PhD Defense by Mohammadreza Arastoo

Title: Lateral Organization of Plasma Membrane Proteins Studied Using Thermoplasmonics

Abstract: The eukaryotic cell membrane is a dynamic structure crowded with different protein species. Protein segregation in this crowded environment is crucial for their proper functioning which guarantees cell homeostasis. This is presumably done by laterally organizing the plasma membrane into nanoscopic local domains so-called ‘lipid rafts’. Rafts are highly dynamic structures (milliseconds) with a very small size (10-200nm). These two traits make them difficult targets to be directly visualized and investigated. Most of the methods reported so far for raft studies involve procedures which can potentially influence protein integrity and/or its localization on the membrane. Motivated by the challenges of raft visualization, we combined cell culturing with optical manipulations and confocal imaging to build a new system of hybrid vesicles to study lipid rafts and their associated proteins. Here, the protein of interest is carried by vesicles derived from viable cells and delivered into a phase-separated vesicle by thermoplasmonic based membrane fusion. To extend the applicability of the thermoplasmonic fusion method for protein research, we investigated how annexin A4 and A5 proteins are recruited to membrane ruptures induced in either cell membranes or model membranes. These experiments provide evidence for how cells respond to a photothermal damage. The use of plasmonic nanoparticles and optical trapping provides novel and efficient tool for investigating protein dynamics in membranes.