Identification And Analyses Of Differentially Expressed MicroRNAs In Liver And Spleen Tissues Between Zebu And Taurine Cattle

As one of the most important domestic animals,domestic cattle provide important food and productive resources for human survival and development,and keep a close relation with the civilization of human beings.Therefore,domestic cattle are always the focus of domestic animal research.Domestic cattle are divided into two subspecies or types-Bos taurus taurus?taurine cattle?and Bos taurus indicus?zebu cattle?.There exist many differences between them,including not only in their physical appearance but also in many physiological processes.Thus it is very important to study and reveal the molecular mechanism of differences between them.microRNAs?miRNAs?are a class of endogenous noncoding small RNAs molecules that have about 22 nt in length.Many studies have shown that miRNAs are common in animals,plants and viruses.miRNAs play a key regulatory role in almost all biological processes of living organisms.In addition,the research on miRNAs of domestic cattle lags behind that on humans and mice.At present,there are no reports on the miRNAs comparative studies between taurine cattle and zebu cattle.Therefore,taurine cattle were selected as reference for the first time,in the present study two important internal organs — liver and spleen were collected from healthy taurine cattle and zebu cattle for miRNAs highthroughput sequencing?five biological replicates for each tissue for each species,a total of 20 samples?.This study aims to search for novel miRNAs and specific miRNAs expressed in domestic cattle and to identify and analyze the differentially expressed miRNAs in liver and spleen tissues between taurine cattle and zebu cattle.The results showed as follows:The number of known miRNAs ranging from 376 to 450 were identified in 20 cattle samples.The number of novel miRNAs ranging from 554 to 832 predicted in 20 cattle samples.The expression level of bta-miR-122 and novel miRNA AC₀00181.1₂7195* was the highest in the liver tissue.However,the expression level of bta-miR-143 and novel miRNA AC₀00168.1₁4386* was the highest in the spleen tissue.Meanwhile,the BLAST analysis indicated that two novel miRNAs AC₀00181.1₂7195* and AC₀00168.1₁4386* share higher homology with human miRNAs hsa-miR-3591-3p and hsa-miR-126-3p,respectively.There were nine known miRNAs recognizable as specifically expressed in domestic cattle,such as bta-miR-2285 c,bta-miR-2336,bta-miR-2440,bta-miR-3431,btamiR-2478,bta-miR-2284 w,bta-miR-6119-3p,bta-miR-2285 t and btamiR-2898.In addition,bta-miR-122,bta-miR-192,bta-miR-194,bta-miR-200 and bta-miR-885 exhibited a tissue specific expression in liver tissue of domestic cattle.Among these miRNAs,bta-miR-192 and bta-miR-194 were verified by RT-qPCR in this experiment.This study showed that the number of differentially expressed known miRNAs and new predicted novel miRNAs between spleen tissues of taurine cattle and zebu cattle were 17 and 13,respectively.However,the number of differentially expressed known miRNAs and novel miRNAs between the liver tissues of taurine cattle and zebu cattle were 13 and 13,respectively.Furthermore,according to the sequencing results,seven miRNAs?bta-miR-135 a,bta-miR-150,bta-miR-155,bta-miR-192,btamiR-194,bta-miR-451 and AC₀00169.1₁4517?were chosen for RTqPCR verification analyses,and the final results showed that the RT-qPCR results were basically consistent with the sequencing results.Homologous comparisons revealed that the majority of differentially expressed novel miRNAs in the liver tissues and the spleen tissues between taurine cattle and zebu cattle had high homology with the members of the bta-miR-2284 and bta-miR-2285 families.The functional enrichment analysis was performed on the target genes of the differentially expressed known miRNAs in the spleen tissues and the liver tissues between taurine cattle and zebu cattle.The results demonstrated that most of these target genes were enriched to the pathways that were associated with domestic cattle metabolism and immunity,such as the insulin pathway,the mTOR signaling pathway,the IFN-? pathway,the IL3 mediated signaling events,the IL5 mediated signaling events.In this study,comparative study was performed on miRNAs of taurine cattle and zebu cattle for the first time,and a large number of novel miRNAs and differentially expressed miRNAs were identified between them.These results not only enriched the information data of domestic cattle miRNAs,but also provided important theoretical basis for further research on the molecular mechanism of the different phenotypes between the taurine cattle and zebu cattle.