Cattle-yak is an excellent hybrid of cattle and yak; they are characterized by better meat quality and stronger adaptability of harsh environments than their parents. However, male sterility of cattle-yak lay restraints on the transmission of heterosis. In this study, next generation sequence technology was performed to profile the testicular tissues transcriptome (lncRNA and mRNA) of cattle, yak, and cattle-yak. We analyzed the features and functions of significant differentially expressed genes among the three breeds. There are 9 DE lncRNAs and 46 DE mRNAs with comparisons of cattle, yak, and cattle-yak. Among them, the upregulated targeting genes, such as IGF1 and VGLL3 of cattle-yak lncRNA, may be related to the derangement of spermatocyte maturation and cell proliferation. Similarly, we found that the LDOC1 gene, which is related to the process of cellular apoptosis, is overexpressed in cattle-yak. GO enrichment analysis demonstrated that the cattle-yak is lacking the regulation of fertilization (GO: 0009566), spermatogenesis process (GO: 0007283), male gamete generation process (GO: 0048232), sexual reproduction (GO: 0019953), and multi-organism reproductive process (GO: 0044703), such processes may play important and positive roles in spermatogenesis and fertilization. Furthermore, the KEGG enrichment analysis showed that the upregulated DEGs of cattle-yak most enriched in Apoptosis (ko04210) and Hippo signaling pathway (ko04390), may lead to excessively dead of cell and inhibit cell growth, resulting in obstruction of meiosis and spermatogenesis processes. This study will enable us to deeper understand the mechanism of male cattle-yak infertility.

Identification of Novel lncRNA and Differentially Expressed Genes (DEGs) of Testicular Tissues among Cattle, Yak, and Cattle-Yak Associated with Male Infertility

The first filial generation of the cattleyaks demonstrates hybrid vigor; however, the male cattleyaks are infertile and restrict productivity and breeding. The discovery of genes in a segment-specific approach offers valuable information and understanding concerning fertility status, yet the biology of cattleyak epididymis is still progressing. Comparative transcriptome analysis was performed on segment pairs of cattleyak epididymis. The caput versus corpus epididymis provided the highest (57.8%) differentially expressed genes (DEGs), corpus versus cauda (25.1%) followed, whereas caput versus cauda pair (17.1%) had the least DEGs. The expression levels of genes coding EPHB6, TLR1, MUC20, MT3, INHBB, TRPV5, EI24, PAOX, KIF12, DEPDC5, and KRT25, which might have the potentials to regulate the homeostasis, innate immunity, differentiation, motility, transport, and sperm maturation-related function in epididymal cells, were downregulated in the distal segment of epididymis. Top enriched KEGG pathways included mTOR, axon guidance, and taste transduction signaling pathways. EIF4B, EPHB6, and TAS2R42 were enriched in the pathways, respectively. Identifying key, new, and unexplored DEGs among the epididymal segments and further analyzing them could boost cattleyak fertility by maximizing sperm quality from genetically better sires and also facilitate better understanding of the epididymal biology.

RNA-Seq reveals the functional specificity of epididymal caput, corpus, and cauda genes of cattleyak.

N6-methyladenosine (m6A) methylation is a type of methylation modification discovered on RNA molecules, mainly on mRNAs, as well as on other RNAs. Similar to DNA methylation, m6A methylation regulates the post-transcriptional expression level of genes without altering their base sequences. It modulates gene expression mainly by affecting the binding of mRNAs to reader proteins, thereby regulating variable splicing, translation efficiency, and stability of mRNAs. Early in the research, the study of m6A-related biological functions was greatly hindered due to the lack of effective detection methods. As second-generation sequencing and bioinformatics develop, several methods have been available to detect and predict m6A methylation sites in recent years. Moreover, m6A methylation is also closely related to the development of lipid metabolism, as shown in current studies. Combined with recent research, this paper reviews the concept, detection, and prediction means of m6A methylation, especially the relationship between m6A and lipid metabolism, providing a new clue to enrich the molecular mechanism of lipid metabolism.

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Progress of m6A Methylation in Lipid Metabolism in Humans and Animals

Abstract β-defensins are adsorbable on the sperm surface in the male reproductive tract (MRT) and enhance sperm functional characteristics. The beta-defensin 129 ( DEFB129 ) antimicrobial peptide is involved in sperm maturation, motility, and fertilization. However, its role in bovine fertility has not been well investigated. This study examines the relationship between the bovine BBD129 gene and Bos indicus x Bos taurus bull fertility. The complete coding sequence of BBD129 mRNA was identified by RNA Ligase Mediated-Rapid Amplification of cDNA End (RLM-RACE) and Sanger sequencing methodologies. It consisted of 582 nucleotides (nts) including 5' untranslated region (UTR) (46nts) and 3'UTR (23nts). It conserves all beta-defensin-like features. The expression level of BBD129 was checked by RT-qPCR and maximal expression was detected in the corpus—epididymis region compared to other parts of MRT. Polymorphism in BBD129 was also confirmed by Sanger sequencing of 254 clones from 5 high fertile (HF) and 6 low fertile (LF) bulls at two positions, 169 T &gt; G and 329A &gt; G, which change the S57A and N110S in the protein sequence respectively. These two mutations give rise to four types of BBD129 haplotypes. The non-mutated TA- BBD129 (169 T/329A) haplotype was substantially more prevalent among high-fertile bulls (P &lt; 0.005), while the double-site mutated GG- BBD129 (169 T &gt; G/329A &gt; G) haplotype was significantly more prevalent among low-fertile bulls ( P &lt; 0.005). The in silico analysis confirmed that the polymorphism in BBD129 results in changes in mRNA secondary structure, protein conformations, protein stability, extracellular-surface availability, post-translational modifications (O-glycosylation and phosphorylation), and affects antibacterial and immunomodulatory capabilities. In conclusion, the mRNA expression of BBD129 in the MRT indicates its region-specific dynamics in sperm maturation. BBD129 polymorphisms were identified as the deciding elements accountable for the changed proteins with impaired functionality, contributing to cross-bred bulls' poor fertility. 

/pdf/analysis-of-amplification-and-association-polymorphisms-in-2698rwn5.pdf

Xin Cai

Papers

Analysis of long non-coding RNAs in epididymis of cattleyak associated with male infertility.