Nanoscopic spine localization of Norbin, an mGluR5 accessory protein
1 Department of Women’s and Children’s Health, Science for Life Laboratory, Karolinska Institutet, 17165 Solna, Sweden
2 Department of Applied Physics, Science for Life Laboratory, Royal Institute of Technology, 10691 Stockholm, Sweden
3 Department of Neuroscience, Karolinska Institutet, 17177 Stockholm, Sweden
BMC Neuroscience 2014, 15:45 doi:10.1186/1471-2202-15-45Published: 26 March 2014
Norbin is a neuron-specific, cytosolic protein that interacts with the metabotropic glutamate receptor 5 (mGluR5) and has a profound impact on mGluR5 signaling. Yet, little is known about its synaptic distribution.
Here we have analyzed the spatial relationship between Norbin, postsynaptic density protein 95 (PSD-95), actin and mGluR5 in spines using super-resolution microscopy. Norbin was found to have a high degree of colocalization with actin and a lower degree of colocalization with PSD-95. Co-immunoprecipitation studies confirmed that interaction occurs between Norbin and actin, but not between Norbin and PSD-95. Norbin was also found to have a high degree of colocalization with the perisynaptically located mGluR5. Findings based on structured illumination microscopy (3D-SIM) of exogenous expressed Norbin-GFP were confirmed by stimulated emission depletion microscopy (STED) of immunolabeled endogenous Norbin.
Norbin associates with actin rather than with PSD-95 in dendritic spines. Results regarding protein localization and colocalization performed with conventional confocal microscopy must be interpreted with great caution. The now available super-resolution microscopy techniques provide more accurate information about sub-cellular protein localization than previously was possible.