Molecular Mechanism Of Cold Resistance Revealed By Adipose Tissue Transcriptome In Mongolian Sheep

Mongolian sheep are mainly distributed in the Mongolian Plateau with harsh environmental conditions.The sheep have become local varieties with excellent quality and distinctive characteristics via long-term natural selection and artificial breeding under harsh environmental conditions.The remarkable feature of Mongolian sheep is that has strong adaptability to severe cold habitat.However,the genetic mechanism of cold resistance is not completely clear.Adipose tissue throughout animal body plays an important role in the process of cold resistant regulation.Therefore,the perirenal,subcutaneous and tail adipose tissues of Mongolian sheep in winter and summer were selected for high-throughput sequencing.Through bioinformatic analysis of transcriptome data from different adipose tissues and validation of molecular and cellular levels,it is expected to reveal the molecular regulatory mechanisms of Mongolian sheep adaptation to cold habitat.The results are as follows:1.The numbers of m RNAs,lnc RNAs,circ RNAs and mi RNAs differentially expressed in perirenal,subcutaneous and tail fat in winter and summer were 5482,4840,6450;6248,5556,6609;3346,2909,3324;79,68 and 46,respectively.The common differentially expressed m RNAs,lnc RNAs,circ RNAs and mi RNAs in different adipose tissues were 1787,1763,704 and 13,respectively.2.GO Category show that the differentially expressed m RNAs and nc RNAs in adipose tissue were significantly enriched in biological processes such as intracellular and extracellular transport and metabolism of substances(including ions,electrons,etc.).KEGG analysis show that 1720,2108 and 2027 m RNAs differentially expressed in perirenal,subcutaneous and tail fats were extremely enriched to 34,48 and 31 signaling pathways related to fatty acid metabolism,oxidative phosphorylation,vitamin A metabolism and so on.Differentially expressed nc RNAs are also mostly focused on material metabolism,oxidative respiration and related pathways.However,there are differences in transcriptional profiles and enrichment pathways between different position's adipose tissues.3.The q RT-PCR statistical analysis of adipose tissue from winter and summer showed that there were differences in gene expression patterns in distinct adipose tissues at the same period.Our results show that there were the various metabolism regulation mechanism in different adipose tissues for better adapt to the cold environment.4.The results of q RT-PCR confirmed that the expression of thermogenic marker gene UCP1 was obviously up-regulated in adipose tissue of Mongolian sheep from winter.By observing the adipose tissue sections,we confirmed that there were beige adipocytes,which was an important'heat source'to resists the cold habitat in adult Mongolian sheep.5.The expression of LIPE and LIPE-OT1 in adipose tissue showed significant correlation(R~2=0.973).Dual-luciferase reporter gene assay identified that LIPE and LIPE-OT1 were the genes targeted for novel?38.LIPE-OT1 promoted the expression and translation of LIPE in the form of competitive endogenous RNA which adsorpted or phagocytosed the novel?38.Upregulated expression of the LIPE mediated the adipose lipolysis and provided more energy for thermogenesis of Mongolian sheep toresistant cold circumstances.6.The candidate genes RDH16 and ANGPTL7 differentially expressed in perirenal and subcutaneous adipose tissue.The q RT-PCR confirmed that RDH16 and ANGPTL7were significantly up-regulated in perirenal and subcutaneous adipose tissues of Mongolian sheep from winter,respectively.Over-expression of RDH16 and ANGPTL7 in fetal perirenal and subcutaneous preadipocytes from Mongolian sheep showed that up-regulation of RDH16 and ANGPTL7 could inhibit the formation of white adipocytes and promote the production of brown/beige adipocytes.Meanwhile the upregulation of RDH16 led to the expression and transcription of FABP4 increased obviously.7.The regulatory pathways in adipose tissue of Mongolian sheep adapt to cold habitats were basicly established.The pathway indicated that cold-induction changes the gene transcription pattern in adipose tissue,and the up-regulated LIPE-OT1 which targeted to bind novel?38 decrease its inhibitory effect on LIPE expression,and promote lipid droplet decomposition to release fatty acids and retinol.The up-regulated expression of RDH16 elevated the retinol hydrolysis and made more retinoic acid.The results block the formation of white adipocytes and increase the formation of thermogenic brown/beige adipocytes.At the same time,the up-regulated expression of RDH16 accelerated the production of FABP4,promoted the transportion of intracellular fatty acids,retinol and its derivatives.Finally,it regulates lipornetabolism processes,and provides more energy sources for the cold resistance of Mongolian sheep.This study provides a scientific basis for molecular breeding of new cold-resistant livestock varieties,and also sheds light on research of human obesity and other metabolic diseases.