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f Acyl coenzyme A (acyl-CoA) thioesterases hydrolyze thioester bonds in acyl-CoA metabolites. The majority of mammalian thioesterases are ␣/␤-hydrolases and have been studied extensively. A second class of Hotdog-fold enzymes has been less well described. Here, we present a structural and functional analysis of a new mammalian mitochondrial thioesterase, Them5. Them5 and its paralog, Them4, adopt the classical Hotdog-fold structure and form homodimers in crystals. In vitro, Them5 shows strong thioesterase activity with long-chain acyl-CoAs. Loss of Them5 specifically alters the remodeling process of the mitochondrial phospholipid cardiolipin. Them5 ؊/؊ mice show deregulation of lipid metabolism and the development of fatty liver, exacerbated by a high-fat diet. Consequently, mitochondrial morphology is affected, and functions such as respiration and ␤-oxidation are impaired. The novel mitochondrial acyl-CoA thioesterase Them5 has a critical and specific role in the cardiolipin remodeling process, connecting it to the development of fatty liver and related conditions. T hioesterases participate in lipid metabolism by hydrolyzing thioester bonds in a variety of substrates, including fatty acidcoenzyme A (CoA) esters and palmitoylated proteins (17,19). Based on structural folds and the catalytic reaction mechanism, thioesterases are subdivided into ␣/␤-hydrolases and Hotdogfold thioesterases. The majority of mammalian thioesterases are the well-studied ␣/␤-hydrolases; the second class of Hotdog-fold enzymes has been described to a lesser extent (19). Of the eight Hotdog-fold proteins identified thus far in mammals, only a few participate in biological processes involving hydrolysis of acylCoAs (5). These are cytoplasmic Acot7 involved in eicosanoid metabolism, Acot11 and Acot12, which have a START domain, and cytoplasmic/mitochondrial Them2, which is induced by peroxisome proliferator-activated receptor ␣ (PPAR␣) and has been recently shown to regulate hepatic metabolism (5,8,16,18). The mammalian Hotdog thioesterases identified thus far have tetrameric or hexameric arrangements (26).Them5 is part of the mitochondrial proteome and is predicted to be a member of the Hotdog-fold family (5, 24). We show here that Them5, a novel thioesterase with strong specificity for long and unsaturated fatty acid-CoA esters, has the classical Hotdogfold with six ␤-sheets wrapped around a central ␣-helix. Them4 and Them5 form independent homodimers in crystals, suggesting that they represent a new group of mammalian hotdog thioesterases.Them5 Ϫ/Ϫ mice exhibit an increase in monolysocardiolipin, one of the specific metabolites of cardiolipin (CL). CL is a mitochondrion-specific phospholipid characterized by three to four linoleic acid chains (C 18:2 ) in its mature forms, particularly in mammals. Proper synthesis and remodeling (i.e., maintenance of the right acyl composition) of these highly unsaturated CL molecules is crucial for mitochondrial physiology, since CL interacts with many mitochondrial proteins. In the Barth syndrome, a...

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BackgroundMitochondria are central to the metabolism of cells and participate in many regulatory and signaling events. They are looked upon as dynamic tubular networks. We showed recently that the Carboxy-Terminal Modulator Protein (CTMP) is a mitochondrial protein that may be released into the cytosol under apoptotic conditions.Methodology/Principal FindingsHere we report an unexpected function of CTMP in mitochondrial homeostasis. In this study, both full length CTMP, and a CTMP mutant refractory to N-terminal cleavage and leading to an immature protein promote clustering of spherical mitochondria, suggesting a role for CTMP in the fission process. Indeed, cellular depletion of CTMP led to accumulation of swollen and interconnected mitochondria, without affecting the mitochondrial fusion process. Importantly, in vivo results support the relevance of these findings, as mitochondria from livers of adult CTMP knockout mice had a similar phenotype to cells depleted of CTMP.Conclusions/SignificanceTogether, these results lead us to propose that CTMP has a major function in mitochondrial dynamics and could be involved in the regulation of mitochondrial functions.