Molecular Dissection of a Heme Regulatory Complex in the Yeast Saccharomyces Cerevisiae

9617472 Zhang真核细胞需要血红素来利用和感知氧，通过呼吸产生细胞能量，以及合成和降解脂质、类固醇和相关化合物。 除了作为酶、细胞色素和球蛋白中的辅基的作用之外，血红素还直接作为使用或感测氧的细胞和分子系统的调节剂，并且控制哺乳动物中多种细胞的生长和分化。 本研究的目的是在酵母血红素调节复合物的分子解剖。 酵母酿酒酵母是唯一的真核生物，其中血红素信号转导的主调节因子，血红素激活蛋白-1（HAP 1），已被鉴定和克隆。 响应于血红素，HAP 1激活编码呼吸功能和控制氧化损伤所需功能的基因的转录。 在没有血红素的情况下，HAP 1与某些其他细胞因子以高分子量复合物形式被隔离。 本研究将验证血红素与HAP 1结合改变HAP 1构象的假设，使其从高分子量复合物中解离，随后以高亲和力与DNA结合并激活转录。 突变和生化分析，包括血红素响应的体外转录系统的开发，将被用来表征血红素与HAP 1的特异性相互作用，HAP 1与高分子量复合物中其他细胞因子的相互作用，以及HAP 1从高分子量复合物中解离介导血红素转录调节的机制。 通过阐明酵母中血红素-HAP 1诱导途径的调控机制，本研究将首次从分子水平描述调控复合物如何介导血红素在真核细胞中的作用，并将为理解高等真核细胞中血红素对基本分子和细胞过程的调控奠定基础。 真核细胞需要血红素来利用和感知氧，通过呼吸产生细胞能量，以及合成和降解脂质、类固醇和相关化合物。 除了作为酶、细胞色素和球蛋白中的辅基的作用之外，血红素还直接作为使用或感测氧的细胞和分子系统的调节剂，并且控制哺乳动物中多种细胞的生长和分化。通过阐明酵母中血红素的诱导途径，本研究将提供第一个血红素调节复合物如何在真核细胞中介导血红素作用的分子描述，并将为理解高等真核细胞中血红素调节基本分子和细胞过程奠定基础。

9617472 Zhang Eukaryotic cells require heme to use and sense oxygen, to generate cellular energy by respiration, and to synthesize and degrade lipids, steroids and related compounds. In addition to its role as a prosthetic group in enzymes, cytochromes and globins, heme acts directly as a regulator of cellular and molecular systems that use or sense oxygen, and controls the growth and differentiation of a wide array of cells in mammals. The goal of this research is the molecular dissection of the heme regulatory complex in yeast. The yeast Saccharomyces cerevisiae is the only eukaryote in which a master regulator of heme signaling, heme activating protein-1 (HAP1), has been identified and cloned. In response to heme, HAP1 activates transcription of genes encoding respiratory functions and functions required for controlling oxidative damage. In the absence of heme, HAP1 is sequestered in a high molecular weight complex with certain other cellular factors. This research will test the hypothesis that the binding of heme to HAP1 changes the conformation of HAP1 causing it to dissociate from the high molecular weight complex and subsequently bind to DNA with high affinity and activate transcription. Both mutational and biochemical analyses, including the development of a heme-responsive in vitro transcription system, will be used to characterize the specific interaction of heme with HAP1, the interaction of HAP1 with other cellular factors in the high molecular weight complex, and the mechanism by which HAP1 dissociation from the high molecular weight complex mediates heme regulation of transcription. By unraveling the mechanism governing the heme-HAP1 induction pathway in yeast, this research will provide the first molecular description of how a regulatory complex mediates heme action in eukaryotic cells, and will lay the basis for understanding heme regulation of fundamental molecular and cellular processes in higher eukaryotic cells. Eukaryotic cells require heme to use and sense oxy gen, to generate cellular energy by respiration, and to synthesize and degrade lipids, steroids and related compounds. In addition to its role as a prosthetic group in enzymes, cytochromes and globins, heme acts directly as a regulator of cellular and molecular systems that use or sense oxygen, and controls the growth and differentiation of a wide array of cells in mammals. By unraveling the heme induction pathway in yeast, this research will provide the first molecular description of how a heme regulatory complex mediates heme action in eukaryotic cells, and will lay the basis for understanding heme regulation of fundamental molecular and cellular processes in higher eukaryotic cells.