Composition and Acidification of the Culture Medium Influences Chronological Aging Similarly in Vineyard and Laboratory Yeast

Chronological aging has been studied extensively in laboratory yeast by culturing cells into stationary phase in synthetic complete medium with 2% glucose as the carbon source. During this process, acidification of the culture medium occurs due to secretion of organic acids, including acetic acid, which limits survival of yeast cells. Dietary restriction or buffering the medium to pH 6 prevents acidification and increases chronological life span. Here we set out to determine whether these effects are specific to laboratory-derived yeast by testing the chronological aging properties of the vineyard yeast strain RM11. Similar to the laboratory strain BY4743 and its haploid derivatives, RM11 and its haploid derivatives displayed increased chronological life span from dietary restriction, buffering the pH of the culture medium, or aging in rich medium. RM11 and BY4743 also displayed generally similar aging and growth characteristics when cultured in a variety of different carbon sources. These data support the idea that mechanisms of chronological aging are similar in both the laboratory and vineyard strains

Effects of Vegetation, Corridor Width and Regional Land Use on Early Successional Birds on Powerline Corridors

Powerline rights-of-way (ROWs) often provide habitat for early successional bird species that have suffered long-term population declines in eastern North America. To determine how the abundance of shrubland birds varies with habitat within ROW corridors and with land use patterns surrounding corridors, we ran Poisson regression models on data from 93 plots on ROWs and compared regression coefficients. We also determined nest success rates on a 1-km stretch of ROW. Seven species of shrubland birds were common in powerline corridors. However, the nest success rates for prairie warbler (Dendroica discolor) and field sparrow (Spizella pusilla) were <21%, which is too low to compensate for estimated annual mortality. Some shrubland bird species were more abundant on narrower ROWs or at sites with lower vegetation or particular types of vegetation, indicating that vegetation management could be refined to favor species of high conservation priority. Also, several species were more abundant in ROWs traversing unfragmented forest than those near residential areas or farmland, indicating that corridors in heavily forested regions may provide better habitat for these species. In the area where we monitored nests, brood parasitism by brown-headed cowbirds (Molothrus ater) occurred more frequently close to a residential area. Although ROWs support dense populations of shrubland birds, those in more heavily developed landscapes may constitute sink habitat. ROWs in extensive forests may contribute more to sustaining populations of early successional birds, and thus may be the best targets for habitat management

Modulation of Astrocytic Mitochondrial Function by Dichloroacetate Improves Survival and Motor Performance in Inherited Amyotrophic Lateral Sclerosis

Mitochondrial dysfunction is one of the pathogenic mechanisms that lead to neurodegeneration in Amyotrophic Lateral Sclerosis (ALS). Astrocytes expressing the ALS-linked SOD1G93A mutation display a decreased mitochondrial respiratory capacity associated to phenotypic changes that cause them to induce motor neuron death. Astrocyte-mediated toxicity can be prevented by mitochondria-targeted antioxidants, indicating a critical role of mitochondria in the neurotoxic phenotype. However, it is presently unknown whether drugs currently used to stimulate mitochondrial metabolism can also modulate ALS progression. Here, we tested the disease-modifying effect of dichloroacetate (DCA), an orphan drug that improves the functional status of mitochondria through the stimulation of the pyruvate dehydrogenase complex activity (PDH). Applied to astrocyte cultures isolated from rats expressing the SOD1G93A mutation, DCA reduced phosphorylation of PDH and improved mitochondrial coupling as expressed by the respiratory control ratio (RCR). Notably, DCA completely prevented the toxicity of SOD1G93A astrocytes to motor neurons in coculture conditions. Chronic administration of DCA (500 mg/L) in the drinking water of mice expressing the SOD1G93A mutation increased survival by 2 weeks compared to untreated mice. Systemic DCA also normalized the reduced RCR value measured in lumbar spinal cord tissue of diseased SOD1G93A mice. A remarkable effect of DCA was the improvement of grip strength performance at the end stage of the disease, which correlated with a recovery of the neuromuscular junction area in extensor digitorum longus muscles. Systemic DCA also decreased astrocyte reactivity and prevented motor neuron loss in SOD1G93A mice. Taken together, our results indicate that improvement of the mitochondrial redox status by DCA leads to a disease-modifying effect, further supporting the therapeutic potential of mitochondria-targeted drugs in ALS

Replicative Age Induces Mitotic Recombination in the Ribosomal RNA Gene Cluster of Saccharomyces cerevisiae

Somatic mutations contribute to the development of age-associated disease. In earlier work, we found that, at high frequency, aging Saccharomyces cerevisiae diploid cells produce daughters without mitochondrial DNA, leading to loss of respiration competence and increased loss of heterozygosity (LOH) in the nuclear genome. Here we used the recently developed Mother Enrichment Program to ask whether aging cells that maintain the ability to produce respiration-competent daughters also experience increased genomic instability. We discovered that this population exhibits a distinct genomic instability phenotype that primarily affects the repeated ribosomal RNA gene array (rDNA array). As diploid cells passed their median replicative life span, recombination rates between rDNA arrays on homologous chromosomes progressively increased, resulting in mutational events that generated LOH at >300 contiguous open reading frames on the right arm of chromosome XII. We show that, while these recombination events were dependent on the replication fork block protein Fob1, the aging process that underlies this phenotype is Fob1-independent. Furthermore, we provide evidence that this aging process is not driven by mechanisms that modulate rDNA recombination in young cells, including loss of cohesion within the rDNA array or loss of Sir2 function. Instead, we suggest that the age-associated increase in rDNA recombination is a response to increasing DNA replication stress generated in aging cells

Androgen Receptor Drives Cellular Senescence

The accepted androgen receptor (AR) role is to promote proliferation and survival of prostate epithelium and thus prostate cancer progression. While growth-inhibitory, tumor-suppressive AR effects have also been documented, the underlying mechanisms are poorly understood. Here, we for the first time link AR anti-cancer action with cell senescence in vitro and in vivo. First, AR-driven senescence was p53-independent. Instead, AR induced p21, which subsequently reduced ΔN isoform of p63. Second, AR activation increased reactive oxygen species (ROS) and thereby suppressed Rb phosphorylation. Both pathways were critical for senescence as was proven by p21 and Rb knock-down and by quenching ROS with N-Acetyl cysteine and p63 silencing also mimicked AR-induced senescence. The two pathways engaged in a cross-talk, likely via PML tumor suppressor, whose localization to senescence-associated chromatin foci was increased by AR activation. All these pathways contributed to growth arrest, which resolved in senescence due to concomitant lack of p53 and high mTOR activity. This is the first demonstration of senescence response caused by a nuclear hormone receptor

Peroxiredoxin 3 Is a Redox-Dependent Target of Thiostrepton in Malignant Mesothelioma Cells

Thiostrepton (TS) is a thiazole antibiotic that inhibits expression of FOXM1, an oncogenic transcription factor required for cell cycle progression and resistance to oncogene-induced oxidative stress. The mechanism of action of TS is unclear and strategies that enhance TS activity will improve its therapeutic potential. Analysis of human tumor specimens showed FOXM1 is broadly expressed in malignant mesothelioma (MM), an intractable tumor associated with asbestos exposure. The mechanism of action of TS was investigated in a cell culture model of human MM. As for other tumor cell types, TS inhibited expression of FOXM1 in MM cells in a dose-dependent manner. Suppression of FOXM1 expression and coincidental activation of ERK1/2 by TS were abrogated by pre-incubation of cells with the antioxidant N-acetyl-L-cysteine (NAC), indicating its mechanism of action in MM cells is redox-dependent. Examination of the mitochondrial thioredoxin reductase 2 (TR2)-thioredoxin 2 (TRX2)-peroxiredoxin 3 (PRX3) antioxidant network revealed that TS modifies the electrophoretic mobility of PRX3. Incubation of recombinant human PRX3 with TS in vitro also resulted in PRX3 with altered electrophoretic mobility. The cellular and recombinant species of modified PRX3 were resistant to dithiothreitol and SDS and suppressed by NAC, indicating that TS covalently adducts cysteine residues in PRX3. Reduction of endogenous mitochondrial TRX2 levels by the cationic triphenylmethane gentian violet (GV) promoted modification of PRX3 by TS and significantly enhanced its cytotoxic activity. Our results indicate TS covalently adducts PRX3, thereby disabling a major mitochondrial antioxidant network that counters chronic mitochondrial oxidative stress. Redox-active compounds like GV that modify the TR2/TRX2 network may significantly enhance the efficacy of TS, thereby providing a combinatorial approach for exploiting redox-dependent perturbations in mitochondrial function as a therapeutic approach in mesothelioma

Gis1 and Rph1 Regulate Glycerol and Acetate Metabolism in Glucose Depleted Yeast Cells

Aging in organisms as diverse as yeast, nematodes, and mammals is delayed by caloric restriction, an effect mediated by the nutrient sensing TOR, RAS/cAMP, and AKT/Sch9 pathways. The transcription factor Gis1 functions downstream of these pathways in extending the lifespan of nutrient restricted yeast cells, but the mechanisms involved are still poorly understood. We have used gene expression microarrays to study the targets of Gis1 and the related protein Rph1 in different growth phases. Our results show that Gis1 and Rph1 act both as repressors and activators, on overlapping sets of genes as well as on distinct targets. Interestingly, both the activities and the target specificities of Gis1 and Rph1 depend on the growth phase. Thus, both proteins are associated with repression during exponential growth, targeting genes with STRE or PDS motifs in their promoters. After the diauxic shift, both become involved in activation, with Gis1 acting primarily on genes with PDS motifs, and Rph1 on genes with STRE motifs. Significantly, Gis1 and Rph1 control a number of genes involved in acetate and glycerol formation, metabolites that have been implicated in aging. Furthermore, several genes involved in acetyl-CoA metabolism are downregulated by Gis1

Could STAT3 provide a link between respiration and cell cycle progression?

Maintaining the intracellular environment is important for the survival and proliferation of eukaryotic cells. How a cell regulates its redox potential during fluctuations in the generation of intracellular reactive oxygen species and exposure to extracellular oxidants is unclear. The recent findings that Signal Transducer and A ctivator of Transcription 3 (STAT3) regulates mitochondrial respiration and can be oxidized directly by peroxide to form multimers may have revealed features of a homeostatic mechanism coupling cell cycle progression to the intracellular redox potential