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Hirschey et al., 2011 Molecular Cell (2011) Presentation By: Humna Fayyaz August 25, 2011

SIRT3 Deficiency and Mitochondrial Protein Hyperacetylation Accelerate the Development of the Metabolic Syndrome. Hirschey et al., 2011 Molecular Cell (2011) Presentation By: Humna Fayyaz August 25, 2011. Metabolic Syndrom e. Defined by metabolic abnormalities including: central obesity

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Hirschey et al., 2011 Molecular Cell (2011) Presentation By: Humna Fayyaz August 25, 2011

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  1. SIRT3 Deficiency and Mitochondrial Protein Hyperacetylation Accelerate the Development of the Metabolic Syndrome • Hirschey et al., 2011 • Molecular Cell (2011) • Presentation By: Humna Fayyaz • August 25, 2011

  2. Metabolic Syndrome • Defined by metabolic abnormalities including: • central obesity • insulin resistance • Diminished cell response to insulin in transport of sugar glucose from blood  muscles and other tissues • hyperlipidemia, hyperglycemia, hypertension

  3. Research suggests: • High-fat diet (HFD) feeding induces: 1.) hepatic mitochondrial protein hyperacetylation • *acetylation: a regulatory post-translational protein modification • controls the enzymatic activity of mitochondrial metabolic enzymes • occurring in liver in which acetyl-CoA combines with metabolic products 2.) Down-regulation of a major mitochondrial protein deacetylase, SIRT3 gene

  4. A: Western blot analysis with acetyllysine-specific antiserum • Mitochondria isolated from livers of WT mice, fed SD, acute HFD, and chronic HFD • Western Blot: to detect specific proteins in a given sample of tissue extract • Based on denaturing conditions (length, 3D structure) • Then probed using specific antibodies • B: WT mice fed a SD or HFD over interval of time • Western blot analysis with antiserum specific for SIRT3

  5. Variability in SIRT3 gene • Tested if variability in SIRT3 gene was correlated to increased susceptibility for developing metabolic dysfunction in humans (Hirschey et al., 2011) • identified a single nucleotide polymorphism (SNP, pronounced snip) in the human SIRT3 gene • SNP could be suggestive of a genetic association with the metabolic syndrome

  6. SNP • Single nucleotide polymorphism: occurs at a particular nucleotide site--DNA molecules in the population often differ in the identity of the nucleotide pair • Example: Figure, difference occurs at a single base-pair location

  7. Process/Results • Used a “gene candidate approach” • *GCA: analyzing the presence of a mutation in cases and controls, leading to the conclusion of the mutation being considered a risk marker or not • Took 834 gene samples from patients with fatty liver disease • tested if certain SNPs in SIRT3 gene increased the patient’s vulnerability to more complex metabolic diseases • Measured 13 SNPs, 9 of which were found in the patient cohort • rs7934919 and rs11246020 in “near-perfect linkage” were associated with metabolic syndrome

  8. Analysis • Haplotype analysis: to see if a set of SNPs are statistically associated • Performed on rs-919 and rs-020 and determined these SNPs are surrogates of each other • Associations for rs-020 were strongest, further evidence provided by regression analysis • rs-020 “A” minor allele associated with increased risk of Metabolic Syndrome

  9. Further Investigation of rs-020 • METSIM (Metabolic Syndrome in Men) cohort of 8000 Finish Men • Using additive, dominant, and recessive genetic models, the association between rs-020 minor allele “A” showed a positive association with criteria for the Metabolic Syndrome • Example of additive genetic model: If risk conferred by an allele is increased r-fold for heterozygous and 2r-fold for homozygous

  10. Nonsynonymous Point Mutation (NPM) • NPM encoded by rs-020 (change from valine isoleucine at reside 208 of the SIRT3 polypeptide) is SIRT3-V208l • Procedure to verify SIRT3-V208l’s behavior • Tested deacetylase activity of WT SIRT3, SIRT3-V208l, and inactive SIRT3-H248Y in E. coli • Initial rates of radioactive release were measured (as a function of [NAD+] • Results: SIRT3-V208l mutation reduces efficiency by 34% compared to WT SIRT3 • Study may lead to explanation of how patients with V208 mutation have increased vulnerability to MetSyn

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