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Oxygen flux analysis to understand the biological function of sirtuins.

Publication ,  Journal Article
Wang, D; Green, MF; McDonnell, E; Hirschey, MD
Published in: Methods Mol Biol
2013

The sirtuins are a family of highly conserved NAD(+)-dependent lysine deacylases with important roles in metabolic regulation. Of the seven mammalian sirtuins, three localize to the mitochondria: SIRT3, SIRT4, and SIRT5. Mitochondrial sirtuins are crucial regulators of the metabolic network that controls energy homeostasis and impacts cancer, obesity, diabetes, mitochondrial diseases, metabolic disorders, and many other human diseases of aging. To best study the mitochondrial function of the sirtuins, we have employed an oxygen flux analyzer as a tool to track and record the extracellular oxygen consumption rate and acidification rate that reflects mitochondrial respiration and glycolysis, respectfully. Here we described the methods using this assay to study the substrate utilization and mitochondrial function in a human hepatocellular carcinoma cell line, Huh7. Additionally, we have generated a stable SIRT4 knocked-down Huh7 cell line. With this cell line, we evaluated how the absence of SIRT4 affects mitochondrial function, glucose utilization, glutamine oxidation, and fatty acid oxidation in these cells.

Duke Scholars

Published In

Methods Mol Biol

DOI

EISSN

1940-6029

Publication Date

2013

Volume

1077

Start / End Page

241 / 258

Location

United States

Related Subject Headings

  • Tumor Cells, Cultured
  • Sirtuins
  • RNA, Small Interfering
  • Oxygen Consumption
  • Oxygen
  • Oxidation-Reduction
  • Mitochondrial Proteins
  • Mitochondria
  • Liver Neoplasms
  • Immunoblotting
 

Citation

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ICMJE
MLA
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Wang, D., Green, M. F., McDonnell, E., & Hirschey, M. D. (2013). Oxygen flux analysis to understand the biological function of sirtuins. Methods Mol Biol, 1077, 241–258. https://doi.org/10.1007/978-1-62703-637-5_16
Wang, Dongning, Michelle F. Green, Eoin McDonnell, and Matthew D. Hirschey. “Oxygen flux analysis to understand the biological function of sirtuins.Methods Mol Biol 1077 (2013): 241–58. https://doi.org/10.1007/978-1-62703-637-5_16.
Wang D, Green MF, McDonnell E, Hirschey MD. Oxygen flux analysis to understand the biological function of sirtuins. Methods Mol Biol. 2013;1077:241–58.
Wang, Dongning, et al. “Oxygen flux analysis to understand the biological function of sirtuins.Methods Mol Biol, vol. 1077, 2013, pp. 241–58. Pubmed, doi:10.1007/978-1-62703-637-5_16.
Wang D, Green MF, McDonnell E, Hirschey MD. Oxygen flux analysis to understand the biological function of sirtuins. Methods Mol Biol. 2013;1077:241–258.

Published In

Methods Mol Biol

DOI

EISSN

1940-6029

Publication Date

2013

Volume

1077

Start / End Page

241 / 258

Location

United States

Related Subject Headings

  • Tumor Cells, Cultured
  • Sirtuins
  • RNA, Small Interfering
  • Oxygen Consumption
  • Oxygen
  • Oxidation-Reduction
  • Mitochondrial Proteins
  • Mitochondria
  • Liver Neoplasms
  • Immunoblotting