Sequence Characteristics Of PiRNAs And Their Expression Differences In Gonadal Tissues Between Monotocous And Polytocous Sheep

piRNA is a kind of non-coding RNA range from 23 to 34 nt.The known functions of piRNAs are mainly involved in inhibition of transposon activity and regulation of gene expression.piRNAs play an important role in many physiological processes such as germ cell differentiation,spermatogenesis,oogenesis,embryonic development,sex determination and so on.But it's role in sheep reproduction is yet unclear.In order to explore the characteristics of piRNAs sequence on sheep gonadal tissue and it's function in sheep reproduction,the small RNAs in testis and ovary of monotocous and polytocous sheep were subjected to Illumina sequencing.After the piRNAs candidate were screened out,the characteristics of piRNAs sequences in sheep gonadal tissue were analyzed.Furthermore,the expression difference of piRNAs in the testis between monotocous and polytocous sheep breeds and in the ovary between monotocous and polytocous Small Ttail Han ewes were analyzed to speculate the role of piRNAs in sheep reproduction.The results are summarized as follows:1.Characteristics of piRNAs sequence in sheep gonadal tissue.(1)The piRNAs reads in testis of monotocous and polytocous sheep showed a unimodal distribution between 26 and 31 nt with a peak at 29 nt,and their tags showed a bimodal distribution between 24 and 31 nt with peaks at 26 and 29 nt.The piRNAs reads in ovary of monotocous and polytocous sheep both peaked at 24 nt and 27~29nt whether at follicular or luteal phase,and their tags showed a decreasing distribution from 24 nt to 33 nt.(2)Statistic results of genomic contexts showed that the piRNAs in ovine testis are mainly aligned to CDS,repeat region,intron and unannotated region,while just a few piRNAs are aligned to 5?UTR,3?UTR and lncRNA.The piRNAs in ovine ovary are mainly mapped to intron,CDS and repeat region,except for the monotocous group in follicular phase.There are a large number of piRNAs were aligned to the unannotated region in other three groups,however just few piRNAs were mapped to the 5?UTR,3?UTR and lncRNA regions in all four groups.(3)There are obvious ping pong signature in the testis of sheep but not in the ovary.(4)The piRNAs in the testis and ovary of sheep were mainly aligned to LINE,SINE and LTR,and some piRNAs were aligned to DNA transposons.The piRNAs derived from different TE in the testis of sheep showed obvious ping pong signature while not in ovary.(5)The piRNAs in ovine testis are mainly distributed on chromosomes 3,7,15,17,18,and 20,and the piRNAs in ovary of sheep are mainly distributed on chromosome 6.(6)The number of piRNAs distributed on the positive strand of DNA is almost equal to that on the negative strand in testis and ovary of sheep.2.A total of 5461 differentially expressed piRNAs were obtained in the testis between monotocous and polytocous sheep breed.Their target genes were significantly enriched in 73 pathways,including the pathway related to the renewal and differentiation of sperm stem cells(PIK3-Akt signaling pathway and Ras signaling pathway),pathway related to blood testis barrier stability(Tight junction pathway),pathway that can affects the energy metabolism and proliferation of Sertoli cell(AMPK signaling pathway).Together,this suggests that the differentially expressed piRNA in testis between monotonous and polytocous sheep may affect ram reproduction through the above pathways.3.In the follicular phase and luteal phase,972 and 1312 differentially expressed piRNAs were obtained in the ovary of between monotocous and polytocous Small Tailed Han ewes respectively.Their target genes were significantly enriched in 83 and 76 pathways,including the pathway that related to the activation and development of primordial follicles such as PIK3-Akt signaling pathway and TGF? signaling pathway,the pathway that related to Oocyte meiosis such as Oocyte meiosis pathway,Purine metabolism pathway and so on,pathway related to the selection of dominant follicles and ovulation such as the Estrogen signaling pathway.Together,this suggests that differentially expressed piRNAs in the ovary may affect the reproduction of ewes through the above pathways.This study revealed the sequence characteristics of piRNAs in sheep gonadal tissues and speculate the pathways through which piRNAs may participate in the regulation of sperm quantity and quality in the testis of sheep and the regulation of oogenesis and ovulation in the ovary of sheep.This study provides a new perspective for further analysis about the epigenetic mechanism of sheep high fertility.