Metals are associated with a range of degenerative conditions and they pose a serious threat to human health. Considerable evidence indicates that reactive oxygen species (ROS) may play a key role in causing the deleterious biological effects of metals. However, this evidence remains inconclusive and the principal cellular and molecular mechanism(s) underlying metal toxicology have yet to be clearly resolved. This work exploits the experimental advantages of the yeast model Saccharomyces cerevisiae to elucidate cellular metal toxicity, so avoiding the limitations imposed on studies of this nature in animal models. Recent advances in functional genomics technologies that are unique to yeast, in conjunction with results leading up to this proposal from our laboratory, underscore S. cerevisiae as the eukaryotic model of choice here. The objective is to test the hypothesis that oxidative mechanisms are the primary cause of metal toxicity in cells. This will be achieved using two principal strategies developed in our laboratory, each highly distinct from previous approaches to this problem. The work will focus on copper, chromium and cadmium as examples of model redox-active and - inactive toxic metals. First, we will explore in greater depth an entirely novel question that is giving us major new insight to metal toxicity: do proteins that protect against oxidative damage determine the differing metal resistances of individual cells within isogenic populations? We have already discovered that the mechanisms underlying this heterogeneous metal resistance are distinct from those giving rise to culture-averaged resistance (the focus of past studies). Moreover, specific antioxidant functions appear to underpin the heterogeneity, and this key issue will be resolved directly in this project. In addition, an innovative genome-wide search will be carried out to find new determinants of heterogeneity. These heterogeneity studies are vital because, for the first time, they allow true evaluation of the roles of genes-of-interest in situ, i.e., pertaining to intact cells in which expression is not manipulated artificially. Second, we will build on our recent successful use of specific oxidative-damage repair enzymes applied to the problem of metal toxicity. Alongside such approaches, this project will introduce powerful new functional-genomics tools to identify the essential molecular-target(s) of metal action in cells. We will determine directly whether oxidative mechanisms cause the inactivation of these targets, and whole-cell toxicity. By testing the hypothesis in the manners described, the proposed studies should advance significantly our understanding of metal toxicology at the cellular level, and concurrently provide greater insight into the impact of ROS on biological systems. ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM057945-09
Application #
7252692
Study Section
Metallobiochemistry Study Section (BMT)
Program Officer
Fabian, Miles
Project Start
1999-05-01
Project End
2009-06-30
Budget Start
2007-07-01
Budget End
2009-06-30
Support Year
9
Fiscal Year
2007
Total Cost
$204,807
Indirect Cost
Name
University of Nottingham
Department
Type
DUNS #
211389598
City
Nottingham
State
Country
United Kingdom
Zip Code
NG7 2-RD
Alhebshi, Alawiah; Sideri, Theodora C; Holland, Sara L et al. (2012) The essential iron-sulfur protein Rli1 is an important target accounting for inhibition of cell growth by reactive oxygen species. Mol Biol Cell 23:3582-90
Holland, Sara L; Ghosh, Ekalabya; Avery, Simon V (2010) Chromate-induced sulfur starvation and mRNA mistranslation in yeast are linked in a common mechanism of Cr toxicity. Toxicol In Vitro 24:1764-7
Holland, Sara L; Avery, Simon V (2009) Actin-mediated endocytosis limits intracellular Cr accumulation and Cr toxicity during chromate stress. Toxicol Sci 111:437-46
Sideri, Theodora C; Willetts, Sylvia A; Avery, Simon V (2009) Methionine sulphoxide reductases protect iron-sulphur clusters from oxidative inactivation in yeast. Microbiology 155:612-23
Nargund, Amrita M; Avery, Simon V; Houghton, John E (2008) Cadmium induces a heterogeneous and caspase-dependent apoptotic response in Saccharomyces cerevisiae. Apoptosis 13:811-21
Payne, Tom; Hanfrey, Colin; Bishop, Amy L et al. (2008) Transcript-specific translational regulation in the unfolded protein response of Saccharomyces cerevisiae. FEBS Lett 582:503-9
Smith, Matthew C A; Sumner, Edward R; Avery, Simon V (2007) Glutathione and Gts1p drive beneficial variability in the cadmium resistances of individual yeast cells. Mol Microbiol 66:699-712
Holland, Sara; Lodwig, Emma; Sideri, Theodora et al. (2007) Application of the comprehensive set of heterozygous yeast deletion mutants to elucidate the molecular basis of cellular chromium toxicity. Genome Biol 8:R268
Bishop, Amy L; Rab, Faiza A; Sumner, Edward R et al. (2007) Phenotypic heterogeneity can enhance rare-cell survival in 'stress-sensitive'yeast populations. Mol Microbiol 63:507-20
Sumner, Edward R; Shanmuganathan, Anupama; Sideri, Theodora C et al. (2005) Oxidative protein damage causes chromium toxicity in yeast. Microbiology 151:1939-48

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