Identification Of Molecular Mechanisms Associated With High-altitude Adaptation In Yak Lung Based On Multi-omics Analysis

Yaks（Bos gruniens）are mainly distributed in the Qinghai-Tibet Plateau and its surrounding plateaus and sub-plateaus in China.After a long time of natural selection,yaks have formed unique phenotypic characteristics and genetic mechanisms of altitude adaptation.Gene expression,as the intermediate medium connecting DNA sequence and physiological phenotype,is important to reveal related molecular pathways and mechanisms.In order to explored the dynamic molecular regulation mechanism of high-altitude adaptation in yak lung,we studied the whole transcriptome and proteome of lung tissues of yak at 3 different altitudes（3400 m,4200 m and 5000 m）by multi-omics analysis,low altitude Zaosheng cattle（1500 m）as control.And the targeting relationship of key genes in high-altitude adaptation was verified by dual luciferase reporter gene assay.The results are as follows:1.A total of 21764 m RNAs,1209 miRNAs,14168 1nc RNAs and 1377 circ RNAs were obtained by whole transcriptome.A total of 4975 significantly differentially expressed m RNAs,75 significantly differentially expressed miRNAs,3326 significantly differentially expressed lnc RNAs and 252 significantly differentially expressed circ RNAs were screened in the comparison between yaks and Zaosheng cattle.A total of 756 significantly differentially expressed m RNAs,83 significantly differentially expressed miRNAs,346significantly differentially expressed lnc RNAs and 64 significantly differentially expressed circ RNAs were screened in the comparison among yaks living at three altitudes.2.Compared with Zaosheng cattle,genes and pathways involved in metabolism and protein stability were significantly enriched in yak lung.Such as key genes ACOT,Abca1,PDIA4,HYOU1,BAX,CAPN1,HSP40,HSP70 and key pathways"long-chain fatty acid metabolic process","‘protein targeting to peroxisome"and"protein processing in the endoplasmic reticulum".In the comparison among yaks living at different altitudes,genes and pathways related to immunity and proliferation were significantly enriched,such as the key gene p15 and the key pathway"FOXO signaling pathway".The gene expression of yak lung showed nonlinear regulation with the altitude.In each altitude gradient,the immune function of yak lung plays an important role.The regulation of metabolism related genes was more active between 3400-4200 m above sea level,and environmental information processing related genes were more important between 4200-5000 m above sea level.3.The target genes of differentially expressed circ RNA was mainly related to metabolism in the comparison between yaks and cattle.The lipid metabolism and reduction of endoplasmic reticulum stress damage related core m RMAs（POR,PRKCSH）and their regulatory relationship（XM＿005892362.2-gma-miR6300＿R2-18L18-PRKCSH and New Gene.14593.1-ata-miR5168-3p-POR）were identified,and revealing that yaks may regulate lipid metabolism through POR regulating PPAR to resist the cold plateau environment.In the comparison among yaks living at different altitudes,the target genes of differentially expressed circ RNA was mainly related to immunity and genetic information processing,maintaining cell homeostasis related core m RMAs（UVSSA,SLTM,PARP4）and their regulatory relationship（New Gene.10538.1-pab-miR396a-PARP4）were identified.4.The expressions of m RNA and protein related to metabolism of yak lung increased with rising altitude,but decreased at the altitude of 4200 meters.The up regulated expression of PPAR related genes may promote the transformation of mesenchymal stem cells into brown adipose tissue（BAT）cells,thus enhancing mitochondrial oxidative phosphorylation to achieve non-shivering thermogenesis and maintain the constant body temperature of yaks in cold plateau environment.It was found that in the ce RNA regulatory network lnc102266115-novel-m0053-3p-RETN with the rising altitude,miRNA（novel-m0053-3p）activity was enhanced after lnc102266115 expression was down regulated,and miRNA combined with target gene RETN to induce the expression of RETN.Inhibited the increase of intracellular Ca²⁺through PLC-ITPR pathway prevents vasoconstriction,thereby preventing the generation of hypoxic pulmonary hypertension.5.Dual luciferase reporter gene assay was used to reveal the targeting relationship between ath-miR8175＿R4-20L20 and ITPR3,novel-m0209-5p and IFIT3 by cloning ITPR3and IFIT3 3’UTR and constructing wild type and mutant recombinant vectors.In summary,high-altitude adaptation related genes of yak lung were nonlinear regulated with the rising altitude,metabolism related genes were more active between 3400-4200 m above sea level,environmental information processing related genes were more important between 4200-5000 m above sea level,and immune process plays an important role in all altitude gradients.The expression of RETN in yaks may be decreased by lnc102266115-novel-m0053-3p-RETN ce RNA regulatory network,preventing the generation of hypoxic pulmonary hypertension through the PLC-ITPR pathway with the rising altitude.This study enriched the reference genome database of yak and provided a reference basis for revealing the adaptive evolutionary mechanism of plateau animals.