One of the most fascinating research problems currently discussed in cell biology and membrane biophysics is that which relates to understanding the relationship between nanoscale membrane heterogeneity (domain formation) and membrane function. In particular, sphingolipid (SL)-cholesterol (CHOL)-rich domains, also known as lipid rafts, have been identified to play a key role in the functionality of biomembranes. Though the important functional role of raft domains has been established, the biophysics of raft assembly remains poorly understood. This research project seeks to address an important question related to this still open topic: What are the biophysical mechanisms that are involved in the recruitment of membrane proteins to raft domains? The goal will be to explore the three most likely mechanisms of recruitment: (1) via binding ligands; (2) via SL-CHOL-rich lipid shells around membrane proteins; and (3) via other raft-associated proteins and lipids. To address the question of protein recruitment to raft domains, the experimental strategy will be to study the recruitment processes in planar model membranes using epifluorescence microscopy, wide-field single molecule fluorescence microscopy, and fluorescence correlation spectroscopy (FCS). The model membrane approach enables membrane protein recruitment studies to be conducted under controlled membrane conditions without the great complexity found in cells. Imaging experiments will be performed on membrane proteins, which are considered to be permanently raft-excluded (transferrin receptor), permanently raft-associated [urokinase plasminogen activator receptor (uPAR)], and temporarily raft-associated (avb3 and a5b1 integrins). The complementary imaging approach will allow the PI to study the protein-raft co-localization (epifluorescence microscopy), the membrane composition-dependent protein lateral mobility (wide-field single molecule fluorescence microscopy), and the formation of small protein-lipid and protein-protein aggregates (FCS).
This project will be an important part of the PI's goal to develop innovative, interdisciplinary research and teaching programs at the interface between classical sciences and engineering disciplines. To reach this goal, research results will be implemented into courses taught by the PI. Furthermore, this project will provide scientific training for one undergraduate and two graduate students. Because interdisciplinary teamwork and communication are key elements of the training program, students of the PI's research group will have the opportunity to participate in collaborative research with students from excellent research groups in Germany. Finally, as co-director of the newly established IUPUI Nanoscale Imaging Center, the PI seeks also to outreach into the local community to promote interdisciplinary research by providing cutting-edge facilities for biomolecular imaging.
