Abstract
The mitochondrial dimeric phospholipid cardiolipin is characterized by a high degree of unsaturation of its acyl chains, which is important for its functional interaction with mitochondrial enzymes. The unusual fatty acid composition of cardiolipin molecular species emerges from a de novo synthesized “premature” species by extensive acyl chain remodeling that involves as yet only partially identified acyltransferases and phospholipases. Recently, the yeast protein Taz1p was shown to function as a transacylase, which catalyzes the reacylation of monolysocardiolipin to mature cardiolipin. A defect in the orthologous human TAZ gene is associated with Barth syndrome, a severe genetic disorder, which may lead to cardiac failure and death in childhood. We now identified the protein encoded by reading frame YGR110W as a mitochondrial phospholipase, which deacylates de novo synthesized cardiolipin. Ygr110wp has a strong substrate preference for palmitic acid residues and functions upstream of Taz1p, to generate monolysocardiolipin for Taz1p-dependent reacylation with unsaturated fatty acids. We therefore rename the Ygr110wp as Cld1p (cardiolipin-specific deacylase 1).
Highlights
EXPERIMENTAL PROCEDURESMedia and Growth Conditions—Yeast strains were grown in YPD medium at 30 °C on a rotary shaker with vigorous aeration
Cardiolipin (CL)2 is a dimeric phospholipid enriched in mitochondrial membranes [1]
In this work we show that the yeast open reading frame YGR110W encodes a mitochondrial protein, which functions as phospholipase A (PLA) in vitro and is involved in generating MLCL in vivo
Summary
Media and Growth Conditions—Yeast strains were grown in YPD medium at 30 °C on a rotary shaker with vigorous aeration. Standard YPD medium contained 1% yeast extract (Difco), 2% glucose (Merck), and 2% Bacto-peptone (Difco). For plate drop tests cells were grown in YPD for 16 h, and cell numbers were estimated using a Casy cell counter (Scharfe Systems). Serial dilutions of cell cultures were prepared in microtiter plates (5 ϫ 105 to 5 ϫ 102 cells per well). Cells were spotted on YPD, YNBLac (0.67% Yeast Nitrogen Base (Difco), vitamins, trace elements [17], and 3% lactic acid, Merck, adjusted to pH 5.5) or YNB synthetic medium containing 3% glycerol (Roth) and 1% ethanol (Merck). Yeast transformants carrying plasmids were grown on uracil-free YNB synthetic medium containing 2%
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