOX8 are essential in the development of bones. In this study, we explored the association between the growth traits and CNVs of SOX5 and SOX8 in 326 Ashidan yaks and detected mRNA expression levels in different tissues. Our results illustrated that CNVs of SOX5 and SOX8 were significantly associated with withers height at 18 months of age and chest girth at 30 months of age (p < 0.05). The CNV combination of SOX5 and SOX8 was significantly associated with withers height at 18 months of age (p < 0.01). SOX5 expression in the lung was significantly higher than in the heart, spleen, kidney, and muscle (p < 0.05). SOX8 expression in the lung was significantly higher than in the liver and muscle (p < 0.05). Our results provide evidence that the CNVs of SOX5 and SOX8 genes could be used as new markers for the selection of yak growth traits. [ABSTRACT FROM AUTHOR]

Sperm-associated antigen 17 (SPAG17) gene encodes a central pair protein, which is involved in flagellar motility, male fertility and skeletal growth in ruminants. The insertions/deletions (indels) and copy number variations (CNVs) influence phenotypic traits by altering the sequences and copy numbers of functional genes, respectively. This study identified a novel 8-bp indel of SPAG17 gene in 1520 individuals from eight different cattle breeds, as well as a novel CNV region in 355 animals. The correlation analysis of indel showed that the individuals of ID genotype had superior performance traits such as body height (p = 0.038) and body slanting length (p = 0.041) as compared to other genotypes in Xianan cattle. For the CNV, different copy numbers were closely related to the body height in Qinchuan (p = 0.045) and body weight in Xianan (p = 0.036) breeds. Importantly, significant difference was observed between the 8-bp indel and the copy number loss in Xianan breed (p < 0.01). These findings indicated that the variations within the bovine SPAG17 gene can be considered as an effective DNA molecular marker for beef cattle breeding. [ABSTRACT FROM AUTHOR]

Simple Summary: In mammals, the testis-specific serine/threonine kinase (TSSK) is essential for spermatogenesis and male fertility. This study aimed to analyze the possible alternative spliceosomes of the TSSK4 gene in the yak testis tissue using PCR amplification and cloning techniques. A total of six alternative spliceosomes were obtained, of which there were only two alternative spliceosomes in the yak testis before sexual maturity and four alternative spliceosomes in the yak testis after sexual maturity. In mammals, the testis-specific serine/threonine kinase (TSSK) is essential for spermatogenesis and male fertility. TSSK4 belongs to the family of the testis-specific serine/threonine-protein kinase (TSSK), with a crucial role in spermatogenesis. This study aimed to analyze the variable spliceosome of the TSSK4 gene in the yak for understanding the regulatory function of the TSSK4 spliceosome in yak testis development using PCR amplification and cloning techniques. The GST pull-down was used for pulling down the protein interacting with TSSK4, and then the protein interacting with TSSK4 was identified using LC–MS/MS. The results of the PCR amplification demonstrated multiple bands of the TSSK4 gene in the yak. The cloning and sequencing yielded a total of six alternative spliceosomes, which included only two alternative spliceosomes before sexual maturity and four alternative spliceosomes after sexual maturity. The sub-cells of the alternative spliceosomes were found to localize in the nucleus before sexual maturity and in the cytoplasm after sexual maturity. The LC–MS/MS analysis of the alternative spliceosome with the highest expression after sexual maturity yielded a total of 223 interacting proteins. The enrichment analysis of the 223 interacting proteins revealed these proteins participate in biological processes, cell composition, and molecular functions. The KEGG analysis indicated that the TSSK4-interacting protein participates in the estrogen signaling pathways, tight junctions, endoplasmic reticulum protein processing, and other signaling pathways. This study cloned the six alternative spliceosomes of the TSSK4 gene laying the foundation for studying the function of each spliceosome in the future. [ABSTRACT FROM AUTHOR]

Background and aim: Yak estrus is a seasonal phenomenon, probably involving epigenetic regulation of synthesis and secretion of sex hormones as well as growth and development of follicles. N6-methyladenosine (m6A) is the most common internal modification of the eukaryotic mRNA. However, there are no detailed reports on the m6A transcriptome map of yak ovary. Therefore, this study aimed to collected the yak ovarian tissues at three different states of anestrus (YO-A), estrus (YO-F), and pregnancy (YO-P), and obtained the full transcriptome m6A map in yak by MeRIP-seq. Results: The HE staining revealed that the number of growing follicles and mature follicles in the ovary during the estrus period was relatively higher than those in the anestrus period and the pregnancy period. The RT-qPCR showed that the expression of METTL3, METTL14, FTO, YTHDC1 were significantly different across different periods in the ovaries, which suggests that m6A may play a regulatory role in ovarian activity. Next, we identified 20,174, 19,747 and 13,523 m6A peaks in the three ovarian samples of YO-A, YO-F and YO-P using the methylated RNA immunoprecipitation sequencing (MeRIP-seq). The m6A peaks are highly enriched in the coding sequence (CDS) region and 3′untranslated region (3′UTR) as well as the conserved sequence of "RRACH." The GO, KEGG and GSEA analysis revealed the involvement of m6A in many physiological activities of the yak's ovary during reproductive cycle. The association analysis found that some genes such as BNC1, HOMER1, BMP15, BMP6, GPX3, and WNT11 were related to ovarian functions. Conclusions: The comparison of the distribution patterns of methylation peaks in the ovarian tissues across different periods further explored the m6A markers related to the regulation of ovarian ovulation and follicular development in the yak ovary. This comprehensive map provides a solid foundation for revealing the potential function of the mRNA m6A modification in the yak ovary. [ABSTRACT FROM AUTHOR]

Residents on the Tibetan Plateau intake a lot of yak subcutaneous fat by diet. Modern healthy diet ideas demand higher unsaturated fatty acids (UFAs), especially polyunsaturated fatty acid (PUFA) content in meat. Here, the gas chromatography (GC) and tandem mass tag (TMT) proteomic approaches were applied to explore the relationship between the proteomic differences and UFA and PUFA content in the subcutaneous fat of yaks with different sex. Compared with male yaks (MYs), the absolute contents of UFAs, monounsaturated fatty acids (MUFAs) and PUFAs in the subcutaneous fat of female yaks (FYs) were all higher (p < 0.01); the relative content of MUFAs and PUFAs in MY subcutaneous fat was higher, and the value of PUFAs/SFAs was above 0.4, so the MY subcutaneous fat is more healthy for consumers. Further studies showed the transcriptional regulation by peroxisome proliferator-activated receptor delta (PPARD) played a key role in the regulation of UFAs, especially PUFA content in yaks of different sex. In FY subcutaneous fat, the higher abundance of the downstream effector proteins in PPAR signal, including acyl-CoA desaturase (SCD), elongation of very-long-chain fatty acids protein 6 (ELOVL6), lipoprotein lipase (LPL), fatty acid-binding protein (FABP1), very-long-chain (3R)-3-hydroxyacyl-CoA dehydratase 3 (HACD3), long-chain fatty acid CoA ligase 5 (ACSL5) and acyl-CoA-binding protein 2 (ACBP2), promoted the UFAs' transport and synthesis. The final result was the higher absolute content of c9-C14:1, c9-C18:1, c9,c12-C18:2n-6, c9, c12, c15-C18:3n-3, c5, c8, c11, c14, c17-C20:5n-3, c4, c7, c10, c13, -c16, c19-C22:6n-3, UFAs, MUFAs and PUFAs in FY subcutaneous fat. Further, LPL, FABP1, HACD3, ACSL1 and ACBP2 were the potential biomarkers for PUFA contents in yak subcutaneous fat. This study provides new insights into the molecular mechanisms associated with UFA contents in yak subcutaneous fat. [ABSTRACT FROM AUTHOR]

• This is the first systematic study of the keratin family in yak. 53 keratins are identified in yak. • The results show that yak keratins have the same conserved domain and sequence as other species keratins. • The study demonstrates the expression pattern of keratins genes throughout the hair cycle and identifies five different expression patterns. • Nine hub keratin genes are identified by WGCNA method in the whole hair cycle. As the largest subgroup of intermediate filament proteins, keratins are divided into two types of subfamily. Currently, the molecular mechanism of keratins in several animals has been reported but is limited in yak. Here, 53 different kinds of keratins were identified in the yak genome, including 23 type I and 30 type II keratins. Bioinformatics analysis in this study revealed that multiple phosphorylation sites were identified among all the family members. And the subcellular localization of these proteins was predicted to be in the nucleus, cytoskeleton, and cytoplasm. All keratin family proteins were unstable and the scores of instability coefficient were higher than 40. Phylogenetic analysis showed that high consistency results of the sequence conservation and grouping were found in the genomes of yak, sheep, cattle, mouse, rat, and human. Based on the expression patterns obtained from the transcriptome data, keratin genes (KRTs) were grouped into five clusters, and results also showed that KRTs were highly activated in skin tissues during the hair cycle in yak. Among the five clusters, Cluster II contained the most KRTs , which was the main expression pattern of the yak hair follicle cycle, followed by Cluster III. These results indicated the transition period from telogen to anagen and catagen to telogen were highly dynamic in yak. Gene expression correlation analysis showed that KRTs exhibited a strong correlation (mainly positive correlation) throughout the hair follicle development cycle. And the identification of hub KRTs in specific modules related to hair follicle development in this study was performed using the Weight Gene Co-Expression Network Analysis (WGCNA). Specific modules that include KRTs were darkgreen (KRT40), darkgrey (KRT5), turquoise (KRT1 , KRT2 , KRT10), bisque4 (KRT4), thistle2 (KRT9 , KRT39), and yellowgreen (KRT24). The interaction network showed that these genes were found to be related to the regulation of cell cycle, melanogenesis, hair follicle development, keratinocyte proliferation. Our study provides theoretical support for the study of the evolutionary relationship and molecular mechanism of keratin family in B. grunnien. [ABSTRACT FROM AUTHOR]

Luteinizing hormone receptor (LHR), prolactin receptor (PRLR), growth hormone (GH) and insulin‐like growth factor 1 (IGF1) have been shown to be key regulators of germ cell development. However, the role of LHR, PRLR, GH and IGF1 in the development of yak testis remains unclear. In this study, we aimed to describe and compare gene expression and protein localization of LHR, PRLR, GH and IGF1 in the development of yak testes. Testes were collected from 6, 24, 36 and 72 months yak, and the kidney, liver, testicular, lung, skeletal muscle, heart and spleen tissues were collected from 36 months yak. The quantitative real‐time PCR (qRT‐PCR) results showed that the expression of these four genes was widely expressed in kidney, liver, testicular, lung, skeletal muscle, heart and spleen, while the LHR and PRLR were highly expressed in the kidney, skeletal muscle and testis, and higher levels of GH and IGF were expressed in spleen and testis. Moreover, the mRNA expression of these genes in adults was higher than in pre‐pubertal yak. In the testis, the LHR‐, PRLR‐, GH‐ and IGF1‐positive signals were detected in the Leydig cells of the 6 months, while the intense positive signals were discovered in Leydig cells, spermatogonia and spermatocytes of the 36 and 72 months. Thus, LHR, PRLR, GH and IGF1 may be involved in the development of spermatids and spermatocytes, and in the regulation of spermatogonia proliferation and Leydig cell function. [ABSTRACT FROM AUTHOR]

AbstractLysozyme like 4 (LYZL4), lysozyme like 6 (LYZL6) and proliferating cell nuclear antigen (PCNA) are implicated in the regulation of testicular function, but there was no research reported available on the expression patterns of LYZL4, LYZL6 and PCNA genes at different developmental stages of yak testes. In this study, we used the qRT-PCR, western blotting and immunohistochemistry estimated the LYZL4, LYZL6 and PCNA gene expression and protein lo-calization at different developmental stages of yak testes. The qPCR results showed that the mRNA expression of LYZL4, LYZL6 and PCNA genes significantly increased with age in the testes of yaks. Western blot results showed that the protein abundance of LYZL4, LYZL6 and PCNA in yak testes was significantly higher after puberty than before puberty. Furthermore, the results of immunohistochemistry indicated that LYZL4, LYZL6 and PCNA may be involved in the regulation of spermatogonia proliferation and Leydig cell function in immature testis. In adult yak testes, LYZL4, LYZL6 and PCNA may involve in the development of round spermatids and primary spermatocytes during testicular development. Our results indicated that LYZL4, LYZL6 and PCNA may be involved in the development of Sertoli cells, Leydig cells and gonocytes in yak testes. [ABSTRACT FROM AUTHOR]

Cattle-yak, which is the hybrid F 1 generation of cattle and yak, demonstrates better production performance compared to yak. However, there is limited research on the molecular mechanisms responsible for the muscle development of cattle-yak. To address this knowledge gap, a comprehensive transcriptomic survey of the longissimus dorsi muscle in cattle-yak was conducted. Three transcript types, namely lncRNAs, miRNAs, and circRNAs, along with protein-coding genes were characterized at two developmental stages (6 m, 18 m) of cattle-yak. The results revealed significant enrichment of these transcripts into pathways related to myoblast differentiation and muscle development signaling. Additionally, the study identified the TCONS00024465/circHIPK3-bta-miR-499- ADAMTS6 regulatory network, which may play a crucial role in the muscle development of cattle-yak by combining the transcriptome data of yak and constructing the ceRNA co-expression network. HEK 293 T cells were used to validate that TCONS00024465 and circHIPK3 are located upstream of bta-miR-499, and can competitively bind to bta-miR-499 as ceRNA. The study also verified that ADAMTS6 regulates skeletal muscle development by inhibiting myoblast proliferation, promoting myoblast differentiation, and positively regulating the apoptosis of myoblasts. Taken together, this study provides new insights into the advantages of cattle-yak production performance and offers a molecular basis for further research on muscle development. • Cattle-yak, as the hybrid animal, exhibits better production performance compared to yak. • The first transcriptome profile of cattle-yak revealed the key role of TCONS00024465/circHIPK3-bta-miR-499- ADAMTS6 regulatory network. • TCONS00024465 and circHIPK3 competitively bind bta-miR-499, thereby targeting ADAMTS6. • ADAMTS6 regulates skeletal muscle development by inhibiting myoblast proliferation and promoting myoblast differentiation. • New insights into the molecular basis of cattle-yak muscle development. [ABSTRACT FROM AUTHOR]

In mammals, epigenetic modifications involving DNA methylation are necessary for the completion of the cell differentiation process. However, the global DNA methylation landscape and its dynamics during yak adipocyte differentiation remain unexplored. Here, we performed whole-genome bisulfite sequencing (WGBS) to asses DNA methylation in yak preadipocytes and adipocytes, combining these results with those of our previous studies on changes in chromatin accessibility and gene expression during yak adipogenesis. The results showed that CG methylation levels were lower in promoter than in exon and intron, and gradually decreasing from the distal regions to transcription start site (TSS). There was a significant negative correlation between CG methylation levels located in promoter and gene expression levels. The 46 genes shared by CG differentially methylated regions (DMRs) and differential chromatin accessibility were significantly enriched in Hedgehog and PI3K-Akt signaling pathways. ATAC-seq peaks with high chromatin accessibility located in both promoter (≤ 2 kb from TSS) and distal (> 2 kb from TSS) regions corresponded to low methylation levels. Taken together, these findings demonstrated that DNA methylation and its interactions with chromatin accessibility regulate gene expression during yak adipocyte differentiation, contributing to the understanding of mechanisms of various epigenetic modifications and their interactions in adipogenesis. [ABSTRACT FROM AUTHOR]

Ruminant rumen plays an important role in the digestibility of cellulose, hemicellulose, starch and fat. In this study, the yaks under graze and stall feeding were chosen as the models of different rumen bacteria and intramuscular fat (IMF). The characteristics of IMF deposition, serum indexes in yaks were detected; the bacteria, metabolites in rumen was explored by 16S rRNA sequencing technology, untargeted metabolomics based on liquid chromatography-mass spectrometer and gas chromatography, respectively; the transcriptome of longissimus thoracis was identified by RNA-Sequencing analysis. Based on above results, a hypothesis that yak IMF deposition is regulated by the combined action of microbiome-gut-brain and muscle axis was proposed. The short-chain fatty acids (SCFAs) and neurotransmitters precursors like acetylcholine produced in yak rumen promoted insulin secretion via central nervous system. These insulin resulted in the high expression of SREBF1 gene by gut-brain axis; SCFAs can directly arrive to muscular tissue via blood circulation system, then activated the expression of PPARγ gene by gut-muscle axis. The expression of lipogenesis gene SCD , FABP3 , CPT1 , FASN and ACC2 was accordingly up-regulated. This study firstly introduce the theory of microbiome-gut-brain/muscle axis into the study of ruminant, and comprehensively expounded the regulatory mechanism of yak IMF deposition. • Microbial-gut-muscle and brain axis in livestock were verified for the frist time. • The insulin secretion via CNS was increased under the regulation of gut-brain axis mediated by SCFAs and neurotransmitters from the microbial fermentation in yaks rumen. • SCFAs also directly arrive to muscular tissue by blood circulation system via gut-muscle axis. • The up-regulated expression of lipogenesis gene PPARγ , SREBF1 , SCD , FABP3 , CPT1 , FASN and ACC2 promotes IMF fat deposition in yaks. • Microbial-gut-muscle and brain axis also regulate the fatty acids composition in yak IMF. [ABSTRACT FROM AUTHOR]

The current study aimed to investigate the damage of long-term high concentrate diet feeding pattern on Yak jejunal structure, physiological function and protein composition during cold season. Twelve Datong male Yak (Bos grunniens) with the same age from cold season were randomly selected and slaughtered to determine Yak jejunal digestive enzyme activity, morphology and protein composition on different feeding patterns in Tibetan Plateau. The results showed that Yak jejunum digestive enzyme activity and morphology of grazing reared group were better than those in the intensively reared group. A total of 96 differentially expressed proteins were identified by label-free Mass Spectrometry (MS), which could be concluded to two predominant themes: protein structure and inflammatory response. Nine differentially expressed proteins were correlated in Yak jejunum damage in different feeding patterns. According to this research, we found that feeding pattern resulted the differences in Yak jejunum physiological function, morphology and protein composition. This fact was confirmed long-term high dietary concentrate feeding could damage the jejunum epithelial morphology and function. [ABSTRACT FROM AUTHOR]

The variety of species can be efficiently increased by interspecific hybridization. However, because the males in the hybrid progeny are usually sterile, this heterosis cannot be employed when other cattle and yaks are hybridized. While some system-level studies have sought to explore the etiological basis for male cattle-yak sterility, no systematic cellular analyses of this phenomenon have yet been performed. Here, single-cell RNA sequencing and UPHLC-MS/MS targeted metabolomics methods were used to study the differences in testicular tissue between 4-year-old male yak and 4-year-old male cattle-yak, providing new and comprehensive insights into the causes of male cattle-yak sterility. Cattle-yak testes samples detected 6 somatic cell types and one mixed germ cell type. Comparisons of these cell types revealed the more significant differences in Sertoli cells (SCs) and [Leydig cells and myoid cells (LCs_MCs)] between yak and cattle-yak samples compared to other somatic cell clusters. Even though the LCs and MCs from yaks and cattle-yaks were derived from the differentiation of the same progenitor cells, a high degree of overlap between LCs and MCs was observed in yak samples. Still, only a small overlap between LCs and MCs was observed in cattle-yak samples. Functional enrichment analyses revealed that genes down-regulated in cattle-yak SCs were primarily enriched in biological activity, whereas up-regulated genes in these cells were enriched for apoptotic activity. Furthermore, the genes of up-regulated in LCs_MCs of cattle-yak were significantly enriched in enzyme inhibitor and molecular function inhibitor activity. On the other hand, the genes of down-regulated in these cells were enriched for signal receptor binding, molecular function regulation, positive regulation of biological processes, and regulation of cell communication activity. The most significant annotated differences between yak and cattle-yak LCs_MCs were associated with cell-to-cell communication. While yak LCs_MCs regulated spermatogenic cells at spermatogonia, spermatocyte, and spermatid levels, no such relationships were found between cattle-yak LCs_MCs and germ cells. This may suggest that the somatic niche in male cattle-yak testes is a microenvironment that is ultimately not favorable for spermatogenesis. [ABSTRACT FROM AUTHOR]

Simple Summary: In this study, yak's TLR4 gene alternative spliceosomes were investigated using PCR amplification and cloning with an aim to improve disease-resistance in yaks and promote efficient utilization of yak's resources. qRT-PCR was used to evaluate the expression levels of two alternatively spliced TLR4 transcripts in seven distinct yak tissues. To predict the function of proteins expressed by each TLR4 spliceosome, TLR4 protein structure and function were analyzed bioinformatically. Besides, two alternative spliceosomes of yak's TLR4 gene were also identified, which were in line with predicted variants of the TLR4 gene in NCBI. These two alternative spliceosomes of the TLR4 gene were expressed in each tissue; however, the expression levels of these spliceosomes were significantly different in different tissue. We also observed that deletion of exon-2 in TLR4 affected the function of the corresponding protein. This study will lay a theoretical foundation for future studies on the role of two variants of yak's TLR4 gene in disease resistance. Besides, data from this study could be analyzed further to explore the molecular mechanism associated with disease-resistance in the yak. In this study, the yak's TLR4 gene alternative spliceosomes were investigated using PCR amplification and cloning to improve disease-resistance in yak and promote efficient utilization of yak's resources. qRT-PCR was used to determine the expression levels of two alternatively spliced transcripts of the TLR4 gene in seven distinct tissues. To predict the function of proteins expressed by each TLR4 spliceosome, bioinformatic analysis of yak's TLR4 protein structure and function was performed, which led to the identification of two alternative spliceosomes of yak's TLR4 gene. The TLR4-X1 sequence length was 2526 bp, and it encoded full-length TLR4 protein (841 amino acids). The sequence length of the exon-2 deleted TLR4-X2 sequence was 1926 bp, and it encoded truncated TLR4 protein (641 amino acids). TLR4-X2 sequence was consistent with the predicted sequence of the TLR4 gene in GenBank. Each tissue showed significantly different expression levels of these two alternative spliceosomes. As per the bioinformatic analysis of the structure and function of TLR4 protein, deletion of exon-2 in the TLR4 gene resulted in frameshift mutations of the reading frame in the corresponding protein, which altered its ligand-binding and active sites. Besides, biological property such as substrate specificity of truncated TLR4 protein was also altered, leading to altered protein function. This study has laid a theoretical foundation for exploring the role of two variants of the TLR4 gene in yak's disease resistance. Besides, this study's data could be analyzed further to explore the molecular mechanism associated with disease-resistance in the yak. [ABSTRACT FROM AUTHOR]

A previous study using next-generation sequencing illustrated and confirmed that the cholinergic receptor muscarinic 3 (CHRM3) gene resides within CNVRs of yak populations that overlap with quantitative trait loci (QTL) relating to meat quality and growth, and plays a prominent role in smooth muscle contraction. This study was conducted to determine the distribution of the CNVs of the CHRM3 gene in five Chinese domestic yak breeds and to investigate the influences of the CHRM3 gene on the gene expression and growth traits of the Datong yak. The data were analyzed by ANOVA using SPSS software. Our results revealed that a gain CNV type in the CHRM3 gene is more highly associated (P < 0.05) with various growth traits in six-month-old and five-year-old Datong yaks. The gain CNV type of the CHRM3 gene more frequently occurred than loss and normal types in all yak breeds. Nevertheless, the CHRM3 gene copy number had a highly significant (P <0.0012) and moderately negative correlation with the mRNA transcription levels were determined in the skeletal muscle of Chinese Datong yaks. Therefore, we concluded that this study provided inclusive information on the application of CNVs of the CHRM3 in growth traits and its possible function. [ABSTRACT FROM AUTHOR]

Simple Summary: Genomic selection is a new technology in animal breeding after the selection according to the best linear unbiased prediction (BLUP) value and marker assisted selection. Genomic selection has gradually been used in practical applications over recent years following the advent of high-density single nucleotide polymorphism (SNP) chips for livestock and poultry. Yak are a critical species on the Qinghai–Tibet Plateau, which is of great significance to herders. The early selection of yak could save feeding costs and shorten the generation interval. In the present study, we estimated the accuracy of genomic prediction compared with different classical models for yak early growth traits. The results of cross-validation indicated that the average predictive accuracy ranged from 0.147 to 0.391. The average correlation coefficient between prediction and true phenotype was 0.4. Genomic selection is a promising breeding strategy that has been used in considerable numbers of breeding projects due to its highly accurate results. Yak are rare mammals that are remarkable because of their ability to survive in the extreme and harsh conditions predominantly at the so-called "roof of the world"—the Qinghai–Tibetan Plateau. In the current study, we conducted an exploration of the feasibility of genomic evaluation and compared the predictive accuracy of early growth traits with five different approaches. In total, four growth traits were measured in 354 yaks, including body weight, withers height, body length, and chest girth in two early stages of development (weaning and yearling). Genotyping was implemented using the Illumina BovineHD BeadChip. The predictive accuracy was calculated through five-fold cross-validation in five classical statistical methods including genomic best linear unbiased prediction (GBLUP) and four Bayesian methods. Body weights at 30 months in the same yak population were also measured to evaluate the prediction at 6 months. The results indicated that the predictive accuracy for the early growth traits of yak ranged from 0.147 to 0.391. Similar performance was found for the GBLUP and Bayesian methods for most growth traits. Among the Bayesian methods, Bayes B outperformed Bayes A in the majority of traits. The average correlation coefficient between the prediction at 6 months using different methods and observations at 30 months was 0.4. These results indicate that genomic prediction is feasible for early growth traits in yak. Considering that genomic selection is necessary in yak breeding projects, the present study provides promising reference for future applications. [ABSTRACT FROM AUTHOR]

Domestic yak (Bos grunniens) is the most crucial livestock in the Qinghai-Tibetan Plateau, providing meat and other necessities for local people. The skeletal muscle of adult livestock is composed of muscle fibers, and fiber composition in muscle has influence on meat qualities, such as tenderness, pH, and color. Real-time quantitative polymerase chain reaction (RT-qPCR) is a powerful tool to evaluate the gene expression of muscle fiber, but the normalization of the data depends on the stability of expressed reference genes. Unfortunately, there is no consensus for an ideal reference gene for data normalization in muscle tissues of yak. In this study, we aimed to assess the stability of 14 commonly used candidate reference genes by using five algorithms (GeNorm, NormFinder, BestKeeper, Delat Ct and Refinder). Our results suggested UXT and PRL13A were the most stable reference genes, while the most commonly used reference gene, GAPDH, was most variably expressed across different muscle tissues. We also found that the extensor digitorum lateralis (EDL), trapezius pars thoracica (TPT), and psoas major (PM) muscle had the higher content of type I muscle fibers and the lowest content of type IIB muscle fibers, while gluteobiceps (GB) muscle had the highest content of type IIB muscle fibers. Our study provides the suitable reference genes for accurate analysis of yak muscle fiber composition. [ABSTRACT FROM AUTHOR]

The changes in semen and cryodamage after the cryopreservation process negatively affect sperm function and motility. However, possible proteomic alterations of yak semen during cryopreservation have not yet been achieved. In this study, we compared proteomes of fresh and frozen thawed yak sperm using iTRAQ combined with LC-MS/MS proteome approach. Totally, 2064 proteins were quantitatively identified, including 161 in fresh sperm that showed significant differences compared to frozen thawed sperm. According to the Gene ontology (GO) enrichment analysis, differentially expressed proteins (DEPs) are predominantly associated with spermatogenesis, tricarboxylic acid cycle, ATP synthesis, and differentiation biological process. Furthermore, the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that DEPs were mainly involved in metabolic pathways related to pyruvate metabolism, carbon metabolism, glycolysis/gluconeogenesis, together with the citrate (TCA) cycle. In the analysis of the protein-protein interaction (PPI) network, 15 potential proteins (PDHB, DLAT, PDHA2, PGK1, TP5C1, etc.) that could be related to the sperm quality of the yaks were obtained. Furthermore, 6 DEPs were validated by parallel reaction monitoring (PRM), confirming that the iTRAQ data were reliable. These results indicate that cryopreservation alters the proteome of yak sperm, which is possibly related to cryodamage and fertilization ability. • The impact of cryopreservation on yak sperm was firstly investigated. • The cryopreservation process alters the yak sperm proteome. • The majority of differentially expressed proteins are related to energy metabolism and capacitation of sperm. • The identified differentially expressed proteins may help us understand the cryoinjury mechanism of sperm. [ABSTRACT FROM AUTHOR]