Abstract
Charged lipid species in the plasma-membrane play an integral role in a variety of signaling pathways, in which they function as second messangers as well as docking sites for proteins. It has been suggested that the spatial organization of charged lipids called phosphoinositides is not random_rather, they form nano-scale clusters that are distinct from lipid rafts. Through super-resolution imaging in live cells, we are investigating the role of these phosphoinositide nanodomains in providing spatial regulation of the PI3K/Akt and IP3 signaling pathways. Using PALM and photochromatic SOFI (pcSOFI) superresolution techniques, we are working to create high resolution, time-resolved lipid maps of the inner-leaflet of the plasma membrane. Lipids are probed using various fluorescent protein-tagged pleckstrin homology domains, which are coimaged with each other, with lipid raft markers, and with protein subunits. The resulting lipid maps will provide information on the how lipids in the plasma membrane are organized, and how this organization helps provide spatial regulation of the PI3K/Akt as well as the IP3 signaling pathways.
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