Nutrient sensing by the mitochondrial transcription machinery dictates oxidative phosphorylation
Lijun Liu, Minwoo Nam, Wei Fan, Thomas E. Akie, David C. Hoaglin, Guangping Gao, John F. Keaney Jr., Marcus P. Cooper
Lijun Liu, Minwoo Nam, Wei Fan, Thomas E. Akie, David C. Hoaglin, Guangping Gao, John F. Keaney Jr., Marcus P. Cooper
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Research Article Metabolism

Nutrient sensing by the mitochondrial transcription machinery dictates oxidative phosphorylation

Abstract

Sirtuin 3 (SIRT3), an important regulator of energy metabolism and lipid oxidation, is induced in fasted liver mitochondria and implicated in metabolic syndrome. In fasted liver, SIRT3-mediated increases in substrate flux depend on oxidative phosphorylation (OXPHOS), but precisely how OXPHOS meets the challenge of increased substrate oxidation in fasted liver remains unclear. Here, we show that liver mitochondria in fasting mice adapt to the demand of increased substrate oxidation by increasing their OXPHOS efficiency. In response to cAMP signaling, SIRT3 deacetylated and activated leucine-rich protein 130 (LRP130; official symbol, LRPPRC), promoting a mitochondrial transcriptional program that enhanced hepatic OXPHOS. Using mass spectrometry, we identified SIRT3-regulated lysine residues in LRP130 that generated a lysine-to-arginine (KR) mutant of LRP130 that mimics deacetylated protein. Compared with wild-type LRP130 protein, expression of the KR mutant increased mitochondrial transcription and OXPHOS in vitro. Indeed, even when SIRT3 activity was abolished, activation of mitochondrial transcription and OXPHOS by the KR mutant remained robust, further highlighting the contribution of LRP130 deacetylation to increased OXPHOS in fasted liver. These data establish a link between nutrient sensing and mitochondrial transcription that regulates OXPHOS in fasted liver and may explain how fasted liver adapts to increased substrate oxidation.

Authors

Lijun Liu, Minwoo Nam, Wei Fan, Thomas E. Akie, David C. Hoaglin, Guangping Gao, John F. Keaney Jr., Marcus P. Cooper

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Figure 7

During the fasted response, SIRT3 deacetylates LRP130 in liver mitochondria.

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During the fasted response, SIRT3 deacetylates LRP130 in liver mitochond...
(A) Representative immunoblot and protein quantification showing reduced acetylation of LRP130 (Ac-LRP130) in mitochondria isolated from fasted mouse liver (n = 3 experiments). (B) Immunoblot and protein quantification showing hyperacetylation of LRP130 in mitochondria isolated from liver deficient for SIRT3 (n = 4). (C) Immunoblot showing deacetylation of LRP130 in H2.35 cells stably expressing SIRT3 upon treatment with 500 μM NAD+. (D) SIRT3 and LRP130 coimmunoprecipitated, using ectopically expressed SIRT3-FLAG protein and endogenous LRP130. (E) Using purified proteins, SIRT3 robustly deacetylated the C terminus of LRP130. Deacetylation of LRP130 was inhibited by 12.5 mM nicotinamide (NAM). Shown is 1 representative of 4 independent experiments. Similar, but less robust, deacetylation was obtained for the N terminus of LRP130 (not shown). (F) Acetylated LRP130 fragments were incubated with control buffer or purified SIRT3 protein, then subjected to mass spectrometry. Lysines showing greater than 50% deacetylation by SIRT3 are graphed (gray and black bars). Percent deacetylation was calculated as 1 – (SIRT3 signal/control signal). 7 lysines were mutated to arginines (gray bars), generating LRP130-7KR, which mimics deacetylated protein. Data are mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, 2-tailed unpaired Student’s t test (A and B).