Quantifying Protein Interactions via near-Quantum Optimal Imaging and MINFLUX at kHz Sampling Rates

About This Grant

This research will develop an advanced optical microscopy system with an unprecedented combined spatial and temporal precision that will dramatically improve our ability to study how proteins cluster and interact on the membranes of living cells. This work has the potential to significantly advance our understanding of cell signaling, which plays a crucial role in biological processes such as wound healing and development. Moreover, it will further provide insights into how dysregulation of cell signaling relates to diseases such as cancer. The highly interdisciplinary nature of the project will provide unique opportunities for students’ training across biology, microscopy, and quantum information science. Students will be in a unique research environment at the University of New Mexico (UNM) across multiple departments and will take advantage of many professional and training activities including workshops, seminars, and networking, within and outside UNM. The undergraduate and graduate students involved in this project will participate in the newly established Quantum Photonics and Quantum Technology (QPAQT) graduate program (an NRT program from NSF), and the “Quantum Undergraduate Research Experience'' (QU-REACH), which creates inclusive research experiences in STEM. The project will design and construct a state-of-the-art microscope that combines two recent breakthroughs in imaging techniques: MINFLUX, which allows high-speed (kHz rates) tracking of fluorophore-tagged proteins, and Modal Imaging, which enables near quantum optimal measurements of the separation between two-point emitters. This combined system will allow for the observation of fast protein interactions and dimerization kinetic rates in the full context and complexity of the living cell membrane at time scales not previously reachable through any other method. The system will be applied to investigate dimerization kinetics related to the oligomer induced signaling from cell membranes, in particular the quantification of receptor tyrosine kinases (RTKs). RTKs play critical roles in normal cell processes such as development and wound healing, and dysregulated RTK signaling drives multiple diseases including cancer. The combination of MINFLUX and modal imaging will allow us to probe RTK diffusional dynamics, oligomer formation and dimer kinetics at unprecedented spatiotemporal resolution on living cells. The study will focus on two RTKs: the epidermal growth factor receptor (EGFR), a widely studied RTK providing a well stablished signaling model, and the less-studied RON receptor. Furthermore, this system will enable future studies of other proteins and membrane architectures, such as cytoskeleton corrals and protein condensates, and their effects on oligomerization and signaling. This project was jointly funded by the Molecular Biophysics Cluster in the Division of Molecular and Cellular Biosciences, and the Established Program to Stimulate Competitive Research (EPSCoR). 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.