Single-cell long- and short-read transcriptomics sheds light on high-altitude adaptation in the porcine cerebral cortex.

The cerebral cortex is highly sensitive to oxygen availability and thus serves as an ideal model for studying neural adaptation to hypoxia. Here, we integrated long-read (Oxford Nanopore) and short-read (Illumina) single-cell RNA sequencing to systematically characterize the cellular composition and transcriptional landscape of the cerebral cortex in high-altitude Diqing Tibetan pigs (DT, ~3200 m) and low-altitude Diannan small-ear pigs (DSE, ~500 m). Single-cell analysis identified nine major cortical cell types and revealed pronounced transcriptional divergence between the breeds. In DT, multiple genes associated with high-altitude hypoxia adaptation and energy homeostasis, including NKAIN2, CTNNA3, ZFP36, and HSP40 family members, were significantly upregulated. Functional enrichment highlighted mitochondrial oxidative phosphorylation and energy metabolism pathways, suggesting metabolic reprogramming under chronic hypoxia. Key transcription factors, including SOX17, NKX6-2, ETV4, and SPI1, displayed cell-type-specific activity in Endo, Oligo, and Micro. Cell-cell communication analysis showed DSE with broader intercellular connectivity, whereas DT exhibited more focused and stronger signaling interactions. Integration of long-read sequencing further identified numerous novel, cell-type-specific transcripts, expanding porcine cortical transcriptome annotation. Collectively, these findings provide a comprehensive framework for understanding cellular and molecular adaptations of the cerebral cortex to prolonged high-altitude hypoxia.