Spatial-temporal patterning of metabotropic glutamate receptor-mediated inositol 1,4,5-triphosphate, calcium, and protein kinase C oscillations - Protein kinase C-dependent receptor phosphorylation is not required

John P. Robarts Research Institute, P. O. Box 5015, 100 Perth Drive, London, Ontario N6A 5K8, Canada.
Journal of Biological Chemistry (Impact Factor: 4.57). 10/2001; 276(38):35900-8. DOI: 10.1074/jbc.M103847200
Source: PubMed


The metabotropic glutamate receptors (mGluR), mGluR1a and mGluR5a, are G protein-coupled receptors that couple via Gq to the hydrolysis of phosphoinositides, the release of Ca2+ from intracellular stores, and the activation of protein kinase C (PKC). We show here that mGluR1/5 activation results in
oscillatory G protein coupling to phospholipase C thereby stimulating oscillations in both inositol 1,4,5-triphosphate formation
and intracellular Ca2+ concentrations. The mGluR1/5-stimulated Ca2+ oscillations are translated into the synchronized repetitive redistribution of PKCβII between the cytosol and plasma membrane.
The frequency at which mGluR1a and mGluR5a subtypes stimulate inositol 1,4,5-triphosphate, Ca2+, and PKCβII oscillations is regulated by the charge of a single amino acid residue localized within their G protein-coupling
domains. However, oscillatory mGluR signaling does not involve the repetitive feedback phosphorylation and desensitization
of mGluR activity, since mutation of the putative PKC consensus sites within the first and second intracellular loops as well
as the carboxyl-terminal tail does not prevent mGluR1a-stimulated PKCβII oscillations. Furthermore, oscillations in Ca2+ continued in the presence of PKC inhibitors, which blocked PKCβII redistribution from the plasma membrane back into the cytosol.
We conclude that oscillatory mGluR signaling represents an intrinsic receptor/G protein coupling property that does not involve
PKC feedback phosphorylation.

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    • "eptor phosphorylation , specifi - cally at Ser 839 , to uncouple the receptor from Gq / 11 and an as yet poorly defined protein phosphatase activity rapidly dephosphorylates the receptor to allow restoration of mGlu5 receptor – Gq / 11 coupling ( Kawabata et al . , 1996 ; Nakahara et al . , 1997 ; Nash et al . , 2002 ; Kim et al . , 2005 ; but see Dale et al . , 2001 ) . Stimulation of the astrocyte mGlu5 receptor initiates robust Ca 2+ oscillations that can reach a frequency of approxi - mately three oscillations per minute . Application of calyculin A or cantharidin , relatively selective inhibitors of the serine / threonine protein phosphatases PP1 / PP2A ( Ishihara et al . , 1989 ; Honkanen , 19"
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    ABSTRACT: Cyclical phosphorylation and dephosphorylation of a key residue within the C-terminal domain of the activated type 5 metabotropic glutamate (mGlu₅) receptor is believed to cause the synchronous, oscillatory changes in inositol 1,4,5-trisphosphate and Ca²⁺ levels observed in a variety of cell types. Here, we have attempted to better define the kinase and phosphatase enzymes involved in this modulation. Ca²⁺ and [³H]inositol phosphate ([³H]IP(x) ) measurements in astrocyte preparations have been used to evaluate the effects of pharmacological inhibition of protein kinase C (PKC) and protein phosphatase activities and small interfering RNA-mediated specific PKC isoenzymic knock-down on mGlu₅ receptor signalling. Ca²⁺ oscillation frequency or [³H]IP(x) accumulation in astrocytes stimulated by mGlu₅ receptors, was concentration-dependently decreased by protein phosphatase-1/2A inhibition or by PKC activation. PKC inhibition also increased [³H]IP(x) accumulation two- to threefold and changed the Ca²⁺ response into a peak-plateau response. However, selective inhibition of conventional PKC isoenzymes or preventing changes in [Ca²⁺](i) concentration by BAPTA-AM loading was without effect on mGlu₅ receptor-stimulated [³H]IP(x) accumulation. Selective knock-down of PKCδ was without effect on glutamate-stimulated Ca²⁺ responses; however, selective PKCε knock-down in astrocytes changed Ca²⁺ responses from oscillatory into peak-plateau type. These data confirm the acute regulation of mGlu₅ receptor signalling by protein kinases and protein phosphatases and provide novel data pinpointing the isoenzymic dependence of this regulation in the native mGlu₅ receptor-expressing rat cortical astrocyte. These data also highlight a potential alternative mechanism by which mGlu₅ receptor signalling might be therapeutically manipulated.
    British Journal of Pharmacology 04/2011; 164(2b):755-71. DOI:10.1111/j.1476-5381.2011.01421.x · 4.84 Impact Factor
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    • "In transfected cells, mGluR1a activation induced a single-peaked Ca 2+ rise whereas mGluR5a activation elicited characteristic Ca 2+ oscillations (Kawabata et al., 1996). The latter depends on the phosphorylation at a single amino acid (Kawabata et al., 1996; Uchino et al., 2004; Kim et al., 2005; but Dale et al., 2001). Early studies suggest that the phosphorylation of the mGluR5a C terminus at T840 by PKC (likely the Ca 2+ -independent PKCδ isoform rather than Ca 2+ dependent PKCγ) is responsible for the generation of Ca 2+ oscillations in mGluR5a-expressing cells (Kawabata et al., 1996; Uchino et al., 2004). "
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    Neuropharmacology 07/2008; 55(4):403-8. DOI:10.1016/j.neuropharm.2008.05.034 · 5.11 Impact Factor
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    • "The mix is highly variable, ranging from no PKC involvement (e.g. Dale et al. 2001) to PKC inhibition late in the process (e.g. Muyderman et al. 1998) to critical involvement (e.g. "
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    The Journal of Physiology 01/2005; 562(Pt 2):553-68. DOI:10.1113/jphysiol.2004.076919 · 5.04 Impact Factor
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