Combinatorial Microscopies for Super-Resolution Imaging of Molecular and Cellular Dynamics

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Understanding and ultimately controlling how molecules assemble into functional structures is critical in fields ranging from materials science and chemistry to structural and cell biology. We have a long-standing interest in understanding the fundamental mechanisms of self-assembly, ranging from the self-association of dye molecules into excitonic structures, to protein and peptide aggregation. Our most recent work has focused on the dynamics of molecular assembly in live cells and specifically how the hetero- and homo-geneous association of membrane receptors impacts cell signaling and cell-cell interactions. Determining the structural and conformational requirements for these complex assembly processes, and the dynamics thereof, is best accomplished by examining these phenomena on the molecular scale, in real-time, and under nominally real-world conditions. We are focused on the design and optimization of combinatorial microscopies for the direct interrogation of molecular and biomolecular self-assembly. ***Our technical objectives are to implement:*** Hyperspectral super-resolution 3-D optical microscopy for tracking sub-cellular dynamics, organelle remodeling, and protein assembly in live cells;*** Super-resolution light sheet microscopy for probing cell-cell and tissue dynamics and structures with sub-micron spatial resolution and high temporal resolution;***These platforms, and other systems in our lab, have unique capabilities for assessing the role of orientation, conformation, and structure on self-assembly over several orders of magnitude in both length and time. Previously, they have allowed us to identify the critical dimensions associated with exciton generation in two-dimensional molecular solids, and determine the spatial, conformational, and dynamic heterogeneity of membrane receptors in live cells. In the present proposal, the unique super-resolution capabilities of the platforms will allow us to (1) assess how protein and peptide sequence alter the dynamics of membrane association and disruption; (2) interrogate in situ complex formation during mitochondrial membrane remodeling; (3) track cytoskeletal protein self-assembly and organization;***This highly interdisciplinary research program embraces photonics, biophysics, chemistry, cell and molecular biology, and engineering. HQP are directly involved in the design, optimization, implementation, and modification of unique and innovative imaging platforms, with a specific focus on open hardware and software design. By being fully involved in the application of these imaging platforms to complex biological questions and phenomena, HQP are not simply "using" black box imaging strategies but  are afforded a unique and powerful opportunity to be fully engaged in all aspects of the research. **