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首页> 外文学位 >Genetic and functional genomic approaches to characterize regulatory proteins in Saccharomyces cerevisiae.
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Genetic and functional genomic approaches to characterize regulatory proteins in Saccharomyces cerevisiae.

机译：遗传和功能基因组学方法来表征酿酒酵母中的调节蛋白。

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摘要

BAR domain proteins participate in a variety of membrane trafficking processes that require membrane deformation, particularly in endocytosis. In yeast, endocytosis is achieved by four distinct protein modules: the coat, WASP/MYO, actin and scission modules. The scission step requires the combined action of the BAR domain proteins, Rvs161 and Rvs167, the phosphoinositide phosphatase, Inp52, and the pulling force of actin polymerization to generate sufficient interfacial force to culminate in scission. I took a structure-motivated mutational approach to dissect the function of the BAR domains in Rvs161 and Rvs167 in vivo and validate the significance of Rvs-membrane interaction during the endocytosis process. I find that residues important for lipid binding and bending are required for the initial recruitment of Rvs to endocytic sites and for their function in vesicle scission.;Kinases and transcription factors (TFs) are key modulators of important signaling pathways. A systematic assessment of genetic interactions involving the combined perturbation of kinases and TFs may elucidate new aspects of the complex kinase-TF regulatory network. I constructed a collection of yeast strains, each of which contains a chromosomally-integrated kinase or TF gene that can be inducibly overexpressed, and used a variety of assays to characterize the phenotypic consequences of their overexpression. To identify novel genetic interactions, I used Synthetic Genetic Array (SGA) analysis to systematically overexpress 239 different TFs in 13 kinase mutant backgrounds. I assessed dosage-dependent interactions between a loss-of-function or a gain-of-function allele of kinase and TF overexpression allele, and obtained a genetic network consisting of 111 confirmed interactions between 72 TF overexpression alleles and 9 kinase mutants. Several of these genetic interactions were further characterized to delineate regulatory relationships between the kinase and TF, suggesting that genetic interactions between kinase and TF overexpression alleles will be a rich source of new functional information.
机译：BAR结构域蛋白参与各种需要膜变形的膜运输过程，尤其是在胞吞作用中。在酵母中，内吞作用是通过四个不同的蛋白质模块实现的：外壳，WASP / MYO，肌动蛋白和分裂模块。分裂步骤需要BAR结构域蛋白Rvs161和Rvs167，磷酸肌醇磷酸酶Inp52和肌动蛋白聚合的拉力共同作用以产生足够的界面力最终达到分裂。我采取了一种以结构为动机的突变方法，在体内解剖了Rvs161和Rvs167中BAR结构域的功能，并验证了内吞过程中Rvs-膜相互作用的重要性。我发现对于Rvs最初募集到内吞位点及其在囊泡切开中的功能而言，对于脂质结合和弯曲很重要的残基是必需的。激酶和转录因子（TFs）是重要信号通路的关键调节剂。对涉及激酶和TFs的扰动的遗传相互作用的系统评估可能阐明了复杂的激酶-TF调控网络的新方面。我构建了一个酵母菌株的集合，每个酵母菌株都包含一个可以诱导型过表达的染色体整合激酶或TF基因，并使用了多种测定方法来表征它们过表达的表型后果。为了确定新的遗传相互作用，我使用合成遗传阵列（SGA）分析在13种激酶突变体背景下系统性地过表达239种不同的TF。我评估了激酶功能丧失或功能获得等位基因与TF过表达等位基因之间的剂量依赖性相互作用，并获得了由72个TF过表达等位基因与9个激酶突变体之间的111种相互作用确定的遗传网络。这些遗传相互作用中的一些被进一步表征为描绘激酶和TF之间的调节关系，这表明激酶和TF过表达等位基因之间的遗传相互作用将成为新功能信息的丰富来源。

著录项

  • 作者

    Youn, Ji-Young.;

  • 作者单位

    University of Toronto (Canada).;

  • 授予单位 University of Toronto (Canada).;
  • 学科 Genetics.
  • 学位 Ph.D.
  • 年度 2013
  • 页码 212 p.
  • 总页数 212
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类
  • 关键词

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