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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., (Copyright © 2023 Elsevier Inc. All rights reserved.)

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., Competing Interests: Declaration of competing interest The authors declare that there is no conflict of interest regarding the publication of this paper., (Copyright © 2023. Published by Elsevier B.V.)

To achieve fertilization, mammalian spermatozoa must undergo capacitation and the acrosome reaction (AR) within the female reproductive tract. However, the effects of cryopreservation on sperm maturation and fertilizing potential have yet to be established. To gain insight into changes in protein levels within sperm cells prepared for use in the context of fertilization, a comprehensive quantitative proteomic profiling approach was used to analyze frozen-thawed Ashidan yak spermatozoa under three sequential conditions: density gradient centrifugation-based purification, incubation in a capacitation medium, and treatment with the calcium ionophore A23187 to facilitate AR induction. In total, 3280 proteins were detected in these yak sperm samples, of which 3074 were quantified, with 68 and 32 being significantly altered following sperm capacitation and AR induction. Differentially abundant capacitation-related proteins were enriched in the metabolism and PPAR signaling pathways, while differentially abundant AR-related proteins were enriched in the AMPK signaling pathway. These data confirmed a role for superoxide dismutase 1 (SOD1) as a regulator of sperm capacitation while also offering indirect evidence that heat shock protein 90 alpha (HSP90AA1) regulates the AR. Together, these findings offer a means whereby sperm fertility-related marker proteins can be effectively identified. Data are available via Proteome Xchange with identifier PXD035038.

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., (© 2022. The Author(s).)

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.

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., (© 2022. The Author(s).)

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]

Yaks (Bos grunniens) are a critical livestock breed in the plateau region, and changing the feeding system of yaks can significantly improve their growth performance. The effects of different feeding regimes on the growth performance and meat quality of yaks were comprehensively compared here. The transcriptome and proteome of the Longissimus dorsi muscle were determined using RNA-seq and Tandem Mass Tag (TMT) techniques. Indoor feeding significantly improved the growth performance (such as the average daily gain and carcass weight) and meat quality characteristics compared with traditional grazing feeding. In the grazing (Group G) vs. in-house fed group (Group HF) comparison, 40 differentially expressed genes/differentially abundant proteins exhibited the same mRNA and protein expression trends. These genes were associated with collagen binding, the lipoxygenase pathway, and the arachidonic acid metabolic process. Parallel reaction monitoring verified whether the TMT results were reliable. Moreover, some pathways, such as the AMPK signaling pathway, FoxO signaling pathway, PPAR signaling pathway, and fatty acid metabolism, were significantly enriched. These results expand our knowledge about meat quality in yaks and provide practical information and more evidence for further insight into the biological mechanisms underlying meat quality traits. [ABSTRACT FROM AUTHOR]