Small RNA sequencing reveals miRNAs important for hypoxic adaptation in Tibetan chicken

Tibetan chicken, an indigenous chicken breed living at the Tibetan Plateau, exhibits specific adaptations to high-altitude environment. However, the molecular mechanism of hypoxic adaptation is still unclear. This study aimed to investigate differentially expressed miRNAs between hypoxia-adapted and lowland chickens through high-throughput RNA sequencing. Quantitative reverse transcription polymerase chain reaction was used to verify the differentially expressed miRNAs and their target genes in chicken embryonic heart tissues and fibroblasts, and luciferase reporter assays was performed to confirm the relationship between miRNAs and target genes. We identified 37 differentially expressed miRNAs, including 20 up- and 17 down-regulated miRNAs in Tibetan chicken embryonic heart tissues compared to that in the lowland chicken. Differentially expressed miRNAs were mainly involved in the biological process of cell cycle arrest, toll-like receptor signaling pathway, and I-kappa B kinase/NF-kappa B signaling. We found that gga-miR-34c-5p was significantly upregulated, while EHHADH, a target gene of gga-miR-34c-5p, was downregulated in Tibetan chicken tissues and hypoxic fibroblasts. Moreover, gga-miR-34c-5p dramatically decreased the luciferase activity of the wild EHHADH, whereas showed no effect on the mutational EHHADH. This study generated the miRNA expression profiles in Tibetan chicken and suggested miR-34c-5p might act as a novel miRNA associated with hypoxic adaptation, facilitating to understand the molecular mechanisms that underlie long-term exposure to hypoxia.