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.

摘要 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。这些研究集中在微生物的过程， 与实验室对微生物致病机理的兴趣有交集事实上，虽然主要目标是阐明生物学 在原子水平上的机制，这些研究也提供了迫切发展的潜在目标， 需要抗菌剂。令人担忧的是，最近的估计表明， 如果不采取措施产生新的治疗方法，到2050年，全世界死于细菌的人数可能超过1 000万。 我们研究的具体过程包括转录，DNA组织和RNA编辑。细菌必须 能够感知和应对环境变化，以生存，在某些情况下， 因此，我们对转录的研究集中在重要的网络上，并解决环境如何影响转录的问题。 通过转录开关发出信号并检测提示。链霉菌属细菌代表了 抗菌药物和其他关键药物，它们是伴随着发展而产生的。因此，理解 它们的发育生命周期几十年来一直受到人们的极大关注，尽管是什么驱动了它们的发育生命周期还是一个谜。 这个过程我们在过去几年的研究表明，这种发育开关是由 第二信使，c-di-GMP，通过两个全球转录调节因子BldD和WhiG发挥作用。这些 调节剂控制链霉菌发展的第一和第二步，分别，但如何c-di-GMP 水平的感应和信号，这些监管机构是未知的，这是一个问题，我们将解决在这一点上， 提议最初的研究揭示了WhiG和c-di-GMP磷酸二酯酶之间的可能联系， 表明共定位作为一种机制，以控制第二个发展步骤，我们将调查。 还将进行研究，以分析c-di-GMP水平，并识别和表征其他c-di-GMP 调节发育调节因子。使用冷冻EM，生物化学和体内研究的组合，我们将 还剖析了革兰氏阳性菌中氮水平被检测的分子机制， 新的谷氨酰胺合成酶-GlnR信号通路，其中代谢通路的中心酶 (GS)直接将养分有效性转换为其主转录调节因子（GlnR）。最后我们将 阐明细菌中第一个SOS非依赖性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虫线粒体中不寻常的RNA编辑过程。 称为动质体RNA（kRNA）编辑。最近发现的一种附属复合体，MRB 1 这一过程需要复杂。然而，这种复合物的结构和作用机制是不确定的。 完全未知我们将获得这种复合物的结构，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学过程， 发现潜在的针对微生物疾病的化疗靶点。