title: Bioenergetic and autophagic control by Sirt3 in response to nutrient deprivation in mouse embryonic fibroblasts.
abstract: Sirt3 (sirtuin 3) is an NAD-dependent deacetylase localized to mitochondria. Sirt3 expression is increased in mouse muscle and liver by starvation, which could protect against the starvation-dependent increase in oxidative stress and protein damage. Damaged proteins and organelles depend on autophagy for removal and this is critical for cell survival, but the role of Sirt3 is unclear. To examine this, we used Sirt3-KO (knockout) mouse embryonic fibroblast cells, and found that, under basal conditions, Sirt3-KO cells exhibited increased autophagy flux compared with WT (wild-type) cells. In response to nutrient deprivation, both WT and KO cells exhibited increased basal and ATP-linked mitochondrial respiration, indicating an increased energy demand. Both cells exhibited lower levels of phosphorylated mTOR (mammalian target of rapamycin) and higher autophagy flux, with KO cells exhibiting lower maximal mitochondrial respiration and reserve capacity, and higher levels of autophagy than WT cells. KO cells exhibit higher phospho-JNK (c-Jun N-terminal kinase) and phospho-c-Jun than WT cells under starvation conditions. However, inhibition of JNK activity in Sirt3-KO cells did not affect LC3-I (light chain 3-I) and LC3-II levels, indicating that Sirt3-regulated autophagy is independent of the JNK pathway. Caspase 3 activation and cell death are significantly higher in Sirt3-KO cells compared with WT cells in response to nutrient deprivation. Inhibition of autophagy by chloroquine exacerbated cell death in both WT and Sirt3-KO cells, and by 3-methyadenine exacerbated cell death in Sirt3-KO cells. These data suggest that nutrient deprivation-induced autophagy plays a protective role in cell survival, and Sirt3 decreases the requirement for enhanced autophagy and improves cellular bioenergetics.
authors.affiliation: Center for Free Radical Biology, University of Alabama at Birmingham, 35294, USA.
authors.forename: Qiuli
authors.initials: Q
authors.lastname: Liang
authors.forename: Gloria A
authors.initials: GA
authors.lastname: Benavides
authors.forename: Athanassios
authors.initials: A
authors.lastname: Vassilopoulos
authors.forename: David
authors.initials: D
authors.lastname: Gius
authors.forename: Victor
authors.initials: V
authors.lastname: Darley-Usmar
authors.forename: Jianhua
authors.initials: J
authors.lastname: Zhang
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001343:Autophagy / Q000187:drug effects; D053148:Caspase 3 / Q000378:metabolism; D002478:Cells, Cultured; D003470:Culture Media / Q000378:metabolism; D004622:Embryo, Mammalian / Q000166:cytology; D004734:Energy Metabolism / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D005347:Fibroblasts / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000187:drug effects; D051379:Mice; D018345:Mice, Knockout; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D010085:Oxidative Phosphorylation / Q000187:drug effects; D010766:Phosphorylation / Q000187:drug effects; D020033:Protein Isoforms / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D056566:Sirtuin 3 / Q000235:genetics / Q000378:metabolism*; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism; D050257:Tubulin Modulators / Q000494:pharmacology
chemical_list: D003470:Culture Media; C480860:Map1lc3b protein, mouse; D008869:Microtubule-Associated Proteins; D020033:Protein Isoforms; C525046:Sirt3 protein, mouse; D050257:Tubulin Modulators; C546843:mTOR protein, mouse; D058570:TOR Serine-Threonine Kinases; C505409:Casp3 protein, mouse; D053148:Caspase 3; D056566:Sirtuin 3