Chemical genetic analysis of intracellular Transport

描述（由申请人提供）：细胞内转运是细胞器和大分子复合物在每个真核细胞的细胞质中正确分布和定位所需的基本生物学过程。我们项目的目标是利用化学遗传学研究细胞内转运的分子机制。这种方法模仿经典遗传学作为发现工具，并涉及选择可以干扰细胞功能的化学探针，鉴定这些探针的靶标，然后使用探针来阐明已知或新蛋白质对细胞功能的贡献。我们相信，化学遗传学方法将是特别有效的，在确定和检查脊椎动物细胞内运输的分子机制，其中遗传筛选的应用是困难的，方法干扰蛋白质功能的时间尺度上慢得多的运输过程可以是有限的使用。我们已经确定了一个强大而有效的系统，将允许选择的化合物，无论是对马达蛋白，或蛋白质最直接参与调节马达蛋白功能在细胞内运输。在本工作中，我们将：（1）利用非洲爪蟾黑素细胞系统鉴定细胞内转运的小分子探针。(2)使用二级筛选来选择直接抑制运动蛋白驱动的细胞器运动或靶向与这些运动最密切相关的调节机制的小分子。(3)识别细胞靶点，检查化学遗传探针用于研究细胞内转运的特异性和普遍适用性。我们的研究发现和验证的小分子探针将成为研究细胞内转运机制的有力工具。这些小分子很可能会导致识别治疗剂的新靶点和改善人类疾病的治疗。

DESCRIPTION (provided by applicant): Intracellular transport is a fundamental biological process that is required for the proper distribution and positioning of organelles and macromolecular complexes in the cytoplasm of every eukaryotic cell. The goal of our project is to examine molecular mechanisms of intracellular transport using chemical genetics. This approach emulates classical genetics as a discovery tool, and involves the selection of chemical probes that can perturb a cellular function, identification of targets of these probes, followed by the use of the probes to clarify the contribution of known or novel proteins to the cellular function. We believe that the chemical genetic approach will be particularly effective in identifying and examining molecular mechanisms of vertebrate intracellular transport, where the application of genetic screens is difficult and approaches Perturbing protein function on time-scales much slower than the transport processes can be of limited use. We have identified a robust and efficient system that will allow the selection of compounds that either act on motor proteins, or proteins most directly involved in regulating motor protein function during intracellular transport. In the course of this work we will: (1) Use the Xenopus melanophore system to identify small molecule probes for intracellular transport. (2) Use secondary screens to select small molecules that either directly inhibit motor protein-driven organelle movement or target the regulatory mechanisms most closely associated with these movements. (3) Identify cellular targets, examine specificity and general applicability of chemical genetic probes for studying intracellular transport. The small molecules probes discovered and validated in our studies will be powerful tools to examine mechanisms of intracellular transport. It is likely that these small molecules will lead to the identification of new targets for therapeutic agents and improved treatments of human diseases.