Noncoding RNAs Expression And Regulations In The Testes Of Immature And Sexually Mature Wandong Cattle(Bos Taurus)

Wandong cattle are local breed and distributed in the Jianghuai watershed area in eastern Anhui province.The breed has high adoptability to extreme weather conditions and survives on raw feed as well as resistant to tick-borne diseases.However,the introduction of germplasm through artificial insemination;and the proper use of genetic prediction in the reproductive performance of Wandong bull,is not well established.Improving reproductive efficiency of local bull via genetical selection is crucial,because individual bull can breed multiple numbers of cows through artificial insemination(AI).Therefore,basic biological understanding and novel molecular techniques are required to propagate the superior genetic bulls.Based on this,3-month-old calves(immature)and 3-year-old Wandong bulls(sexually mature)were selected.Testicular tissues were collected for transcriptome sequencing analysis to explore the regulatory role of transcription factors in testicular development of Wandong cattle,and the following conclusions were drawn:1.A total of 21250 mRNA and 20533 lncRNAs were found in calf and bull testicles,among which 7908 lncRNA(P<0.05)and 5122 mRNA(P<0.05)differentially expressed between two developmental stages of bulls.In addition,GO analysis and KEGG analysis showed that these genes were enriched in lysine degradation,cell cycle,propionic acid metabolism,adhesion and cell adhesion pathways.The mRNA source genes(CCDC83,DMRTC2,HSPA2,IQCG,PACRG,SPO11,EHHADH,SPP1,NSD2,ACTN4)and lncRNA(COX7A2,COX6B2,TRIM37,PRM2,INHBA,ERBB4,SDHA,ATP6VOA2,FGF9,TCF21)are closely related to sexual maturation and spermatogenesis in bulls.2.A total of 17013 circRNA were found in calf and bull testicles,among which 681 circRNA(P<0.05)were differentially expressed.Among the differentially expressed circRNA,579 of them are up-regulated and another 103 are down-regulated.GO and KEGG analysis revealed the involvement of multiple target genes in biological processes,cell components and molecular functions.In addition,signaling pathways of the source genes of circRNA were analyzed,which were enriched in cell cycle,adhesion links,tight junction,and cell degradation pathways.The source genes such as ATM,CCNA1,GSK3B,KMT2E,NSD2,SUCLG2,QKI,HOMER1 and SNAP91 are closely related to sexual maturation and spermatogenesis in bulls.The binding interactions between differential circRNA and miRNAs are constructed using bioinformatics methods.3.A total of 754 miRNA were found in calf and bull testicles,among which 98 were newly identified,while,152 miRNA were differentially expressed.Among the differentially expressed miRNA,64 of them were significantly up-regulated and another 87 were significantly down-regulated(P<0.05).The target genes of miRNAs,such as JAG2,BCL6,CFAP157,PHC2,TYRO3,SEPTIN6 and BSP3 were analyzed for their signaling pathways.These candidate genes are involved in TNF signaling pathway,T cell receptor,PI3KAkt and other signaling pathways,affecting the functions of testicular development and spermatogenesis.The differentially expressed miRNAs such as MIR425,MIR98,MIR34C,MIR18A,MIR18A,MIR136,MIR15A,MIR1388 and MIR210 are closely related to sexual maturation and spermatogenesis in bulls.In this study,mRNA,LncRNA,circRNA and miRNA in testis of Wandong cattle were comprehensively analyzed,providing a reference for further understanding of the molecular mechanism of the interaction between mRNA-lncRNA-circRNA-miRNA,which is related to testis development at immature and mature stages.In the next step of the study,the marker genes related to semen quality in bulls,or candidate gene functional studies required to explore the genes regulating testicular development and spermatogenesis in local bulls and their regulatory mechanisms.