CAREER: A Toolkit to Modulate the Mitochondrial Calcium Uptake Machinery

线粒体是细胞的重要组成部分，因为它们产生细胞所需的大部分化学能，并影响细胞的生存和死亡途径。钙离子进出线粒体对这些细胞成分的功能至关重要；相反，线粒体钙水平的失调与几种人类疾病有关。了解钙离子如何影响线粒体的功能取决于有适当的工具来选择性地减少线粒体对钙的摄取。有了这个奖项，化学部的生命过程化学项目资助康奈尔大学的Justin Wilson教授开发基于线粒体钙摄取小分子调节剂的工具。这些工具对于研究钙在生物学和人类健康和疾病中的重要作用的更大的研究团体是有价值的。在威尔逊教授的实验室进行研究的研究生获得了生物无机化学的知识，并掌握了合成化学和细胞生物学的技能，为他们成为下一代科学家做准备。威尔逊教授还致力于加强农村社区的STEM教育，为K-12 STEM教师组织外展活动和讲习班。本研究项目开发线粒体钙单转运蛋白（MCU）的抑制剂，这是线粒体中唯一已知的钙转运蛋白。MCU于2011年被最终确定。它在调节线粒体钙水平的生物学效应中的确切作用仍不确定。努力阐明MCU在介导细胞过程中的作用将通过在一系列生物条件和设置中操作的选择性MCU调节器工具包得到加强。唯一已知的MCU选择性抑制剂是双核氧桥钌配合物Ru360。然而，这种化合物的实验使用受到几个因素的限制。该复合物仅对研究分离的线粒体或通透细胞有用，因为它不能穿透细胞；分离的线粒体或通透化细胞太简单，无法将结果和结论外推到体内系统。此外，Ru360的作用机制尚不清楚，这阻碍了改进类似物的设计。Wilson教授基于Ru360开发MCU抑制剂。这些抑制剂的开发首先是基于对Ru360功能机制的深入了解。通过核磁共振和EPR光谱研究了该化合物与mcu抑制活性相关的生物分子的相互作用。从机理研究中获得的见解被应用于新型钌基MCU抑制剂的设计，该抑制剂具有增强的细胞渗透性和水稳定性。这些新的抑制剂在活的哺乳动物细胞中进行了评估，以验证它们的mcu抑制活性在复杂的生物系统中保留。这些新工具可用于阐明MCU在介导生理和病理生理细胞过程中的作用和重要性，从而更深入地了解钙的基本细胞生物学及其对人类健康的重要性。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Mitochondria are critical components of a cell because they produce most of the chemical energy the cell needs and influence the cell survival and death pathways. Calcium ion entry in, and exit from, mitochondria is crucial for the function of these cellular components; conversely, deregulation of mitochondrial calcium levels was implicated in several human diseases. The understanding of how the calcium ions influence the function of the mitochondria depends on having appropriate tools to selectively decrease the uptake of calcium by mitochondria. With this award, the Chemistry of Life Processes Program in the Chemistry Division is funding Professor Justin Wilson from Cornell University to develop such tools based on small-molecule regulators of mitochondrial calcium uptake. These tools are valuable to the larger research community that studies the important roles of calcium in biology and in human health and disease. The graduate students who conduct research in Professor Wilson's lab gain knowledge of bioinorganic chemistry and acquire skills in synthetic chemistry and cell biology that prepare them to become members of the next generation of scientists. Professor Wilson also works on enhancing STEM education in rural communities by organizing outreach activities and workshops for K-12 STEM teachers. This research project develops inhibitors of the mitochondrial calcium uniporter (MCU), which is the only known calcium transporter protein in mitochondria. The MCU was conclusively identified in 2011. Its precise role in modulating the biological effects of mitochondrial calcium levels remains uncertain. Efforts to elucidate the roles of the MCU in mediating cellular processes would be strengthened by a toolkit of selective MCU modulators that operate in a range of biological conditions and settings. The only known selective inhibitor for the MCU is the dinuclear oxo-bridged ruthenium complex Ru360. However, the experimental use of this compound is limited by several factors. The complex is useful only for studying isolated mitochondria or permeabilized cells because it cannot penetrate into cells; isolated mitochondria or permeabilized cells are too simple to afford the extrapolation of results and conclusions to in vivo systems. Furthermore, the mechanism of action of Ru360 is unknown, which prevents the design of improved analogues. Professor Wilson develops MCU inhibitors based on Ru360. The development of these inhibitors is first based on gaining mechanistic insight with respect to how Ru360 functions. The interactions of this compound with biomolecules relevant to its MCU-inhibitory activity is studied by NMR and EPR spectroscopy. The insight gained from the mechanistic studies is applied in the design of new ruthenium-based MCU inhibitors with enhanced cell permeability and aqueous stability. These new inhibitors are evaluated in living mammalian cells to verify that their MCU-inhibitory activity is retained in complex biological systems. The new tools may be used to elucidate the role and importance of the MCU in mediating physiological and pathophysiological cellular processes, thereby leading to deeper insight into the fundamental cell biology of calcium and its importance in human health.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.