Protein kinase C phosphorylation of the metabotropic glutamate receptor mGluR5 on Serine 839 regulates Ca2+ oscillations.
The activation of Group 1 metabotropic glutamate receptors, mGluR5 and mGluR1alpha, triggers intracellular calcium release; however, mGluR5 activation is unique in that it elicits Ca2+ oscillations. A short region of the mGluR5 C terminus is the critical determinant and differs from the analogous region of mGluR1alpha by a single amino acid residue, Thr-840, which is an aspartic acid (Asp-854) in mGluR1alpha. Previous studies show that mGluR5-elicited Ca2+ oscillations require protein kinase C (PKC)-dependent phosphorylation and identify Thr-840 as the phosphorylation site. However, direct phosphorylation of mGluR5 has not been studied in detail. We have used biochemical analyses to directly investigate the phosphorylation of the mGluR5 C terminus. We showed that Ser-839 on mGluR5 is directly phosphorylated by PKC, whereas Thr-840 plays a permissive role. Although Ser-839 is conserved in mGluR1alpha (Ser-853), it is not phosphorylated, as the adjacent residue (Asp-854) is not permissive; however, mutagenesis of Asp-854 to a permissive alanine residue allows phosphorylation of Ser-853 on mGluR1alpha. We investigated the physiological consequences of mGluR5 Ser-839 phosphorylation using Ca2+ imaging. Mutations that eliminate Ser-839 phosphorylation prevent the characteristic mGluR5-dependent Ca2+ oscillations. However, mutation of Thr-840 to alanine, which prevents potential Thr-840 phosphorylation but is still permissive for Ser-839 phosphorylation, has no effect on Ca2+ oscillations. Thus, we showed that it is phosphorylation of Ser-839, not Thr-840, that is absolutely required for the unique Ca2+ oscillations produced by mGluR5 activation. The Thr-840 residue is important only in that it is permissive for the PKC-dependent phosphorylation of Ser-839.
Kim
Chul Hoon
CH
NINDS, National Institutes of Health, Bethesda, Maryland 20892, USA.
Braud
Stephanie
S
Isaac
John T R
JT
Roche
Katherine W
KW
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D020013:Calcium Signaling / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D011817:Rabbits; D064529:Receptor, Metabotropic Glutamate 5; D018094:Receptors, Metabotropic Glutamate / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D012694:Serine / Q000276:immunology / Q000378:metabolism
D000906:Antibodies; C578935:GRM5 protein, human; D064529:Receptor, Metabotropic Glutamate 5; D018094:Receptors, Metabotropic Glutamate; D012694:Serine; D011493:Protein Kinase C