AUTHOR=Becirovic Elvir , Böhm Sybille , Nguyen Ong N. P. , Riedmayr Lisa M. , Hammelmann Verena , Schön Christian , Butz Elisabeth S. , Wahl-Schott Christian , Biel Martin , Michalakis Stylianos 

TITLE=AAV Vectors for FRET-Based Analysis of Protein-Protein Interactions in Photoreceptor Outer Segments

JOURNAL=Frontiers in Neuroscience

VOLUME=10

YEAR=2016

URL=https://www.frontiersin.org/journals/neuroscience/articles/10.3389/fnins.2016.00356

DOI=10.3389/fnins.2016.00356

ISSN=1662-453X

ABSTRACT=<p>Fluorescence resonance energy transfer (FRET) is a powerful method for the detection and quantification of stationary and dynamic protein-protein interactions. Technical limitations have hampered systematic <italic>in vivo</italic> FRET experiments to study protein-protein interactions in their native environment. Here, we describe a rapid and robust protocol that combines adeno-associated virus (AAV) vector-mediated <italic>in vivo</italic> delivery of genetically encoded FRET partners with <italic>ex vivo</italic> FRET measurements. The method was established on acutely isolated outer segments of murine rod and cone photoreceptors and relies on the high co-transduction efficiency of retinal photoreceptors by co-delivered AAV vectors. The procedure can be used for the systematic analysis of protein-protein interactions of wild type or mutant outer segment proteins in their native environment. Conclusively, our protocol can help to characterize the physiological and pathophysiological relevance of photoreceptor specific proteins and, in principle, should also be transferable to other cell types.</p>