Aging in yeast

Research project P 14574 Aging in Yeast Michael BREITENBACH 26.6.2000 Yeast mother cell-specific aging was discovered more than 40 years ago and has been intensively investigated during the last five years. The yeast aging model has enormous a4vantages compared to other model organisms for aging research: 1. The yeast cell has been the first eukaryotic cell for which a complete genome: sequence was available. This enables us to use systematic methods of functional genomics, in particular it is now possible to compare transcriptional activities of all 6200 yeast genes in young and senescent yeast cells with well established methods. Since 1999 the complete genome sequence of the nematode, C. elegans, is also known which is a prominent model organism for aging research. This enables us to systematically compare aging-specific genes in the two genomes. 2.Yeast displays the fastest known aging process. The life span of yeast mother cells is typically 20-30 generations or 2-3 days. 3.Yeast methods are extremely well developed with respect to classical genetics, reverse genetics, structural biology, and cell physiology and the body of knowledge in all these fields is enormous. It is easy to manipulate respiration of yeast cells and therefore the production of reactive oxygen species (ROS) through glucose repression, anaerobiosis and the generation of cytoplasmic petite mutations enabling us to study the influence of oxygen toxicity on yeast aging- The aging phenotypes of yeast cells and higher cells are remarkably similar with respect to cell size, cell polarity, surface deformations, a slowing down of protein synthesis and of the cell cycle and the influence of activated alleles of the ras oncogenes, which activate the aging process in both systems. The aims of our project are i) the investigation of the role of ROS in the yeast aging process. In a reoent paper (Nestelbacher et al., Exp. Gerontol. 35, p.63-70, 2000) such a role was suggested based on life span determinations in catalase mutants, in increased oxygen conditions, and in the presence of the antioxidant, glutathione. Additional genetic and biochemical experiments are now planned to show the involvement of ROS in the aging process in a more direct way. ii) We have shown that activated ras mutations accelerate the aging process of yeast cells as they do in primary fibroblast cultures. We are now proposing to analyse in detail extragenic suppressors of the ras mutations which we have isolated. This may lead us back to the cellular ROS detoxification pathways. iii) We will perform a systematic and genome wide comparison of transcription levels in young and senescent cells in at least two genetic backgrounds. This will lead to identification of co-regulated groups of genes that could have functionally to do with the aging process. Some of these genes will be analyzed systematically for phenotypes. Methods developed by us in the EUROFAN program will be used. One of the genes identified previously by differential display techniques could have a role in ROS detoxification.

Presently we are experiencing a world-wide renaissance of aging research that is without doubt caused by the awareness of increasing socio-economic problems related to the dramatic increase in life expectancy and health over the last 50 years and the concomitantly decreasing age of retirement, the decreasing number of children in the industrialized countries, the problems of migration, and the scarcity of jobs for young people. This is an enormous challenge to the EU and to other developed countries. What is needed is a steady state-model for economy (which does not exist), but also an improved understanding of the aging process itself, both biochemically and from the standpoint of social sciences. Researchers realise how little we know about the physiology and biochemistry of aging. It is not even possible to discriminate with certainty between "healthy aging" and the pathology of aging- related diseases. Studies in humans for obvious reasons cannot clarify the most urgent questions of aging research because of the long generation time, the poor genetics in an outbred population and for ethical reasons. Moreover, human cells in culture, as shown by a very large body of evidence, can only address a partial aspect of organismic aging, and so we are left with model systems, of which the mouse, the fly, the worm and also bakers`s yeast, Saccharomyces cerevisiae are the most important ones. The aim of the present project was to produce detailed evidence for the "oxygen theory of aging" which is also called "radical theory of aging" in the yeast model system. Oxygen is not only necessary for respiration and hence for survival, it also produces oxygen radicals and the toxic products derived from them, which must be constantly detoxified in order to protect a healthy cell. Numerous human pathologies are related to oxygen toxicity. As all cells except anaerobes face this problem of oxygen toxicity and as accumulation of oxidative damage has been demonstrated in senescent individuals, oxygen toxicity could be a "public mechanism of aging". It is the study of those public mechanisms of aging (in contrast to the "private" ones) which could promote our understanding of aging in general and which could be applicable to the understanding of human aging. We have shown that senescent yeast mother cells in their aging process create endogenous oxidative stress which is not triggered by outside oxidants and eventually undergo programmed cells death (apoptosis). Further we have shown that activating the RAS/cAMP pathway of yeast through an oncogenic point mutation of the RAS2 gene induces oxidative stress and premature aging in yeast, as it does in human cells. Finally we have analyzed on DNA microarrays which yeast genes are transcriptionally over- or underexpresed in aging cells. An analysis of apoptotic yeast cells resulted in a similar pattern of gene expression, confirming our earlier findings about senescence in yeast.