Review
Mitochondrial sirtuins

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Abstract

Sirtuins have emerged as important proteins in aging, stress resistance and metabolic regulation. Three sirtuins, SIRT3, 4 and 5, are located within the mitochondrial matrix. SIRT3 and SIRT5 are NAD+-dependent deacetylases that remove acetyl groups from acetyllysine-modified proteins and yield 2′-O-acetyl-ADP-ribose and nicotinamide. SIRT4 can transfer the ADP-ribose group from NAD+ onto acceptor proteins. Recent findings reveal that a large fraction of mitochondrial proteins are acetylated and that mitochondrial protein acetylation is modulated by nutritional status. This and the identification of targets for SIRT3, 4 and 5 support the model that mitochondrial sirtuins are metabolic sensors that modulate the activity of metabolic enzymes via protein deacetylation or mono-ADP-ribosylation. Here, we review and discuss recent progress in the study of mitochondrial sirtuins and their targets.

Introduction

Mitochondria are double-membrane bound organelles that play a central role in energy production. Increasing evidence suggests a key role for mitochondrial dysfunction in diabetes, aging, neurodegenerative disorders, and cancer [1], [2], [3]. Many mitochondrial proteins are multiply acetylated [4], [5] and mitochondrial protein acetylation levels are modulated during calorie restriction and fasting [4], [6]. It has been well established that protein acetylation is dynamic and regulated by the competing enzymatic activities of protein acetyltransferases and protein deacetylases.

Histone/protein deacetylases are enzymes that catalyze the removal of acetyl groups from the ε-amino group of lysine residues and are classified into three groups. Sirtuins, the class III histone deacetylases, are homologous to the yeast transcriptional repressor, Sir2. Unlike previously characterized class I and II histone deacetylases, which catalyze the simple hydrolysis of acetyllysine [7], [8], Sir2 deacetylates lysine residues in a novel chemical reaction that consumes nicotinamide adenine dinucleotide (NAD+) and generates nicotinamide, O-acetyl-ADP-ribose (OAADRr), and the deacetylated substrate [9], [10], [11]. Seven sirtuins have been identified in the human genome [12], [13]. They share a conserved Sir2 catalytic core domain and exhibit variable amino- and carboxyl-terminal extensions that contribute to their unique subcellular localization and may also regulate their catalytic activity. The subcellular distribution, substrate specificity, and cellular functions of sirtuins are quite diverse. SIRT1 is found in the nucleus, where it functions as a transcriptional repressor via histone deacetylation. SIRT1 also regulates transcription by modifying the acetylation levels of transcription factors, such as MyoD, FOXO, p53, and NF-κB [14], [15], [16], [17], [18], [19], [20]. The SIRT2 protein is found in the cytoplasm, where it associates with microtubules and deacetylates lysine 40 of α-tubulin [21]. SIRT6 functions primarily as a histone H3K9 deacetylase and regulates telomeric chromatin [22]. SIRT7 is localized in the nucleolus and functions as a positive regulator of RNA polymerase I transcription [23].

Interestingly, three sirtuins, SIRT3, SIRT4, and SIRT5, are located in mitochondria (Table 1). Here, we review the emerging roles for each of the mitochondrial sirtuins, including their known substrates and functions, and discuss their possible biological roles under normal and pathological conditions.

Section snippets

Reversible mitochondrial protein acetylation

A novel experimental approach combining immuno-affinity purification of peptides carrying acetylated lysine residues after proteolytic digestion of mitochondrial extracts and mass spectroscopic analysis has led to the identification of a large number of mitochondrial acetylated proteins. At least 20% of mitochondrial proteins are acetylated. An early proteomic survey identified 277 acetylation sites on 133 proteins in liver mitochondria from fed and fasted mouse [4]. Proteins from all major

SIRT3

SIRT3 is a soluble protein located in the mitochondrial matrix [31], [32], [33]. The mitochondrial-targeting peptide of human SIRT3 at the N-terminus of the precursor protein is cleaved off after import into mitochondria [31]. The cleavage can be catalyzed by mitochondrial processing peptidase (MPP) in vitro. Two arginine residues, Arg 99 and Arg 100, are essential for cleavage [31] and are conserved among many species [34]. Full protein deacetylase activity of human SIRT3 is obtained only

SIRT4

SIRT4 is localized in the mitochondrial matrix and its first 28 amino acids are removed after import into mitochondria [53], [54]. Expression levels of SIRT4 are relatively high in the liver, heart, kidney and brain [53]. While SIRT3 plays a major role in deacetylation of mitochondrial proteins, SIRT4 has no detectable NAD+-dependent deacetylase activity in vitro [53], [55]. In agreement with these findings, mice lacking SIRT4 show no significant change in mitochondrial protein acetylation

SIRT5

SIRT5 is also localized in the mitochondrial matrix and the N-terminal 36 amino acids are cleaved after import into mitochondria [49], [71]. SIRT5 is broadly expressed in various tissues, with relatively high level in the heart, muscle, brain, liver and kidney [71], [72]. The SIRT5 gene is localized to a chromosomal region associated with malignancies [72].

SIRT5 exhibits weak but detectable deacetylase activity against an acetylated histone H4 peptide [73], as well as chemically acetylated

NAD+ metabolism in mitochondria

Using NAD+ as a cofactor, sirtuins are likely to be sensors for fluctuations of NAD+ in responses to nutrient deprivation. Changes in NAD+/NADH ratios during metabolic stress is well-documented [78], [79]. Recent studies have measured approximately 2- to 2.5-fold increases in NAD+ during prolonged fasting [49], [51]. NAD+ has also been shown to increase (approximately 1.5-fold) during CR [49]. Thus, the measured accumulation in NAD+ during metabolic stress could be responsible for concerted

O-Acetyl-ADP-ribose (OAADPr)

OAADPr is produced by sirtuins during the deacetylation reaction by the cleavage of NAD+ into nicotinamide and ADP-ribose (ADPr). The latter serves as an acceptor for the acetyl group generating OAADPr. Various enzyme activities, such as hydrolases or esterases, can degrade OAADPr into free ADPr plus acetate [91], [92]. ADPr can be further hydrolyzed into AMP [92]. Both OAADPr and ADPr can act as second messengers. Microinjection of OAADPr or ADPr causes a delay/block in oocyte maturation [93].

Conclusions

Because of their mitochondrial subcellular localization and their dependence on NAD+, SIRT3, SIRT4 and SIRT5 are in a position to sense and regulate various mitochondrial metabolic pathways. Indeed, a metabolic regulatory role is emerging for the mitochondrial sirtuins. SIRT3 and SIRT5, the two known mitochondrial deacetylases, both regulate unique metabolic enzymes in the mitochondrial matrix, suggesting a specific non-overlapping role for each protein. SIRT4, a mitochondrial enzyme

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    These authors contributed equally to this work.

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