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In multicellular organisms, alternative splicing (AS) is central to the regulation of multiple biological processes. To further elucidate the adaptive strategy of AS in the lungs of Tibetan pigs in response to hypoxia, we identified and analyzed five basic AS types and 59,930 AS events in 18,179 genes. We found that approximately 65.10% of the total expressed genes underwent AS in the lungs of Tibetan pigs at a high altitude (TH). The frequencies of AS events were similar among the different groups (5.06-5.30 events in each gene on average). Skipped exons (SEs) were the predominant type of AS event, followed by mutually exclusive exons (MXEs), alternative 3' splice sites (A3SSs) and alternative 5' splice sites (A5SSs). Retained introns (RIs), the remaining type of AS event, showed lower frequencies. Further comparison analysis of differentially expressed genes (DEGs) and differentially spliced genes (DSGs) identified 2,209 differential splicing events in the above 18,000 expressed genes, including 918 increased and 1,291 decreased splicing events between the TH and Tibetan pigs at a low altitude (TL) groups. We identified 227 hypoxia-related genes involved in lung development that were differentially regulated through AS. Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis clearly identified many DEGs and DSGs at high or low altitude. Seven pathways in the top 20 enriched KEGG terms overlapped for the DEGs and DSGs, including the chemokine signaling pathway, B cell receptor signaling pathway, and cytokine-cytokine receptor interaction, which exert many immunoregulatory and inflammatory actions critical to the lung under hypoxia. Twelve pathways overlapped in hypoxic DEGs and DSGs and included antigen processing, presentation and biosynthesis. ​GO analysis of the DEGs and DSGs among the four groups showed that numerous GO terms were enriched in the biological category, and the proportion of genes with downregulated expression was greater among 227 hypoxic genes than that of all genes. The results suggest that AS plays an essential role in the regulation of gene expression during hypoxia and that numerous genes involved in lung development are differentially regulated through AS., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Due to their long-term colonization of and widespread distribution in plateau environments, yaks can serve as an ideal natural animal model for the adaptive evolution of other plateau species, including humans. Some studies reported that the lung and heart are two key organs that show adaptive transcriptional changes in response to high altitudes, and most of the genes that show differential expression in lung tissue across different altitudes display nonlinear regulation. To explore the molecular mechanisms that are activated in yak lung tissue in response to hypoxia, the mRNAs, lncRNAs and miRNAs of lung tissue from 9 yaks living at three different altitudes (3400 m, 4200 m and 5000 m), with three repetitions per altitude, were sequenced. Two Zaosheng cattle from 1500 m were selected as low-altitude control. A total of 21,764 mRNAs, 14,168 lncRNAs and 1209 miRNAs (305 known and 904 novel miRNAs) were identified. In a comparison of yaks and cattle, 4975 mRNAs, 3326 lncRNAs and 75 miRNAs were differentially expressed. A total of 756 mRNAs, 346 lncRNAs and 83 miRNAs were found to be differentially expressed among yaks living at three different altitudes (fold change ≥ 2 and P-value < 0.05). The differentially expressed genes between yaks and cattle were functionally enriched in long-chain fatty acid metabolic process and protein processing, while the differentially expressed genes among yaks living at three different altitudes were enriched in immune response and the cell cycle. Furthermore, competing endogenous RNA (ceRNA) networks were investigated to illustrate the roles of ceRNAs in this process, the result was also support the GO and KEGG analysis. The present research provides important genomic insights for discovering the mechanisms that are activated in response to hypoxia in yak lung tissue.

The Tibetan pig is an endemic economic animal in the plateau region of China, and has a unique adaptation mechanism to the plateau hypoxic environment. Research into microRNAs (miRNAs) involved in the mechanism underlying hypoxia adaptation of Tibetan pig is very limited. Therefore, we isolated alveolar type II epithelial (ATII) cells from the lungs of the Tibetan pig, cultured them in normoxia/hypoxia (21% O2; 2% O2) for 48 h, and performed high-throughput sequencing analysis. We identified a hypoxic stress-related ssc-miR-141 and predicted its target genes. The target genes of ssc-miR-141 were mainly enriched in mitogen-activated protein kinase (MAPK), autophagy-animal, and Ras signaling pathways. Further, we confirmed that PDCD4 may serve as the target gene of ssc-miR-141. Real-time quantitative polymerase chain reaction (RT-qPCR) analysis was performed to confirm the expression levels of ssc-miR-141 and PDCD4, and a dual-luciferase gene reporter system was used to verify the targeted linkage of ssc-miR-141 to PDCD4. The results showed that the expression level of ssc-miR-141 in the hypoxia group was higher than that in the normoxia group, while the expression level of PDCD4 tended to show the opposite trend and significantly decreased under hypoxia. These findings suggest that ssc-miR-141 is associated with hypoxia adaptation and provide a new insight into the role of miRNAs from ATII cells of Tibetan pig in hypoxia adaptation. [ABSTRACT FROM AUTHOR]

We compared the differences in testicular morphology and vascular anatomy between plateau‐type Tibetan sheep and low‐altitude Small‐tailed Han sheep to explore any adaptations of Tibetan sheep to high altitudes and hypoxia factors. Testicular artery architecture specimens of plateau‐type Tibetan sheep and lowland Small‐tailed Han sheep were used to compare the physical parameters of the testis, testicular artery architecture, arterial diameter, testicular leaflets and the characteristics of epididymal head arterioles. The results showed that the weight, major axis, minor axis, overall length, caput major axis and caput minor axis in Tibetan sheep were larger than those of Small‐tailed Han sheep; however, the caput major axis of epididymal head in Tibetan sheep was smaller than that of Small‐tailed Han sheep. Compared with Small‐tailed Han sheep, the testicular knot‐like arteries of Tibetan sheep had denser spiral distribution and more small arteries, and the diameters of centripetal arteries, knot‐like arteries, centrifugal arteries and epididymal microarterioles were also thicker. The 'spindle' indentation on the surface of the microarteriole of the head of the testicular leaflets and the epididymis in plateau Tibetan sheep was relatively shallow. The results showed that the physical testicular characteristics, testicular artery construction and arteriole surface structure of Tibetan sheep were adapted for the plateau environment. [ABSTRACT FROM AUTHOR]

Tibetan pigs show a widespread distribution in plateau environments and exhibit striking physiological and phenotypic differences from others pigs for adaptation to hypoxic conditions. However, the regulation of mRNAs and metabolites as well as their functions in the alveolar type II epithelial (ATII) cells of Tibetan pigs remain undefined. Herein, we carried out integrated metabolomic and transcriptomic profiling of ATII cells between Tibetan pigs and Landrace pigs across environments with different oxygen levels to delineate their signature pathways. We observed that the differentially accumulated metabolites (DAMs) and differentially expressed genes (DEGs) profiles displayed marked synergy of hypoxia-related signature pathways in either Tibetan pigs or Landrace pigs. A total of 1,470 DEGs shared between normoxic (TN, ATII cells of Tibetan pigs were cultured under 21% O2; LN, ATII cells of Landrace pigs were cultured under 21% O2) and hypoxic (TL, ATII cells of Tibetan pigs were cultured under 2% O2; LL, ATII cells of Landrace pigs were cultured under 2% O2) groups and 240 DAMs were identified. Functional enrichment assessment indicated that the hypoxia-related genes and metabolites were primarily involved in glycolysis and aldosterone synthesis and secretion. We subsequently constructed an interaction network of mRNAs and metabolites related to hypoxia, such as guanosine-3′, 5′-cyclic monophosphate, Gly-Tyr, and phenylacetylglycine. These results indicated that mitogen-activated protein kinase (MAPK) signaling, aldosterone synthesis and secretion, and differences in the regulation of MCM and adenosine may play vital roles in the better adaptation of Tibetan pigs to hypoxic environments relative to Landrace pigs. This work provides a new perspective and enhances our understanding of mRNAs and metabolites that are activated in response to hypoxia in the ATII cells of Tibetan pigs. [ABSTRACT FROM AUTHOR]

To adapt to a low-oxygen environment, Tibetan pigs have developed a series of unique characteristics and can transport oxygen more effectively; however, the regulation of the associated processes in high-altitude animals remains elusive. We performed mRNA-seq and miRNA-seq, and we constructed coexpression regulatory networks of the lung tissues of Tibetan and Landrace pigs. HBB, AGT, COL1A 2, and EPHX 1 were identified as major regulators of hypoxia-induced genes that regulate blood pressure and circulation, and they were enriched in pathways related to signal transduction and angiogenesis, such as HIF -1, PI 3 K - Akt, mTOR , and AMPK. HBB may promote the combination of hemoglobin and oxygen as well as angiogenesis for high-altitude adaptation in Tibetan pigs. The expression of MMP 2 showed a similar tendency of alveolar septum thickness among the four groups. These results indicated that MMP 2 activity may lead to widening of the alveolar wall and septum, alveolar structure damage, and collapse of alveolar space with remarkable fibrosis. These findings provide a perspective on hypoxia-adaptive genes in the lungs in addition to insights into potential candidate genes in Tibetan pigs for further research in the field of high-altitude adaptation. [ABSTRACT FROM AUTHOR]

Yak is one of the largest native mammalian species at the Himalayas, the highest plateau area in the world with an average elevation of >4,000 m above the sea level. Yak is well adapted to high altitude environment with a set of physiological features for a more efficient blood flow for oxygen delivery under hypobaric hypoxia. Yet, the genetic mechanism underlying its adaptation remains elusive. We conducted a cross-tissue, cross-altitude, and cross-species study to characterize the transcriptomic landscape of domestic yaks. The generated multi-tissue transcriptomic data greatly improved the current yak genome annotation by identifying tens of thousands novel transcripts. We found that among the eight tested tissues (lung, heart, kidney, liver, spleen, muscle, testis, and brain), lung and heart are two key organs showing adaptive transcriptional changes and >90% of the cross-altitude differentially expressed genes in lung display a nonlinear regulation. Pathways related to cell survival and proliferation are enriched, including PI3K-Akt, HIF-1, focal adhesion, and ECM–receptor interaction. These findings, in combination with the comprehensive transcriptome data set, are valuable to understanding the genetic mechanism of hypoxic adaptation in yak. [ABSTRACT FROM AUTHOR]