Screening Polytocous Candidate Genes In Small Tail Han Sheep Based On Hypothalamic Multi-omics Analysis

The litter size in sheep is a quantitative trait controlled by minor-polygene,and the lower litter size is an important factor significantly limiting sheep industry development.Recent years,as the researches going on,many genetic markers associated with litter size have been discovered,however the mechanism affecting litter size remains to be explored.Our previous studies suggested that there are many differences between polytocous and monotocous ewes without FecB mutation in hormone secretions such as progesterone,estrogen,and follicle-stimulating hormone,which indicated that hormone activities may be key factor influencing litter size.Considering the key roles of hypothalamus on initiating hormones secretion,this study divides all the selected sheep into polytocous sheep in the follicular phase(PF),polytocous sheep in the luteal phase(PL),monotocous sheep in the follicular phase(MF)and monotocous sheep in the luteal phase(ML),and aims to analyze the molecular mechanism affecting sheep reproduction in Small Tail Han sheep without FecB mutation via transcriptomics and proteomics.We expect to determine the candidate genes except for FecB.The main content and conclusions of this study were listed as followed:In this study,hypothalamic RNA-seq was used for the first time to screen candidate genes in sheep without FecB mutation.The transcriptomics data suggested that differentially expressed genes(DEGs)like PRL,POMC,GNRH1,GH and TTR were highly enriched in pathways such as Jak-STAT signaling pathway,and participated in many processes like hypothalamic gonadal hormone secretion.By analyzing interactive networks of lncRNA-mRNA,we found that FOXG1 which affects GnRH secretion was regulated by lncRNA MSTRG.95128,and both of which were up-regulated in PF vs.MF,while were down-regulated in PF vs.MF.The results suggested that MSTRG.95128 may influence GnRH secretion by modulating the functions of FOXG1.By analyzing interactive networks of miRNA-mRNA,we predicted that several key DEGs functioned in GnRH activities were regulated by DE miRNAs,such as TRH was co-regulated by oar-miR-379-5p,oar-miR-30 b,oar-miR-152,oar-miR-495-3p,oar-miR-143,oar-miR-106 b,oar-miR-218 a and oar-miR-148 a,and TTR was regulated by oar-miR-432.Therefore,these DE miRNAs may further affect hypothalamic hormone activities and even litter size via modulating functions of target genes like TRH and TTR.In order to further pinpoint the candidate genes affecting litter size in hypothalamus of sheep,hypothalamic TMT-based proteomics analysis was carried out in this study.The results suggested that several differentially abundant proteins(DAPs)like GH,HDAC4,IGF1 R,HSD11B1 and TTR functioned in hormone activity,PI3K-AKT and JAK-STAT signaling pathways,and these DAPs may further modulate GnRH activities directly or indirectly by regulating activities of insulin,leptin,and other proteins.Integrating transcriptomes and proteomics data,we speculated that LGALS3,ASPA and TTR could be the candidate genes affecting litter size in sheep without FecB mutation.In this study,for the first time,we explored the genes and proteins related to polytocous trait in Small Tail Han sheep without FecB mutation via hypothalamic transcriptomics and proteomics.All in all,this study may provide basis for understanding the prolific molecular mechanism of sheep and breeding new sheep breeds with higher fecundity.