细胞因子-1α启动因子(HIF-1α)是各种疾病中的一个关键治疗靶点，包括癌症和纤维化。我们之前证明，转染包括polyI：C和哺乳动物正交再病毒的双链RNA(dsRNA)显著降低了培养细胞中HIF-1α蛋白水平；然而，如何dsRNA诱导HIF-1α蛋白水平下调仍未明确。在本研究中，我们研究了dsRNA介导的HIF-1α蛋白水平下调的机制。我们发现，在dsRNA转染细胞中，敲低和敲除蛋白激酶R(PKR)显著恢复了HIF-1α蛋白水平，表明PKR参与了dsRNA介导的HIF-1α下调。蛋白酶体抑制剂显著恢复了dsRNA转染细胞中的HIF-1α蛋白水平。dsRNA转染后，HIF-1α的泛素化水平增加。这些发现表明，在dsRNA转染后，通过泛素-蛋白酶体途径促进了HIF-1α的降解，并且依赖于PKR。除HIF-1α外，转染dsRNA后还下调了几种蛋白质的表达，包括CDK4和HER2。这些数据为揭示dsRNA介导的细胞毒性机制提供了重要线索，并为dsRNA的治疗应用提供了重要参考。©2023. 作者。

Hypoxia inducible factor-1α (HIF-1α) is a crucial therapeutic target in various diseases, including cancer and fibrosis. We previously demonstrated that transfection with double-stranded RNA (dsRNA), including polyI:C and the dsRNA genome of mammalian orthoreovirus, resulted in significant reduction in HIF-1α protein levels in cultured cells; however, it remained to be elucidated how dsRNA induced down-regulation of HIF-1α protein levels. In this study, we examined the mechanism of dsRNA-mediated down-regulation of HIF-1α protein levels. We found that among the various cellular factors involved in dsRNA-mediated innate immunity, knockdown and knockout of protein kinase R (PKR) significantly restored HIF-1α protein levels in dsRNA-transfected cells, indicating that PKR was involved in dsRNA-mediated down-regulation of HIF-1α. Proteasome inhibitors significantly restored the HIF-1α protein levels in dsRNA-transfected cells. Ubiquitination levels of HIF-1α were increased by transfection with dsRNA. These findings indicated that degradation of HIF-1α in a ubiquitin-proteasome pathway was promoted in a PKR-dependent manner following dsRNA transfection. Expression of not only HIF-1α but also several proteins, including CDK4 and HER2, was down-regulated following dsRNA transfection. These data provide important clues for elucidation of the mechanism of dsRNA-mediated cellular toxicity, as well as for therapeutic application of dsRNA.© 2023. The Author(s).