Faculty Bibliography

Quantitative and qualitative alterations of mitochondrial cardiolipin have been implicated in the pathogenesis of Barth syndrome, an X-linked cardioskeletal myopathy caused by a deficiency in tafazzin, an enzyme in the cardiolipin remodeling pathway. We have generated and previously reported a tafazzin-deficient Drosophila model of Barth syndrome that is characterized by low cardiolipin concentration, abnormal cardiolipin fatty acyl composition, abnormal mitochondria, and poor motor function. Here, we first show that tafazzin deficiency in Drosophila disrupts the final stage of spermatogenesis, spermatid individualization, and causes male sterility. This phenotype can be genetically suppressed by inactivation of the gene encoding a calcium-independent phospholipase A(2), iPLA2-VIA, which also prevents cardiolipin depletion/monolysocardiolipin accumulation, although in wild-type flies inactivation of the iPLA2-VIA does not affect the molecular composition of cardiolipin. Furthermore, we show that treatment of Barth syndrome patients' lymphoblasts in tissue culture with the iPLA(2) inhibitor, bromoenol lactone, partially restores their cardiolipin homeostasis. Taken together, these findings establish a causal role of cardiolipin deficiency in the pathogenesis of Barth syndrome and identify iPLA2-VIA as an important enzyme in cardiolipin deacylation, and as a potential target for therapeutic intervention

In this article, the formation of prokaryotic and eukaryotic cardiolipin is reviewed in light of its biological function. I begin with a detailed account of the structure of cardiolipin, its stereochemistry, and the resulting physical properties, and I present structural analogs of cardiolipin that occur in some organisms. Then I continue to discuss i) the de novo formation of cardiolipin, ii) its acyl remodeling, iii) the assembly of cardiolipin into biological membranes, and iv) the degradation of cardiolipin, which may be involved in apoptosis and mitochondrial fusion. Thus, this article covers the entire metabolic cycle of this unique phospholipid. It is shown that mitochondria produce cardiolipin species with a high degree of structural uniformity and molecular symmetry, among which there is often a dominant form with four identical acyl chains. The subsequent assembly of cardiolipin into functional membranes is largely unknown, but the analysis of crystal structures of membrane proteins has revealed a first glimpse into the underlying principles of cardiolipin-protein interactions. Disturbances of cardiolipin metabolism are crucial in the pathophysiology of human Barth syndrome and perhaps also play a role in diabetes and ischemic heart disease

Bis(monoacylglycero)phosphate (BMP) is a unique lipid enriched in the late endosomes participating in the trafficking of lipids and proteins through this organelle. The de novo biosynthesis of BMP has not been clearly demonstrated. We investigated whether phosphatidylglycerol (PG) and cardiolipin (CL) could serve as precursors of de novo BMP synthesis using two different cellular models: CHO cells deficient in phosphatidylglycerophosphate (PGP) synthase, the enzyme responsible for the first step of PG synthesis; and human lymphoblasts from patients with Barth syndrome (BTHS), characterized by mutations in tafazzin, an enzyme implicated in the deacylation-reacylation cycle of CL. The biosynthesis of both PG and BMP was reduced significantly in the PGP synthase-deficient CHO mutants. Furthermore, overexpression of PGP synthase in the deficient mutants induced an increase of BMP biosynthesis. In contrast to CHO mutants, BMP biosynthesis and its fatty acid composition were not altered in BTHS lymphoblasts. Our results thus suggest that in mammalian cells, PG, but not CL, is a precursor of the de novo biosynthesis of BMP. Despite the decrease of de novo synthesis, the cellular content of BMP remained unchanged in CHO mutants, suggesting that other pathway(s) than de novo biosynthesis are also used for BMP synthesis

Barth syndrome (BTHS) is a mitochondrial disorder that is caused by mutations in the tafazzin gene, which affects phospholipid composition. To determine whether this defect leads to alterations in the internal three-dimensional organization of mitochondrial membranes, we applied electron microscopic tomography to lymphoblast mitochondria from BTHS patients and controls. Tomograms were formed from 50 and 150 nm sections of chemically fixed lymphoblasts and the data were used to manually segment volumes of relevant structural details. Normal lymphoblast mitochondria contained well-aligned, lamellar cristae with slot-like junctions to the inner boundary membrane. In BTHS, mitochondrial size was more variable and the total mitochondrial volume per cell increased mainly due to clusters of fragmented mitochondria inside nuclear invaginations. However, mitochondria showed reduced cristae density, less cristae alignment, and inhomogeneous cristae distribution. Three-dimensional reconstruction of BTHS mitochondria revealed zones of adhesion of the opposing inner membranes, causing obliteration of the intracrista space. We found small isolated patches of adhesion as well as extended adhesion zones, resulting in sheets of collapsed cristae packaged in multiple concentric layers. We also found large tubular structures (diameter 30-150 nm) that appeared to be derivatives of the adhesion zones. The data suggest that mitochondrial abnormalities of BTHS involve adhesions of inner mitochondrial membranes with subsequent collapse of the intracristae space

Tafazzin is a putative enzyme that is involved in cardiolipin metabolism, it may carry mutations responsible for Barth syndrome. To identify the biochemical reaction catalyzed by tafazzin, we expressed the full-length isoform of Drosophila melanogaster tafazzin in a baculovirus-Sf9 insect cell system. Tafazzin expression induced a new enzymatic function in Sf9 cell mitochondria, namely 1-palmitoyl-2-[14C]linoleoyl-phosphatidylcholine:monolysocardiolipin linoleoyltransferase. We also found evidence for the reverse reaction, because tafazzin expression caused transfer of acyl groups from phospholipids to 1-[14C]palmitoyl-2-lyso-phosphatidylcholine. An affinity-purified tafazzin construct, tagged with the maltose-binding protein, catalyzed both forward and reverse transacylations between cardiolipin and phosphatidylcholine, but was unable to utilize CoA or acyl-CoA as substrates. Whereas tafazzin supported transacylations between various phospholipid-lysophospholipid pairs, it showed the highest rate for the phosphatidylcholine-cardiolipin transacylation. Transacylation activities were about 10-fold higher for linoleoyl groups than for oleoyl groups, and they were negligible for arachidonoyl groups. The data show that Drosophila tafazzin is a CoA-independent, acyl-specific phospholipid transacylase with substrate preference for cardiolipin and phosphatidylcholine