Intracellular transport and organelle biology at the nanoscale: A multidimensional super-resolution approach

项目摘要 我的研究采用了一种独特的方法， 与细胞生物学的基本发现并行的进展。在方法开发方面， 在实现超分辨率显微镜亚10纳米分辨率的早期成功，我最近的工作， 开创了多维和多功能超分辨率显微镜的概念，其中 细胞内功能参数，包括局部化学极性、pH、扩散率和蛋白质活性， 以纳米分辨率和单分子灵敏度绘制出来。有了这些能力，我的实验室 已经非常成功地揭示了隐藏的亚细胞结构和过程，以及它们的基础。 生物物理规则，从哺乳动物细胞骨架，细胞内运输， 细胞器形态学和生物发生，到膜生物学。我们未来的研究将继续推动 方法开发和生物学发现之间的协同作用。主要方向包括电荷调制 蛋白质的相互作用和对细胞器腔内扩散的影响， 活细胞，细胞器pH动力学和蛋白质运输中的作用，以及 内质网出口位点与转运载体的生物发生有关。此外，通过整合超分辨率 利用纤维束扫描电镜，我们将获得我们最近发现的异常薄的管状细胞器的整体照片。 发现并进一步证实了它们的功能和生物起源。另外，我们正在开发一种新工具， 单分子电泳显微镜，在超分辨率水平上定量蛋白质电荷状态。 总之，通过不断开发增强能力的显微镜工具及其战术应用， 基本的生物学问题，我们将继续改变我们如何理解复杂的范式， 细胞的动态行为。

Project Summary My research takes a unique approach in which the development of next-generation microscopy methods progresses in parallel with fundamental discoveries in cell biology. On the method-development front, besides earlier success in achieving sub-10 nm resolution for super-resolution microscopy, my recent work has pioneered the concept of multidimensional and multifunctional super-resolution microscopy, in which intracellular functional parameters, including local chemical polarity, pH, diffusivity, and protein activity, are mapped out at nanometer resolution and single-molecule sensitivity. Empowered by such capabilities, my lab has been highly successful in unveiling hidden subcellular structures and processes, as well as their underlying biophysical rules, for diverse systems ranging from the mammalian cytoskeleton, intracellular transport, organelle morphology and biogenesis, to membrane biology. Our future research continues to push forward the synergy between method development and biological discoveries. Major directions include charge-modulated protein interactions and effects on diffusion inside the organelle lumen, superdiffusion and subdiffusion in the living cell, organelle pH dynamics and role in protein trafficking, and the structure and physical properties of the ER exit site in relationship with the biogenesis of transport carriers. Moreover, by integrating super-resolution microscopy with FIB-SEM, we will obtain holistic pictures of the unusually thin tubular organelles we recently discovered and further substantiate their functions and biogenesis. Separately, we are developing a new tool, single-molecule electrophoresis microscopy, to quantify protein charge states at the super-resolution level. Together, through the continued development of empowering microscopy tools and their tactical application to fundamental biological questions, we will continue shifting the paradigms of how we understand the complex, dynamic behavior of the cell.