Deciphering fundamental biological processes involving protein-nucleic acid interactions at the molecular level

破译涉及分子水平上蛋白质-核酸相互作用的基本生物过程

• 批准号: 10622948
• 负责人: Maria Schumacher
• 金额: $ 26.84万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-01-01 至 2027-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10622948
• 关键词: Address; Anti-Bacterial Agents; Antimicrobial Resistance; Bacteria; Biochemistry; Biological; Biological Process; Cessation of life; Complex; Cryoelectron Microscopy; Cues; DNA; DNA Repair Pathway; Development; Enzymes; Genetic Transcription; Glutamate-Ammonia Ligase; Goals; Gram-Positive Bacteria; Health; Human; Investigation; Life Cycle Stages; Link; Metabolic Pathway; Microbe; Mitochondria; Molecular; Multi-Drug Resistance; Nitrogen; Nucleic Acids; Nutrient availability; Pathogenesis; Pharmaceutical Preparations; Process; Proteins; Protozoa; RNA Editing; Second Messenger Systems; Signal Pathway; Signal Transduction; Source; Streptomyces; Structure; Therapeutic; antimicrobial drug; bacterial resistance; combat; drug resistant microorganism; environmental change; in vivo; interest; microbial; microbial disease; novel; phosphoric diester hydrolase; rational design; therapeutic target

ABSTRACT The central goal of the Schumacher lab is to deduce molecular principles governing fundamental biological processes involving protein-nucleic acid interactions. These investigations focus on processes in microbes and intersect with the lab’s interests in microbial pathogenesis. Indeed, while the main goal is to elucidate biological mechanisms at the atomic level, these studies also provide potential targets for the development of urgently needed antimicrobial agents. Alarmingly, recent estimates suggest that deaths from antimicrobial resistance bacteria may exceed 10 million deaths worldwide by 2050 if steps are not taken to generate new treatments. The specific processes we investigate include transcription, DNA organization and RNA editing. Bacteria must be able to sense and respond to environmental changes for their survival and in some cases, proper development, so our studies on transcription focus on important networks and address how environmental cues are signaled and detected by transcription switches. Streptomyces bacteria represent the main source of antibacterial and other key drugs, which they generate concomitant with development. Thus, understanding their developmental lifecycle has been of significant interest for decades, although it is a mystery what drives this process. Our studies in the last few years have revealed that this developmental switch is controlled by the second messenger, c-di-GMP, functioning through two global transcription regulators, BldD and WhiG. These regulators control the first and second steps in Streptomyces development, respectively, but how c-di-GMP levels are sensed and signaled to these regulators are unknown and is a question we will address in this proposal. Initial studies unveiled a possible link between WhiG and a c-di-GMP phosphodiesterase, possibly indicating colocalization as a mechanism to control the second developmental step, which we will investigate. Studies will also be performed to analyze c-di-GMP levels and identify and characterize additional c-di-GMP modulated developmental regulators. Using a combination of cryo-EM, biochemistry and in vivo studies, we will also dissect the molecular mechanism by which nitrogen levels are sensed in Gram-positive bacteria by the novel Glutamine Synthetase-GlnR signaling pathway whereby the central enzyme for a metabolic pathway (GS) directly transduces nutrient availability to its master transcription regulator (GlnR). Finally, we will elucidate the signal and mechanism behind the first SOS-independent DNA repair pathway in bacteria. Another focus of the lab is the unusual RNA editing process in the mitochondria of kinetoplastid parasitic protozoans called kinetoplastid RNA (kRNA) editing. A recently identified accessory complex, the MRB1 complex, is required for this process. However, the structure and mechanisms of action of this complex are completely unknown. We will obtain structures of this complex and dissect its various molecular functions in editing. These combined studies will elucidate fundamental biological processes at the molecular level, leading to the discovery of potential chemotherapeutic targets against microbial diseases.

抽象的 舒马赫实验室的中心目标是推导出控制基本生物学的分子原理 涉及蛋白质-核酸相互作用的过程。这些研究重点关注微生物和 与实验室对微生物发病机制的兴趣相交叉。事实上，虽然主要目标是阐明生物学 除了在原子水平上的机制之外，这些研究还为紧急开发提供了潜在的目标 需要抗菌剂。令人担忧的是，最近的估计表明，抗生素耐药性导致的死亡 如果不采取措施开发新的治疗方法，到 2050 年，全球因细菌死亡的人数可能会超过 1000 万人。 我们研究的具体过程包括转录、DNA 组织和 RNA 编辑。细菌必须 能够感知并响应生存所需的环境变化，并且在某些情况下，能够适当地 的发展，因此我们对转录的研究重点关注重要网络并解决环境如何 线索由转录开关发出信号并检测。链霉菌是细菌的主要来源 抗菌药物和其他关键药物，它们在发育过程中产生。因此，了解 几十年来，它们的发育生命周期一直引起人们的极大兴趣，尽管驱动它们的因素仍然是个谜。 这个过程。我们过去几年的研究表明，这种发育开关是由 第二信使 c-di-GMP，通过两个全局转录调节因子 BldD 和 WhiG 发挥作用。这些 监管机构分别控制链霉菌发育的第一步和第二步，但 c-di-GMP 如何控制 感知到的水平并向这些监管机构发出信号是未知的，这是我们将在本文中解决的一个问题 提议。初步研究揭示了 WhiG 和 c-di-GMP 磷酸二酯酶之间可能存在的联系 表明共定位是控制第二个发育步骤的机制，我们将对此进行研究。 还将进行研究来分析 c-di-GMP 水平并识别和表征其他 c-di-GMP 调节发育调节因子。结合冷冻电镜、生物化学和体内研究，我们将 还剖析了革兰氏阳性菌中氮水平感知的分子机制 新型谷氨酰胺合成酶-GlnR 信号通路，是代谢通路的中心酶 (GS) 直接将营养物质可用性转导至其主转录调节因子 (GlnR)。最后，我们将 阐明细菌中第一个不依赖于 SOS 的 DNA 修复途径背后的信号和机制。 该实验室的另一个重点是动质体寄生线粒体中不寻常的 RNA 编辑过程 原生动物称为动质体 RNA (kRNA) 编辑。最近发现的一个辅助复合物，MRB1 复杂，是这个过程所必需的。然而，该复合物的结构和作用机制是 完全未知。我们将获得该复合物的结构并剖析其各种分子功能 编辑。这些综合研究将在分子水平上阐明基本的生物过程，从而引领 发现针对微生物疾病的潜在化疗靶点。