Analysis of the role of the ubiquitin system in the regulation of yeast carbohydrate metabolism

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Every organism, whether a human or a single cell, has the ability to adapt its metabolism between times of feast and famine. Abundant nutrients enable energy stores to be laid down and later released when times are lean. Cells and organisms efficiently respond to nutrient abundance by altering the operation of their metabolic pathways through the rapid and reversible activation of key enzymes that are turned on or off in response to nutrient signals. One such enzyme in mammals is called AMP-dependent protein kinase (AMPK), that responds to cellular ATP/AMP levels, turning energy consuming metabolic pathways off or on to preserve a constant energy level. Our studies focus on the evolutionarily related yeast AMPK enzyme, SNF1 kinase, which responds similarly to glucose levels, its favorite energy source. This simple yeast system will be used to explore the molecular regulation of this enzyme family. We find that SNF1 kinase has a small protein called ubiquitin (Ub) attached to it, which may affect its stability, cellular location, and activity, all processes that could readily contribute to regulation of this enzyme. Phosphorylation of SNF1 kinase and nuclear localization are required for activity, but the molecular mechanisms controlling these events remain unknown. To shed light on the enzymes and potential triggers that cause attachment of Ub to SNF1 kinase, and the result of that attachment when the glucose supplies change, we will examine changes to SNF1 kinase protein levels, nuclear localization using fluorescent protein tags and microscopy, and metabolic adaptation to different sugar sources when we block SNF1 kinase ubiquitination. Elucidating the role of Ub in SNF1 kinase function may provide insight into how human metabolism is regulated. Inefficient metabolism contributes to metabolic syndrome, encompassing diabetes, obesity, and accelerated heart disease. This is rising in prevalence and affecting increasingly younger people in Canada. Two agents that enhance AMPK activity (exercise and Metformin) improve metabolic efficiency, suggesting that our findings have the potential to greatly benefit human metabolism.