Sphingolipids and their metabolites are emerging as key second messengers in signal transduction pathways essential for protection against heat, hyperosmolarity, infection, DNA damage, and other stresses. The physiological significance of most sphingolipid second messengers has yet to be established and we have only a rudimentary understanding of how the signals are generated, what physiological processes they regulated, and how their half-lives are controlled. This proposal seeks to determine the role of sphingolipids in the budding yeast heat stress response. The applicants PI hypothesizes that sphingolipid metabolites act as second messengers during heat stress and regulate signal transduction pathways.
Specific Aim 1 is to use HPLC and radio-labeling methods in conjunction with mutant strains and inhibitory drugs to determine which sphingolipid metabolite signals induction of TPS2 expression and accumulation of trehalose, known thermoprotectant.
Specific Aim 2 is to use reporter genes and molecular biological approaches to determine which members of the MSN family of transcription factors respond to sphingolipid signals and activate TPS2 expression.
Specific Aim 3 is to use biochemical and molecular biological methods to determine the function of phytospingosine- resistance genes in heat stress and growth inhibition. One of these genes is a novel MAP kinases and three others are uncharacterized components of the ubiquitination system. They appear to be components of an uncharacterized signal transduction pathway. The results of these experiments will enhance our understanding of the roles sphingolipids play in human responses to heat, hyperosmolarity, infection, ischemia/reperfusion, DNA-damage, and oxidative stress. In addition, these studies will help us to understand how sphingolipid second messengers regulate cell growth, apoptosis, and activation of immune and reticuloendothelial cells. This new knowledge may identify novel targets for pharmacological intervention.
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