Background: Cells lacking both cardiolipin and mitochondrial phosphatidylethanolamine are inviable, suggesting that these lipids have overlapping functions. Results: The loss of both lipids leads to decreased mitochondrial fusion and fragmented mitochondria. Conclusion: One overlapping function of these lipids is in mitochondrial fusion. Significance: Decreased mitochondrial fusion may partly explain the variation in clinical presentation observed in Barth syndrome.

Saccharomyces cerevisiae mitochondria contain enzymes required for synthesis of the phospholipids cardiolipin (CL) and phosphatidylethanolamine (PE), which are enriched in mitochondrial membranes. Previous studies indicated that PE may compensate for the lack of CL, and vice versa. These data suggest that PE and CL have overlapping functions and that the absence of both lipids may be lethal. To address this hypothesis, we determined whether the crd1⌬ mutant, which lacks CL, was viable in genetic backgrounds in which PE synthesis was genetically blocked. Deletion of the mitochondrial PE pathway gene PSD1 was synthetically lethal with the crd1⌬ mutant, whereas deletion of the Golgi and endoplasmic reticulum pathway genes PSD2 and DPL1 did not result in synthetic lethality. A 20-fold reduction in phosphatidylcholine did not affect the growth of crd1⌬ cells. Supplementation with ethanolamine, which led to increased PE synthesis, or with propanolamine, which led to synthesis of the novel phospholipid phosphatidylpropanolamine, failed to rescue the synthetic lethality of the crd1⌬psd1⌬ cells. These results suggest that mitochondrial biosynthesis of PE is essential for the viability of yeast mutants lacking CL.The phospholipid composition of the mitochondrial membrane is unique in that it is highly enriched in cardiolipin (CL) 2 and phosphatidylethanolamine (PE) (1, 2). CL is a dimeric glycerophospholipid that is synthesized exclusively in mitochondria and plays an important role in oxidative phosphorylation and mitochondrial membrane biogenesis (3). In contrast, PE biosynthesis occurs via multiple pathways (4). PE is commonly present in all subcellular membranes, although PE levels are highest in the mitochondrial membrane (2). PE and CL have similar physical properties in that they have a propensity toward the formation of nonbilayer, inverted hexagonal (H II ) phase structures (5, 6). The local, transient formation of nonbilayer structures is thought to play an important role in vital cellular processes, such as vesicle formation, vesicle-mediated protein trafficking, and membrane fusion (7). In addition, nonbilayer lipids affect integration of proteins into the membrane, their lateral movement within the membrane, and the function and folding of certain integral membrane proteins (8).In the yeast Saccharomyces cerevisiae, phospholipid biosynthesis is compartmentalized in various subcellular organelles, including the Golgi body, endoplasmic reticulum, and mitochondria (Fig. 1). CL biosynthesis occurs in three steps, all catalyzed by enzymes present in the mitochondria. The first step, catalyzed by phosphatidylglycerol phosphate (PGP) synthase, is the synthesis of PGP from CDP-diacylglycerol and glycerol 3-phosphate. PGP phosphatase dephosphorylates PGP to phosphatidylglycerol. In the final step, CL synthase catalyzes the formation of CL from phosphatidylglycerol and CDP-diacylglycerol (9, 10). PGP synthase and CL synthase have been characterized in yeast, and the genes encoding these enzymes, PGS1 (11, 12) and CRD1 ...

Huntington's disease is initiated by the expression of a CAG repeat-encoded polyglutamine region in full-length huntingtin, with dominant effects that vary continuously with CAG size. The mechanism could involve a simple gain of function or a more complex gain of function coupled to a loss of function (e.g. dominant negative-graded loss of function). To distinguish these alternatives, we compared genome-wide gene expression changes correlated with CAG size across an allelic series of heterozygous CAG knock-in mouse embryonic stem (ES) cell lines (Hdh(Q20/7), Hdh(Q50/7), Hdh(Q91/7), Hdh(Q111/7)), to genes differentially expressed between Hdh(ex4/5/ex4/5) huntingtin null and wild-type (Hdh(Q7/7)) parental ES cells. The set of 73 genes whose expression varied continuously with CAG length had minimal overlap with the 754-member huntingtin-null gene set but the two were not completely unconnected. Rather, the 172 CAG length-correlated pathways and 238 huntingtin-null significant pathways clustered into 13 shared categories at the network level. A closer examination of the energy metabolism and the lipid/sterol/lipoprotein metabolism categories revealed that CAG length-correlated genes and huntingtin-null-altered genes either were different members of the same pathways or were in unique, but interconnected pathways. Thus, varying the polyglutamine size in full-length huntingtin produced gene expression changes that were distinct from, but related to, the effects of lack of huntingtin. These findings support a simple gain-of-function mechanism acting through a property of the full-length huntingtin protein and point to CAG-correlative approaches to discover its effects. Moreover, for therapeutic strategies based on huntingtin suppression, our data highlight processes that may be more sensitive to the disease trigger than to decreased huntingtin levels.

Huntingtin is a large HEAT repeat protein first identified in humans, where a polyglutamine tract expansion near the amino terminus causes a gain-of-function mechanism that leads to selective neuronal loss in Huntington's disease (HD). Genetic evidence in humans and knock-in mouse models suggests that this gain-of-function involves an increase or deregulation of some aspect of huntingtin's normal function(s), which remains poorly understood. As huntingtin shows evolutionary conservation, a powerful approach to discovering its normal biochemical role(s) is to study the effects caused by its deficiency in a model organism with a short life-cycle that comprises both cellular and multicellular developmental stages. To facilitate studies aimed at detailed knowledge of huntingtin's normal function(s), we generated a null mutant of hd, the HD ortholog in Dictyostelium discoideum. Dictyostelium cells lacking endogenous huntingtin were viable but during development did not exhibit the typical polarized morphology of Dictyostelium cells, streamed poorly to form aggregates by accretion rather than chemotaxis, showed disorganized F-actin staining, exhibited extreme sensitivity to hypoosmotic stress, and failed to form EDTA-resistant cell–cell contacts. Surprisingly, chemotactic streaming could be rescued in the presence of the bivalent cations Ca2+ or Mg2+ but not pulses of cAMP. Although hd − cells completed development, it was delayed and proceeded asynchronously, producing small fruiting bodies with round, defective spores that germinated spontaneously within a glassy sorus. When developed as chimeras with wild-type cells, hd − cells failed to populate the pre-spore region of the slug. In Dictyostelium, huntingtin deficiency is compatible with survival of the organism but renders cells sensitive to low osmolarity, which produces pleiotropic cell autonomous defects that affect cAMP signaling and as a consequence development. Thus, Dictyostelium provides a novel haploid organism model for genetic, cell biological, and biochemical studies to delineate the functions of the HD protein.

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