Grant: Discovery Grants Program - Individual

Glucose is the preferred carbon source that supports life in most organisms. The yeast Saccharomyces cerevisiae converts glucose into ethanol and carbon dioxide to generate energy. What happens when glucose becomes limiting or is absent? Cells adapt. Yeast can use ethanol as an alternative carbon source when glucose is depleted, thereby allowing the cell to survive. However, increasing concentrations of ethanol negatively impacts cellular processes and can become toxic to the cell. Yeast performing commercially important alcoholic fermentations during wine, beer and bio-ethanol production commonly experience glucose depletion and increasing levels of ethanol as the fermentation proceeds. These conditions influence the yeast's physiology and metabolism that ultimately decrease its fermentation efficiency. Understanding the response of yeast to glucose depletion and ethanol in its environment will provide novel insight into the fundamental process of carbon metabolism regulation with potential benefits to ethanol-producing industries. In an abundant glucose environment the yeast genes needed for fermentative growth are actively expressed, while those needed for growth in the absence of glucose using a non-fermentable carbon source, like ethanol, are inhibited; this is known as glucose repression. This transcriptional regulation reverses when glucose is depleted and ethanol is the sole carbon source. Similarly, protein abundance, activity and intracellular trafficking can also be regulated by the prevailing carbon conditions. These adaptive transcription and/or protein regulation responses are governed by signaling mechanisms. Despite the identification of several regulatory proteins and mechanisms that govern the adaptation of yeast to glucose depletion, several important aspects of these regulatory mechanisms remain unresolved. We will use examples of adaptive transcription and protein regulation as a means to study the function of new regulatory proteins in carbon signaling. The Vid30 protein complex (Vid30c) is an E3 ubiquitin ligase known to link a small molecule, ubiquitin, to target proteins that ultimately result in the degradation of the target protein. We showed recently that the Vid30c was needed for the molecular adaptation to glucose depletion. In addition to its known involvement in the degradation of the hexose transporter Hxt3, the Vid30c is also needed for the transcriptional activation of glucose repressed genes when ethanol is the sole carbon source. The molecular and regulatory mechanisms used by the Vid30c to mediate these specific events in ethanol are currently unknown. The goal of the proposed research is to delineate the mechanisms of function of the Vid30c during the yeast's adaptation to ethanol. We will use a combination of yeast genetic, molecular and cellular biology approaches to identify and characterize specific proteins, including regulatory proteins and direct targets of the Vid30c, that mediate the Vid30c-dependent activation of glucose-repressed genes and the degradation of Hxt3 during the adaptation to ethanol. In addition, we will identify specific proteins that determine the functional specificity of the Vid30c in ethanol. The proposed research will provide novel fundamental insight into an aspect of carbon regulation in yeast that is currently poorly understood. Resolving the molecular adaptation of yeasts to glucose depletion and increasing ethanol conditions can lead to the development of strategies to increase the fermentation efficiency of this commercially important organism.