Mapping of Calmodulin and Gβγ Binding Domains within the C-terminal Region of the Metabotropic Glutamate Receptor 7A

Oussama El Far,Elisa Bofill-Cardona,José M Airas,Vincent O'Connor,Stefan Boehm,Michael Freissmuth,Christian Nanoff,Heinrich Betz

doi:10.1074/jbc.m102573200

Abstract

Ca(2+)/calmodulin (Ca(2+)/CaM) and the betagamma subunits of heterotrimeric G-proteins (Gbetagamma) have recently been shown to interact in a mutually exclusive fashion with the intracellular C terminus of the presynaptic metabotropic glutamate receptor 7 (mGluR 7). Here, we further characterized the core CaM and Gbetagamma binding sequences. In contrast to a previous report, we find that the CaM binding motif localized in the N-terminal region of the cytoplasmic tail domain of mGluR 7 is conserved in the related group III mGluRs 4A and 8 and allows these receptors to also bind Ca(2+)/CaM. Mutational analysis of the Ca(2+)/CaM binding motif is consistent with group III receptors containing a conventional CaM binding site formed by an amphipathic alpha-helix. Substitutions adjacent to the core CaM target sequence selectively prevent Gbetagamma binding, suggesting that the CaM-dependent regulation of signal transduction involves determinants that overlap with but are different from those mediating Gbetagamma recruitment. In addition, we present evidence that Gbetagamma uses distinct nonoverlapping interfaces for interaction with the mGluR 7 C-terminal tail and the effector enzyme adenylyl cyclase II, respectively. Although Gbetagamma-mediated signaling is abolished in receptors lacking the core CaM binding sequence, alpha subunit activation, as assayed by agonist-dependent GTPgammaS binding, was not affected. This suggests that Ca(2+)/CaM may alter the mode of group III mGluR signaling from mono- (alpha) to bidirectional (alpha and betagamma) activation of downstream effector cascades.

Highlights

Glutamate receptors mediate excitatory neurotransmission at most synapses in the central nervous system
In contrast to a previous report, we find that the CaM binding motif localized in the N-terminal region of the cytoplasmic tail domain of metabotropic glutamate receptors (mGluRs) 7 is conserved in the related group III mGluRs 4A and 8 and allows these receptors to bind Ca2؉/CaM
Our previous [9] and the present results demonstrate that Ca2ϩ/CaM and G␤␥ share some common binding determinants that reside in the N-terminal region of the metabotropic glutamate receptor 7 (mGluR 7) C-tail that follows the last transmembrane segment

Summary

Introduction

Glutamate receptors mediate excitatory neurotransmission at most synapses in the central nervous system. Ca2ϩ/CaM binding to this region of the receptor promotes G protein-mediated signaling by displacing G␤␥ subunits from the C-terminal tail [9].

Results

Conclusion