Ageing results from complex genetically and epigenetically programmed processes that are elicited in part by noxious or stressful events that cause programmed cell death. Here, we report that administration of spermidine, a natural polyamine whose intracellular concentration declines during human ageing, markedly extended the lifespan of yeast, flies and worms, and human immune cells. In addition, spermidine administration potently inhibited oxidative stress in ageing mice. In ageing yeast, spermidine treatment triggered epigenetic deacetylation of histone H3 through inhibition of histone acetyltransferases (HAT), suppressing oxidative stress and necrosis. Conversely, depletion of endogenous polyamines led to hyperacetylation, generation of reactive oxygen species, early necrotic death and decreased lifespan. The altered acetylation status of the chromatin led to significant upregulation of various autophagy-related transcripts, triggering autophagy in yeast, flies, worms and human cells. Finally, we found that enhanced autophagy is crucial for polyamine-induced suppression of necrosis and enhanced longevity.

Endonuclease G (EndoG) is located in mitochondria yet translocates into the nucleus of apoptotic cells during human degenerative diseases. Nonetheless, a direct involvement of EndoG in cell-death execution remains equivocal, and the mechanism for mitochondrio-nuclear translocation is not known. Here, we show that the yeast homolog of EndoG (Nuc1p) can efficiently trigger apoptotic cell death when excluded from mitochondria. Nuc1p induces apoptosis in yeast independently of metacaspase or of apoptosis inducing factor. Instead, the permeability transition pore, karyopherin Kap123p, and histone H2B interact with Nuc1p and are required for cell death upon Nuc1p overexpression, suggesting a pathway in which mitochondrial pore opening, nuclear import, and chromatin association are successively involved in EndoG-mediated death. Deletion of NUC1 diminishes apoptotic death when mitochondrial respiration is increased but enhances necrotic death when oxidative phosphorylation is repressed, pointing to dual--lethal and vital--roles for EndoG.

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). Cardiolipin (CL)2 is a dimeric phospholipid specifically enriched in mitochondrial membranes (1). It plays an important role in mitochondrial structure and function and stabilizes respiratory chain super complexes and individual electron transport complexes (2-4). The CL biosynthetic pathway is well characterized in the yeast Saccharomyces cerevisiae, and the enzymes catalyzing the three sequential reactions involved are all associated with mitochondrial membranes (4, 5). CL synthesis shares the same precursor, CDP-diacylglycerol, with the main cellular phospholipids, yet it differs significantly from other phospholipids in its acyl-chain composition that is characterized by a high degree of unsaturated fatty acids. Although cardiolipin was believed to be essential to support mitochondrial function, mutants defective in cardiolipin synthase (Crd1p) are viable and display only moderate defects of mitochondrial function (6, 7). More severe is a defect in the first committed step of CL synthesis, catalyzed by phosphatidylglycerolphosphate synthase Pgs1p/Pel1p (8). pgs1 mutants are temperature-sensitive, unable to grow on non-fermentable carbon sources for growth, and petite lethal, i.e. dependent on intact mitochondrial DNA for survival (8). More subtle mitochondrial phenotypes emerge from alterations of cardiolipin acyl-chain remodeling. "Premature" cardiolipin synthesized by Crd1p undergoes significant remodeling of its acyl-chain composition (9). Mature CL in yeast contains mainly palmitoleic acid (C16:1) and oleic acid (C18:1), which distinguishes CL from most other phospholipids by its high degree of unsaturation (4, 5, 10). In humans, a severe genetic disorder, the Barth syndrome, is associated with defective CL acyl-chain remodeling (11,12). This disease is caused by mutations in the TAZ gene encoding Tafazzin, for which a functional ortholog also exis...

The lysosomal endoprotease cathepsin D (CatD) is an essential player in general protein turnover and specific peptide processing. CatD-deficiency is associated with neurodegenerative diseases, whereas elevated CatD levels correlate with tumor malignancy and cancer cell survival. Here, we show that the CatD ortholog of the budding yeast Saccharomyces cerevisiae (Pep4p) harbors a dual cytoprotective function, composed of an anti-apoptotic part, conferred by its proteolytic capacity, and an anti-necrotic part, which resides in the protein's proteolytically inactive propeptide. Thus, deletion of PEP4 resulted in both apoptotic and necrotic cell death during chronological aging. Conversely, prolonged overexpression of Pep4p extended chronological lifespan specifically through the protein's anti-necrotic function. This function, which triggered histone hypoacetylation, was dependent on polyamine biosynthesis and was exerted via enhanced intracellular levels of putrescine, spermidine and its precursor S-adenosyl-methionine. Altogether, these data discriminate two pro-survival functions of yeast CatD and provide first insight into the physiological regulation of programmed necrosis in yeast.

The yeast vacuole plays a crucial role in cell homeostasis including pH regulation and degradation of proteins and organelles. Class C VPS genes code for proteins essential for vacuolar and endosomal vesicle fusion, their deletion results in the absence of a detectable vacuole. We found that single gene deletions of class C VPS genes result in a drastically enhanced sensitivity to treatment with acetic acid whereas sensitivity towards H2O2 remains largely unaffected. Interestingly acetic acid treatment known as an established inducer of yeast apoptosis leads to necrosis in class C VPS deletion strains. Their intracellular pH drops from 6.7 to 5.5 after acetic acid treatment, while in wild type the pH drops to just 6.3. When the intracellular pH in wild type is lowered below pH 5.5 using a higher concentration of acetic acid, the survival rate is similarly low as in the class C VPS mutants, however, the death phenotype is predominantly apoptotic. Hence, the vacuole not only prevents acetic acid induced cell death by buffering the cytosolic pH, but it also has a proapoptotic function.