From Feast to Famine: Regulating carbon metabolism in yeast

葡萄糖是大多数生物体支持生命的首选碳源。酿酒酵母将葡萄糖转化为乙醇和二氧化碳以产生能量。当葡萄糖限制或缺乏时会发生什么？细胞适应。当葡萄糖耗尽时，酵母可以使用乙醇作为替代碳源，从而使细胞存活。然而，乙醇浓度的增加会对细胞过程产生负面影响，并可能对细胞产生毒性。在葡萄酒、啤酒和生物乙醇生产过程中进行商业上重要的酒精发酵的酵母通常会经历葡萄糖消耗和乙醇水平的增加。这些条件影响酵母的生理和代谢，最终降低其发酵效率。了解酵母对其环境中葡萄糖消耗和乙醇的反应将为碳代谢调节的基本过程提供新的见解，并对乙醇生产行业有潜在的好处。在葡萄糖丰富的环境中，发酵生长所需的酵母基因被积极表达，而那些在没有葡萄糖的情况下使用不可发酵碳源（如乙醇）生长所需的基因则被抑制；这被称为葡萄糖抑制。当葡萄糖消耗殆尽，乙醇是唯一的碳源时，这种转录调节就会逆转。同样，蛋白质丰度、活性和细胞内运输也可以由普遍的碳条件调节。这些适应性转录和/或蛋白质调节反应是由信号机制控制的。尽管已经确定了几种调节蛋白和调节酵母适应葡萄糖消耗的机制，但这些调节机制的几个重要方面仍未得到解决。我们将以适应性转录和蛋白质调控为例，研究新的调节蛋白在碳信号传导中的功能。Vid30蛋白复合物（Vid30c）是一种E3泛素连接酶，已知将小分子泛素连接到靶蛋白，最终导致靶蛋白降解。我们最近证明了Vid30c对于葡萄糖耗竭的分子适应是必需的。除了已知参与己糖转运体Hxt3的降解外，当乙醇是唯一的碳源时，葡萄糖抑制基因的转录激活也需要Vid30c。Vid30c在乙醇中介导这些特定事件的分子和调控机制目前尚不清楚。本研究的目的是描述酵母适应乙醇过程中Vid30c的功能机制。我们将结合酵母遗传学、分子生物学和细胞生物学的方法来鉴定和表征特定的蛋白质，包括Vid30c的调节蛋白和直接靶点，这些蛋白介导了葡萄糖抑制基因的Vid30c依赖性激活和对乙醇的适应过程中Hxt3的降解。此外，我们将确定确定乙醇中Vid30c功能特异性的特定蛋白质。提出的研究将提供新的基本见解，以碳调控方面的酵母，目前知之甚少。解决酵母对葡萄糖消耗和增加乙醇条件的分子适应可以导致提高这种商业上重要生物的发酵效率的策略的发展。

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.