Inhibition of the ubiquitin-proteasome system reduces the abundance of pyruvate dehydrogenase kinase 1 in cultured myotubes

Kociper, Blaž (author), Škorja, Nives (author), Ogrizek, Ivana (author), Miš, Katarina (author), Pirkmajer, Sergej (author)

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.

• Language: English
• Keywords: chloroquine, hypoxia-inducible factor-1α, HIF-1α, MG132, myotubes, PYR-41, pyruvate dehydrogenase kinase 1, PDK1
• Work type: Article
• Typology: 1.01 - Original Scientific Article
• Organization: MF - Faculty of Medicine
• Publication status: Published
• Publication version: Version of Record
• Year: 2024
• Number of pages: Str. 155–169
• Numbering: Vol. 45, iss. 3
• PID: 20.500.12556/RUL-160122
• UDC: 616-092
• ISSN on article: 1573-2657
• DOI: 10.1007/s10974-024-09679-3
• COBISS.SI-ID: 203573251
• Publication date in RUL: 21.08.2024

Record is a part of a journal

• Title: Journal of muscle research and cell motility
• Shortened title: J. muscle res. cell motil.
• Publisher: Springer Nature
• ISSN: 1573-2657
• COBISS.SI-ID: 513197337

Licences

• License: CC BY 4.0, Creative Commons Attribution 4.0 International
• Link: http://creativecommons.org/licenses/by/4.0/
• Description: This is the standard Creative Commons license that gives others maximum freedom to do what they want with the work as long as they credit the author.

Secondary language

• Language: Slovenian
• Keywords: klorokin, hipoksijsko inducibilni faktor-1α, HIF-1α, miocevke, piruvat dehidrogenaza kinaza 1, PDK1

Projects

• Funder: ARIS - Slovenian Research and Innovation Agency
• Project number: P3-0043
• Name: Molekularni mehanizmi razvoja in delovanja skeletne mišice

• Funder: ARIS - Slovenian Research and Innovation Agency
• Project number: J7-3153
• Name: Molekularni mehanizmi specifičnosti pri uravnavanju izločanja in delovanja citokinov mišičnega izvora