CAREER: Molecular mechanisms of pH sensitive proteins, pathways, and cell behaviors

维持细胞内pH值（pHi）在正常的细胞生理学中至关重要，包括代谢、细胞生长和细胞分裂，即使pHi的微小变化也会对细胞过程产生巨大影响。虽然pHi在调节生物学中的作用是明确的，但感知pHi这些微小变化的蛋白质和生物分子在很大程度上是未知的。此外，没有很好的工具来操作和测量单个细胞中的pHi，从而限制了该领域的进展。在这个项目中，PI将确定驱动pH敏感蛋白的分子机制，揭示pH动力学如何驱动单细胞行为，并开发下一代工具来改变活细胞中的pHi。该项目将进一步加深我们对pHi如何调节单细胞生物学的理解，并将揭示蛋白质如何感知和响应pHi的分子机制。操纵细胞pHi的新工具也将适用于其他领域，包括了解pHi如何驱动病理过程。为了这个项目更广泛的影响，PI将开发一个变革性的教育计划，以探索科学研究作为一种创造性的努力。这包括为本科生开发基于项目的学习课程，并在中学艺术课堂上实施基于活动的STEAM（科学、技术、工程、艺术和数学）教育模块。这些模块将培养创新思维，加强跨学科技能，并将科学作为一种创造性的努力建立积极的联系，这已被证明可以鼓励学生学习STEM。人体细胞通常维持在7.0 - 7.2之间的细胞内pH值（pHi），但当pHi值增加到7.6时，可以发出细胞分裂、移动或改变代谢的信号。虽然pHi在调节生物学中的作用是明确的，但感知pHi这些微小变化的蛋白质和生物分子在很大程度上是未知的。因此，该项目将解决ph敏感生物学领域的主要空白。首先，PI将使用最近验证的光遗传学工具来探索时空pH动态如何调节单细胞行为，包括细胞迁移和极性变化。其次，PI将利用钠质子交换剂（NHE1）作为蛋白质工程的基础，以开发一套下一代光遗传学工具，将pHi提高数小时至数天。第三，PI将通过电离残基网络确定ph感应的分子机制，特别是在调节ph敏感生物学的各种含sh2结构域的信号蛋白中。这项工作将揭示pHi动力学如何驱动单细胞生物学，并支持未来研究pH依赖性细胞途径和pH敏感蛋白的基本设计原则的工作。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。

Maintaining intracellular pH (pHi) is vital in normal cellular physiology including metabolism, cellular growth, and cellular division, with even small changes in pHi having a drastic effect on cellular processes. While the role for pHi in regulating biology is clear, the proteins and biomolecules that sense these small changes in pHi are largely unknown. Furthermore, there are no good tools to manipulate and measure pHi in single cells thereby limiting progress in this field. In this project, the PI will determine the molecular mechanisms driving pH sensitive proteins, reveal how pH dynamics drive single-cell behaviors, and develop next-generation tools for changing pHi in living cells. This project will further our understanding of how pHi regulates single-cell biology and will reveal the molecular mechanisms of how proteins sense and respond to pHi. The new tools to manipulate cell pHi will also be applicable to other fields, including understanding how pHi drives pathological processes. For the Broader Impacts of this project, the PI will develop a transformative education program to explore scientific research as a creative endeavor. This includes developing project-based learning curriculum for undergraduate students and implementing activity-based learning modules for STEAM (Science, Technology, Engineering, Art, and Mathematics) education in middle school art classrooms. These modules will cultivate an innovation mindset, strengthen interdisciplinary skills, and build positive associations of science as a creative endeavor, which has been shown to encourage students to pursue STEM.Human cells normally maintain an intracellular pH (pHi) between 7.0 and 7.2 but increases in pHi to ~7.6 can signal cells to divide, to move, or to change their metabolism. While the role for pHi in regulating biology is clear, the proteins and biomolecules that sense these small changes in pHi are largely unknown. Thus, this project will address major gaps in the field of pH-sensitive biology. First, the PI will use recently validated optogenetic tools to probe how spatiotemporal pH dynamics regulate single-cell behaviors including cell migration and polarity changes. Second, the PI will leverage the sodium-proton exchanger (NHE1) as a foundation for protein engineering in order to develop a suite of next-generation optogenetic tools to raise pHi for hours to days. Third, the PI will determine the molecular mechanisms underlying pH-sensing by ionizable residue networks, specifically in various SH2-domain-containing signaling proteins that regulate pH-sensitive biology. This work will reveal how pHi dynamics drive single cell biology as well as support future work investigating pH-dependent cellular pathways and essential design principles of pH sensitive proteins.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.