Yeast mother cell-specific ageing, genetic (in)stability, and the somatic mutation theory of ageing

Yeast mother cell-specific ageing is characterized by a limited capacity to produce daughter cells. The replicative lifespan is determined by the number of cell cycles a mother cell has undergone, not by calendar time, and in a population of cells its distribution follows the Gompertz law. Daughter cells reset their clock to zero and enjoy the full lifespan characteristic for the strain. This kind of replicative ageing of a cell population based on asymmetric cell divisions is investigated as a model for the ageing of a stem cell population in higher organisms. The simple fact that the daughter cells can reset their clock to zero precludes the accumulation of chromosomal mutations as the cause of ageing, because semiconservative replication would lead to the same mutations in the daughters. However, nature is more complicated than that because, (i) the very last daughters of old mothers do not reset the clock; and (ii) mutations in mitochondrial DNA could play a role in ageing due to the large copy number in the cell and a possible asymmetric distribution of damaged mitochondrial DNA between mother and daughter cell. Investigation of the loss of heterozygosity in diploid cells at the end of their mother cell-specific lifespan has shown that genomic rearrangements do occur in old mother cells. However, it is not clear if this kind of genomic instability is causative for the ageing process. Damaged material other than DNA, for instance misfolded, oxidized or otherwise damaged proteins, seem to play a major role in ageing, depending on the balance between production and removal through various repair processes, for instance several kinds of proteolysis and autophagy. We are reviewing here the evidence for genetic change and its causality in the mother cell-specific ageing process of yeast

Dynamic rerouting of the carbohydrate flux is key to counteracting oxidative stress

<p>Abstract</p> <p>Background</p> <p>Eukaryotic cells have evolved various response mechanisms to counteract the deleterious consequences of oxidative stress. Among these processes, metabolic alterations seem to play an important role.</p> <p>Results</p> <p>We recently discovered that yeast cells with reduced activity of the key glycolytic enzyme triosephosphate isomerase exhibit an increased resistance to the thiol-oxidizing reagent diamide. Here we show that this phenotype is conserved in <it>Caenorhabditis elegans </it>and that the underlying mechanism is based on a redirection of the metabolic flux from glycolysis to the pentose phosphate pathway, altering the redox equilibrium of the cytoplasmic NADP(H) pool. Remarkably, another key glycolytic enzyme, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), is known to be inactivated in response to various oxidant treatments, and we show that this provokes a similar redirection of the metabolic flux.</p> <p>Conclusion</p> <p>The naturally occurring inactivation of GAPDH functions as a metabolic switch for rerouting the carbohydrate flux to counteract oxidative stress. As a consequence, altering the homoeostasis of cytoplasmic metabolites is a fundamental mechanism for balancing the redox state of eukaryotic cells under stress conditions.</p

Triose Phosphate Isomerase Deficiency Is Caused by Altered Dimerization–Not Catalytic Inactivity–of the Mutant Enzymes

Triosephosphate isomerase (TPI) deficiency is an autosomal recessive disorder caused by various mutations in the gene encoding the key glycolytic enzyme TPI. A drastic decrease in TPI activity and an increased level of its substrate, dihydroxyacetone phosphate, have been measured in unpurified cell extracts of affected individuals. These observations allowed concluding that the different mutations in the TPI alleles result in catalytically inactive enzymes. However, despite a high occurrence of TPI null alleles within several human populations, the frequency of this disorder is exceptionally rare. In order to address this apparent discrepancy, we generated a yeast model allowing us to perform comparative in vivo analyses of the enzymatic and functional properties of the different enzyme variants. We discovered that the majority of these variants exhibit no reduced catalytic activity per se. Instead, we observed, the dimerization behavior of TPI is influenced by the particular mutations investigated, and by the use of a potential alternative translation initiation site in the TPI gene. Additionally, we demonstrated that the overexpression of the most frequent TPI variant, Glu104Asp, which displays altered dimerization features, results in diminished endogenous TPI levels in mammalian cells. Thus, our results reveal that enzyme deregulation attributable to aberrant dimerization of TPI, rather than direct catalytic inactivation of the enzyme, underlies the pathogenesis of TPI deficiency. Finally, we discovered that yeast cells expressing a TPI variant exhibiting reduced catalytic activity are more resistant against oxidative stress caused by the thiol-oxidizing reagent diamide. This observed advantage might serve to explain the high allelic frequency of TPI null alleles detected among human populations

Induction of autophagy by spermidine promotes longevity.

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Structural model of human TPI. (Upper panel)

<div><p>The pathogenic TPI variants Cys41Tyr, Glu104Asp, Gly122Arg, Ile170Val and Phe240Leu were assigned to the crystal structure of human TPI generated by Kinoshita et al <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0000030#pone.0000030-Kinoshita1" target="_blank">[38]</a>; NT: amino terminus. </p> <p> <b>(Lower panel)</b> Appearance of pathogenic TPI variants among TPI deficiency patients (c.h.: compound heterozygous; n.d.: not determined).</p></div

Consequences of mutations within human TPI.

<div><p>Homozygous mutations within TPI affecting enzyme dimerization cause TPI deficiency, whereas homozygous mutations resulting in an inactive <i>TPI</i> allele are lethal.</p> <p>Compound heterozygous individuals having inherited one inactive and one allele defective in dimerization properties will develop TPI deficiency, whereas heterozygote individuals having inherited a heterozygote null allele have an evolutionary advantage. (+: wild-type TPI; −: TPI with aberrant dimerization property; 0: allele encoding no or a catalytically inactive TPI).</p></div

Human wild-type and pathogenic TPI variants can substitute for yeast TPI1.

<div><p> <b>A)</b> MR100 yeast cells (<i>Δtpi1</i>) were transformed with the various p416GPD-based expression plasmids encoding wild-type human TPI as well as the pathogenic variants Met1_AAG, Cys41Tyr, Glu104Asp, Gly122Arg, Ile170Val or Phe240Leu, respectively, and plated on minimal SC ^-leu-ura medium supplemented with 3% ethanol/0.1% glucose.</p> <p>Afterwards, single yeast clones were selected and grown as represented by the schemes on SC ^-leu-ura medium plates supplemented either with 2% glucose or with 3% ethanol/0.1% glucose at 30°C. <b>B)</b> MR100 <i>Δtpi1</i> yeast cells expressing wild-type TPI or the different pathogenic TPI variants were grown until logarithmic phase.</p> <p>Then, the same cell number of each culture was spotted as 5-fold serial dilutions onto glucose media or onto glucose media supplemented with different concentrations of lithium chloride. </p> <p>Plates were incubated for 3 days at 30°C and growth of the different yeast strains was analyzed.</p></div

MR100 Δ<i>tpi1</i> yeast cells expressing the Ile170Val TPI variant are hyperresistant to diamide.

<div><p>MR100 <i>Δtpi1</i> yeast cells expressing wild-type as well as the pathogenic TPI variants Cys41Tyr, Glu104Asp, Gly122Arg, Ile170Val or Phe240Leu were grown to stationary phase, serially diluted to OD₆₀₀ values of 3.0, 1.0, 0.3, 0.1 and spotted onto SC medium plates containing different concentrations of diamide.</p> <p>Sensitivity/resistance was determined by comparing the growth between the different yeast strains after incubating the plates for 3 days at 28°C.</p></div