Interaction with NEGR1 affects mGlu5 cell signalling and the neuroprotection induced by the positive allosteric modulator VU0409551.

Abstract

Background and purpose: The metabotropic glutamate receptor mGlu₅ plays an important role in neuronal survival and synaptic plasticity. We and others have shown that mGlu₅ positive allosteric modulators (PAMs) are biased agonists, triggering the activation of neuroprotective pathways and promoting neuronal survival. As mGlu₅ biased pharmacology arises from drug-induced interactions with different protein partners, in this study, we analysed the mGlu₅ interactome to identify novel binding partners capable of modulating receptor cell signalling.

Experimental approach: To identify new mGlu₅ partners, we employed mass spectrometry, followed by co-immunoprecipitation experiments, which confirmed the interaction between mGlu₅ and the neuronal growth regulator 1 (NEGR1), a cell adhesion molecule. We next evaluated how NEGR1 affects mGlu₅ cell signalling and cellular distribution using lentivirus transduction on primary neuronal cultures to knockdown NEGR1 expression. We also assessed whether NEGR1 plays a role in the neuroprotective effects of the mGlu5 PAM, VU0409551, by evaluating apoptotic cell death and neurite degeneration.

Key results: The interaction between NEGR1 and mGlu₅ is important for mGlu₅ synaptic localization and for its coupling with Homer, thereby modulating cell signalling pathways activated by mGlu₅ receptors. For instance, NEGR1 knockdown led to decreased mGlu₅-dependent activation of ERK1/2 and reduced expression of ARC/ARG3.1 and BDNF. Importantly, NEGR1 knockdown affected mGlu₅ pharmacology, as the neuroprotective effect of VU0409551 was not observed in neurons expressing low levels of NEGR1.

Conclusions and implications: These results indicate that NEGR1 is essential for mGlu₅-biased activation of neuroprotective cell signalling pathways and for the neuroprotective effects triggered by VU0409551.