Illuminating cellular dark matter through the development of novel chemical tools

通过开发新型化学工具照亮细胞暗物质

• 批准号: 10581981
• 负责人: Neal Krishna Devaraj
• 金额: $ 8.98万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10581981
• 关键词: Affect; Amyotrophic Lateral Sclerosis; Biological; Biology; C9ORF72; Cell physiology; Cells; Cellular biology; Chemicals; Development; Disease; Genetic; Goals; Image; In Situ; Knowledge; Laboratories; Lead; Lipids; Modification; Molecular; Nucleotides; Pathogenicity; Pathway interactions; Permeability; Play; Proteins; RNA; Research; Role; Site; Sphingolipids; Technology; Therapeutic Intervention; Vision; cell behavior; cytotoxic; dark matter; human disease; improved; interest; novel; programs; protein complex; recruit; success; tool

PROJECT SUMMARY There is a vast repertoire of species within cells for which we have a poor understanding of their function and biomolecular interactions. These species can be referred to as the “dark matter” of biology, as their mechanism of action is hidden from conventional observation. Our laboratory seeks to illuminate the function of cellular “dark matter” through the development of new chemical technology. The proposed research program will pursue two major research thrusts. First, we plan to develop tools for site-specific RNA modification, and apply these tools for the manipulation, imaging, and isolation of disease relevant RNA protein complexes. We will create tools for use in live cells and develop the ability to covalently recruit proteins to RNA, forming RNA-protein macromolecular conjugates. The technology will be applied to study RNAs implicated in disease. Specifically, we are interested in characterizing the pathways of pathogenicity for the C9orf72 nucleotide repeat expansion RNA, which is thought to play a major role in genetic amyotrophic lateral sclerosis (ALS). In the second thrust, we will carry out the in situ synthesis of lipid species within living cells, with the goal of uncovering the molecular mechanism by which enigmatic lipid species affect cell behavior. We plan to develop approaches enabling the selective and bioorthogonal delivery of sphingolipids to living cells. Building upon technology previously developed in our lab, we will deliver cell permeable lipid precursors which will spontaneously assemble into functional lipids within the cell. Leveraging this approach, we will create photoaffinity probes for the pulldown of sphingolipid-interacting proteins, with the goal of elucidating the protein partners of the non-canonical deoxysphingolipid 1-deoxydihydroceramide, which is cytotoxic and implicated in several diseases. Realization of our research program goals would improve our knowledge of cell biology and lead to the development of new tools for interrogating RNA and lipid species. Our long-term vision is to create and apply technology that enables improved mechanistic understanding of biomolecular interactions, leading to an increased understanding of human disease, and accelerating the development of possible therapeutic interventions.

项目概要 细胞内有大量的物种，我们对它们的功能了解甚少， 这些物种可以被称为生物学的“暗物质”，因为它们的机制。 我们的实验室试图阐明细胞“黑暗”的功能。 拟议的研究计划将通过开发新的化学技术来实现两项目标。 首先，我们计划开发用于位点特异性RNA修饰的工具，并应用这些工具。 我们将创建用于操作、成像和分离疾病相关 RNA 蛋白复合物的工具。 在活细胞中使用并开发将蛋白质共价招募至 RNA 的能力，形成 RNA-蛋白质 该技术将用于研究与疾病有关的 RNA。 我们感兴趣的是表征 C9orf72 核苷酸重复扩增的致病性途径 RNA被认为在遗传性肌萎缩侧索硬化症（ALS）中发挥着重要作用。 我们将在活细胞内进行脂质种类的原位合成，目的是揭示分子 我们计划开发一些方法来实现神秘的脂质物种影响细胞行为的机制。 基于先前的技术，选择性和生物正交地将鞘脂递送至活细胞。 我们的实验室开发的，我们将提供细胞可渗透的脂质前体，这些前体将自发地组装成 利用这种方法，我们将创建用于下拉的光亲和探针。 鞘脂相互作用蛋白，目的是阐明非经典的蛋白质伴侣 脱氧鞘脂 1-脱氧二氢神经酰胺，具有细胞毒性，与多种疾病有关。 我们的研究计划目标将提高我们对细胞生物学的了解，并导致新细胞的开发 我们的长期愿景是创造和应用能够实现 RNA 和脂质种类的技术。 提高对生物分子相互作用的机制理解，从而加深对 人类疾病，并加速开发可能的治疗干预措施。