A Relationship between Protein Kinase C Phosphorylation and Calmodulin Binding to the Metabotropic Glutamate Receptor Subtype 7

Abstract

Metabotropic glutamate receptor subtype 7 (mGluR7) is coupled to the inhibitory cyclic AMP cascade and is selectively activated by a glutamate analogue, L-2-amino-4-phosphonobutyrate. Among L-2-amino-4-phosphonobutyrate-sensitive mGluR subtypes, mGluR7 is highly concentrated at the presynaptic terminals and is thought to play an important role in modulation of glutamatergic synaptic transmission by presynaptic inhibition of glutamate release. To gain further insight into the intracellular signaling mechanisms of mGluR7, with the aid of glutathione S-transferase fusion affinity chromatography, we attempted to identify proteins that interact with the intracellular carboxyl terminus of mGluR7. Here, we report that calmodulin (CaM) directly binds to the carboxyl terminus of mGluR7 in a Ca 2؉ -dependent manner. The CaM-binding domain is located immediately following the 7th transmembrane segment. We also show that the CaM-binding domain of mGluR7 is phosphorylated by protein kinase C (PKC). This phosphorylation is inhibited by the binding of Ca 2؉ /CaM to the receptor. Conversely, the Ca 2؉ /CaM binding is prevented by PKC phosphorylation. Collectively, these results suggest that mGluR7 serves to cross-link the cyclic AMP, Ca 2؉ , and PKC phosphorylation signal transduction cascades.

Calmodulin dynamically regulates the trafficking of the metabotropic glutamate receptor mGluR5

“…CaM has been shown to bind to several GPCRs (18)(19)(20)(21)(26)(27)(28), and in some cases, CaM is implicated in the regulation of GPCR trafficking. For example, agonist-induced endocytosis of the 5-HT 1A receptor is prevented by pharmacological inhibitors of CaM (29).…”

Tamalin Is a Scaffold Protein That Interacts with Multiple Neuronal Proteins in Distinct Modes of Protein-Protein Association

“…Glutathione-S-transferase (GST) Pull-down Assay, Immunoprecipitation, and Yeast Two-hybrid Screening-Protein purification and pulldown assay were performed as described previously (24,30). For pulldown assay, 10 g of GST fusion proteins were immobilized on glutathione-Sepharose 4B beads (25 l) and incubated with either supernatants (100 g) containing recombinant tamalin transiently expressed in COS-7 cells or the purified recombinant thioredoxin fusion proteins (10 g).…”

Physical Interaction of Calmodulin with the 5-Hydroxytryptamine2C Receptor C-Terminus Is Essential for G Protein-independent, Arrestin-dependent Receptor Signaling

“…For example, a set of postsynaptic density 95/disc-large/zona occludens proteins bind to the SSV motif on the distal C-terminal 5-HT 2C receptor and have a key role in receptor desensitization and internalization after prolonged agonist exposure (Gavarini et al, 2006). The 5-HT 2C receptor C-terminus also interacts with calmodulin (CaM) (Becamel et al, 2002), a ubiquitous modulator of Ca 2ϩ -dependent processes that has been found to interact with many other GPCRs, including the dopamine D 2 , metabotropic glutamate mGlu5 and mGlu7, -opioid, vasopressin V2, angiotensin II AT 1A , and serotonin 5-HT 1A and 5-HT 2A receptors (Minakami et al, 1997;Nakajima et al, 1999;O'Connor et al, 1999;Thomas et al, 1999;Wang et al, 1999;Bofill-Cardona et al, 2000;Nickols et al, 2004;Turner et al, 2004;Turner and Raymond, 2005).…”

“…Depending on the GPCR, CaM interaction with the receptor can affect its coupling with the G protein or its phosphorylation by protein kinase C (Nakajima et al, 1999;O'Connor et al, 1999;Wang et al, 1999;Bofill-Cardona et al, 2000;Turner et al, 2004;Turner and Raymond, 2005), suggesting a possible role in receptor desensitization. CaM can also directly contribute to receptor signaling (Della Rocca et al, 1999;O'Connor et al, 1999;Nickols et al, 2004).…”

“…For example, a set of postsynaptic density 95/disc-large/zona occludens proteins bind to the SSV motif on the distal C-terminal 5-HT 2C receptor and have a key role in receptor desensitization and internalization after prolonged agonist exposure (Gavarini et al, 2006). The 5-HT 2C receptor C-terminus also interacts with calmodulin (CaM) (Becamel et al, 2002), a ubiquitous modulator of Ca 2ϩ -dependent processes that has been found to interact with many other GPCRs, including the dopamine D 2 , metabotropic glutamate mGlu5 and mGlu7, -opioid, vasopressin V2, angiotensin II AT 1A , and serotonin 5-HT 1A and 5-HT 2A receptors (Minakami et al, 1997;Nakajima et al, 1999;O'Connor et al, 1999;Thomas et al, 1999;Wang et al, 1999;Bofill-Cardona et al, 2000;Nickols et al, 2004;Turner et al, 2004;Turner and Raymond, 2005).The roles that interaction with CaM plays in receptor function are remarkably diverse. Depending on the GPCR, CaM interaction with the receptor can affect its coupling with the G protein or its phosphorylation by protein kinase C (Nakajima et al, 1999;O'Connor et al, 1999;Wang et al, 1999;Bofill-Cardona et al, 2000;Turner et al, 2004;Turner and Raymond, 2005), suggesting a possible role in receptor desensitization.…”