Light shaping for unraveling protein oligomerization

Many vital processes in our bodies begin on the surface of our cells, where a wide variety of proteins come together and interact. Understanding how these proteins group together to form so-called oligomers and when they do so is crucial to understanding how cells communicate and respond to their environment. However, even with the most advanced microscopes, scientists still have difficulty observing these interactions clearly. Although the molecules can be observed as a whole, they are so close together that it is practically impossible to make statements about the state of individual proteins. To solve this problem, our team is developing a completely new imaging technique called Single Molecule Imaging upon Patterned Photobleaching (SMIPP). This method uses precisely shaped laser light to temporarily switch off or bleach small areas of the cell surface. We then observe how individual proteins migrate back into these bleached areas and can analyse them in detail. By repeating this process with a variety of carefully designed light patterns, we can gain unprecedented insight into the behaviour and organisation of proteins. The technology behind SMIPP is state-of-the-art. We use dynamic holograms to shape light in real time. This allows us to not only control exactly where the light hits the cell, but also to determine the angle of incidence so that only the cell membrane is bleached without damaging the cell interior with the intense laser light. As a first application, we will investigate the serotonin transporter (SERT), a protein that helps regulate mood and plays a central role in the action of many antidepressants. Understanding how SERT molecules interact could reveal new details about how these drugs work, leading to better treatments. This joint project brings together two teams of experts from Innsbruck and Vienna: one led by Alexander Jesacher at the Medical University of Innsbruck, which focuses on the development of microscopy and holography methods, and another led by Gerhard Schütz at the Vienna University of Technology, which is dedicated to biophysical issues.