Spatial Regulation of Plasma Membrane Signaling

Cells sense and interact with their surroundings through signaling processes that take place on the plasma membrane. Signals transduced through the membrane are integrated by the cell to make decisions about cellular responses and cell fate. Antigen engagement by immune receptors, for example, can provoke dramatic cellular activation and downstream effector responses, while defects in signaling can lead to autoimmune disease or immunodeficiency. Signal transduction often depends on spatial re-organization of receptors that leads to new interactions with downstream signaling partners. This occurs within the plasma membrane which itself is a complex mixture of proteins and lipids with a heterogeneous and dynamic structure. As a result, receptors and signaling molecules encounter various kinds of membrane microenvironments that can organize, compartmentalize, and locally regulate the biochemical steps of signaling. In T cells, activation through the T cell receptor (TCR) is clearly highly coordinated in space. TCRs engaged with an antigen-presenting surface first form microcluster assemblies that initiate the activation signal. But, key mechanistic details of receptor triggering and regulation are lacking. A major obstacle is that both microclusters and the surrounding heterogenous membrane are structured on sub-micron length scales that are inaccessible to conventional microscopy. My research program aims to understand how receptor signaling complexes are assembled and regulated by the heterogeneous membrane using super-resolution fluorescence imaging. Super-resolution localization microscopy (SMLM) is a powerful tool to study the spatial regulation of membrane receptor signaling because it can measure organization of biomolecules on length scales that are relevant to both signaling assemblies and membrane structure. In multi-color live cell experiments, we can produce quantitative measurements of interactions and dynamics of membrane components that are sensitive enough to detect lipid-mediated organization of plasma membrane components. We will use this technique to address the questions of how membrane signaling “compartments”, or local membrane microenvironments, are generated in the plasma membrane, how receptor signaling cascades couple to membrane compartments, and how membrane compartments influence the activity of signaling proteins. In addition to native TCRs, the research outlined in this proposal uses engineered immune receptors known as chimeric antigen receptors (CARs) as a simplified, modular, modifiable, model receptor to study spatial regulation of immune signaling. In parallel, we will develop new super-resolution probes and imaging modalities to access spatial information about signaling activity (e.g. kinase activity, phosphorylation) and physical properties of membrane microenvironments (e.g. surface charge, and lipid composition). Project Number: 1R35GM162563-01 | Fiscal Year: 2026 | NIH Institute/Center: National Institute of General Medical Sciences (NIGMS) | Principal Investigator: Sarah Shelby | Institution: UNIVERSITY OF TENNESSEE KNOXVILLE, KNOXVILLE, TN | Award Amount: $413,069 | Activity Code: R35 | Study Section: Special Emphasis Panel[ZRG1 CDB-N (55)] View on NIH RePORTER: https://reporter.nih.gov/project-details/1R35GM16256301