20.500.12556/RUL-160122 Inhibition of the ubiquitin-proteasome system reduces the abundance of pyruvate dehydrogenase kinase 1 in cultured myotubes Pyruvate dehydrogenase kinase (PDK), which phosphorylates the pyruvate dehydrogenase complex, regulates glucose metabolism in skeletal muscle. PDK1, an isozyme whose expression is controlled by hypoxia-inducible factor-1α (HIF-1α), is thought to play a role in muscle adaptation to hypoxia. While transcriptional upregulation of PDK1 by HIF-1α is well characterised, mechanisms controlling proteolysis of PDK1 in skeletal muscle have not been thoroughly investigated. Proteasome inhibitor MG132 paradoxically reduced the abundance of PDK1 in human cancer cells and rat L6 myotubes, suggesting that MG132 might direct PDK1 towards autophagic degradation. The objectives of our current study were to determine (1) whether MG132 suppresses PDK1 levels in primary human myotubes, (2) whether chloroquine, an inhibitor of autophagy, prevents MG132-induced suppression of PDK1 in L6 myotubes, and (3) whether PYR-41, an inhibitor of ubiquitination, suppresses PDK1 in L6 myotubes. Using qPCR and/or immunoblotting, we found that despite markedly upregulating HIF-1α protein, MG132 did not alter the PDK1 expression in cultured primary human myotubes, while it suppressed both PDK1 mRNA and protein in L6 myotubes. The PDK1 levels in L6 myotubes were suppressed also during co-treatment with chloroquine and MG132. PYR-41 markedly increased the abundance of HIF-1α in primary human and L6 myotubes, while reducing the abundance of PDK1. In L6 myotubes treated with PYR-41, chloroquine increased the abundance of the epidermal growth factor receptor, but did not prevent the suppression of PDK1. Collectively, our results suggest that cultured myotubes degrade PDK1 via a pathway that cannot be inhibited by MG132, PYR-41, and/or chloroquine. chloroquine hypoxia-inducible factor-1α HIF-1α MG132 myotubes PYR-41 pyruvate dehydrogenase kinase 1 PDK1 klorokin hipoksijsko inducibilni faktor-1α HIF-1α miocevke piruvat dehidrogenaza kinaza 1 PDK1 true false true Angleški jezik Slovenski jezik Članek v reviji 2024-08-21 11:34:53 2024-08-21 11:34:57 2024-08-22 03:58:11 0000-00-00 00:00:00 2024 0 0 Str. 155–169 iss. 3 Vol. 45 2024 0000-00-00 Zaloznikova Objavljeno NiDoloceno 0000-00-00 0000-00-00 0000-00-00 616-092 1573-2657 10.1007/s10974-024-09679-3 203573251 s10974-024-09679-3.pdf s10974-024-09679-3.pdf 1 F8AAE35EBE6D1A4C8A8B939C88510D72 ad6a4cfbb15c9cf0289a0d2f7d86f2cd1b8dc4588d13938ad224174e318375cf b094532b-5fa0-11ef-8f74-0050569b8976 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=188603 https://link.springer.com/article/10.1007/s10974-024-09679-3 1 6c5da597-5fa0-11ef-8f74-0050569b8976 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=188602 Medicinska fakulteta 0 0 0