REGULATION OF YEAST INVERTASE GENE EXPRESSION

葡萄糖(碳分解代谢)抑制是一种全球调控 原核生物中控制多种基因表达的系统 和真核生物。我们的目标是在分子水平上理解 控制全球对葡萄糖反应的机制 可用性。我们选择研究酿酒酵母 (酵母)因为这种小的真核生物具有强大的遗传学。 在以前的工作中，我们已经确定了对 SUC2(转化酶)基因和其他葡萄糖抗性基因的调控 基因。 我们之前已经证明SNF1基因编码一种蛋白质 从葡萄糖抑制中释放所必需的激酶。为了获得 洞察这种蛋白激酶的功能和调节， 对SNF1基因进行体外诱变。生化 建议对SNF1蛋白激酶进行研究以检测其 活性、磷酸化和调节。来鉴定那些 在功能上与SNF1相关，包括编码 蛋白激酶的调控因子和靶标，基因外 SNF1突变的抑制者将被分离出来；特别努力将 旨在恢复等位基因特异性抑制基因 识别与物质相互作用的产物的基因 SNF1蛋白激酶。SNF4和SSN6基因产物为 SNF1蛋白激酶的调节剂或靶标的候选者。 特定的抗血清将被提高，这些基因产物将被 特色化的。对另外两个调控基因的分子分析， SSN1和CID1。基因研究将发现新的 作为我们的证据，表达SUC2所需的基因是有计划的 这表明还有更多的基因有待识别。最后，在 体外诱变将被用来识别负调控 在SUC2上游区域的位点，凝胶延迟分析将 被用来研究SUC2序列与 调节蛋白。

Glucose (carbon catabolite) repression is a global regulatory system that governs the expression of many genes in prokaryotes and eukaryotes. Our goal is to understand at the molecular level the mechanisms controlling the global response to glucose availability. We have chosen to study Saccharomyces cerevisiae (yeast) because of the powerful genetics of this small eukaryote. In previous work we have identified regulatory genes essential for control of the SUC2 (invertase) gene and other glucose-repressible genes. We have previously shown that the SNF1 gene encodes a protein kinase essential for release from glucose repression. To obtain insight into the function and regulation of this protein kinase, the SNF1 gene will be subjected to in vitro mutagenesis. Biochemical studies of the SNF1 protein kinase are proposed to examine its activity, phosphorylation, and regulation. To identify genes that are functionally related to SNF1, including those that encode regulators and targets of the protein kinase, extragenic suppressors of SNF1 mutations will be isolated; special effort will be directed towards recovery of allele-specific suppressors to identify genes with products that interact physically with the SNF1 protein kinase. The SNF4 and SSN6 gene products are candidates for a regulator or target of the SNF1 protein kinase. Specific antisera will be raised and these gene products will be characterized. Molecular analysis of two other regulatory genes, SSN1 and CID1, is proposed. Genetic studies to identify new genes required for SUC2 expression are planned, as our evidence suggests that additional genes remain to be identified. Finally, in vitro mutagenesis will be used to identify negative regulatory sites in the SUC2 upstream region, and gel retardation assays will be used to examine interactions of SUC2 sequences with regulatory proteins.