Proteomics and Expression of HIF2α/BNIP3L Signaling in Yak Brains at Different Altitudes

The yak, a unique inhabitant of low-oxygen environments, exhibits brain adaptability to hypoxic conditions. However, the impact of hypoxia on yak brain proteomics and the expression of the HIF2α/BNIP3L signaling pathway remains unexplored. This study utilized TMT-based proteomics analysis to identify differentially expressed proteins (DEPs) in the cerebral cortexes of 9-month-old yaks at high (<i>n</i> = 3) and low (<i>n</i> = 3) altitudes. Additionally, qRT-PCR, Western blot, immunohistochemistry, and immunofluorescence were used to analyze HIF2α, BNIP3L, Beclin1, LC3-II, and cleaved caspase-3 expression in various brain regions from both altitude groups. KEGG analysis revealed that the DEPs were mainly concentrated in the synthesis and metabolism, DNA replication, and repair pathways. Specifically, the autophagy in KEGG attracted our attention due to its absence in other animals. HIF2α, BNIP3L, Beclin1, and LC3-II in the autophagy pathway increased significantly. Furthermore, the results of qRT-PCR and Western blot analysis showed that, at the same altitude, the mRNA and protein levels of HIF2α, BNIP3L, LC3-II, and Beclin1 in the cerebral cortexes and hippocampi of yaks were significantly higher than those in the thalami, medulla oblongatae, and cerebella (<i>p</i> < 0.05), while the expression of cleaved caspase-3 was not significantly different among the regions (<i>p</i> > 0.05). Additionally, within the same brain region, the expression levels of HIF2α, BNIP3L, Beclin1, and LC3-II in high-altitude yaks were higher than those in low-altitude yaks. Moreover, there was no difference in the cleaved caspase-3 mRNA and protein expression between the high-altitude and low-altitude yaks. Immunohistochemistry revealed that HIF2α-positive signaling was expressed in the nucleus and cytoplasm of neurons, while BNIP3L, LC3-II, Beclin1, and cleaved caspase-3 were concentrated in the cytoplasm. The immunofluorescence results showed that HIF2α, BNIP3L, LC3-II, Beclin1, cleaved caspase-3, and NeuN were co-located in the neurons of the cerebral cortex, hippocampus, thalamus, medulla oblongata, and cerebellum, respectively. This study offers a complete characterization of the yak cerebral cortex proteome at different altitudes. The higher expression of HIF2α, BNIP3L, Beclin1, and LC3-II in the cerebral cortexes and hippocampi of yaks indicates that these brain regions are more resistant to hypoxia. In addition, the increased HIF2α/BNIP3L signaling in the high-altitude yaks may enhance brain tissue adaptation to hypoxic conditions.