Nanoscopic spine localization of Norbin, an mGluR5 accessory protein.

Linda Westin,Maria Lindskog,Anita Aperia,Matthias Reuss,Hjalmar Brismar

doi:10.1186/1471-2202-15-45

Abstract

BackgroundNorbin 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.ResultsHere 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.ConclusionsNorbin 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.

Highlights

Norbin is a neuron-specific, cytosolic protein that interacts with the metabotropic glutamate receptor 5 and has a profound impact on mGluR5 signaling
We have previously reported that Norbin acts as an accessory protein to the metabotropic glutamate receptor 5
The endogenous expression pattern of Norbin was identified in hippocampal neurons immunolabeled for Norbin and double-stained for postsynaptic density (PSD)-95, used as a marker for excitatory synapses (Figure 2A,B)

Summary

Introduction

Norbin is a neuron-specific, cytosolic protein that interacts with the metabotropic glutamate receptor 5 (mGluR5) and has a profound impact on mGluR5 signaling. Norbin/Neurochondrin is a cytosolic, neuron-specific, somatodendritic protein abundantly expressed in the central nervous system (CNS) [1]. We have previously reported that Norbin acts as an accessory protein to the metabotropic glutamate receptor 5 (mGluR5). The interaction between Norbin and mGluR5 results in increased cell surface expression of mGluR5 and augmentation of mGluR5 mediated signaling. Accessory proteins in spines play an essential role in regulating the availability and confinement of glutamate receptors [5,6]. Little is known about the localization of Norbin inside the spine

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Discussion

Conclusion