Targeted activation of conventional and novel protein kinases C through differential translocation patterns

Xin Hui; Gregor Reither; Lars Kaestner; Peter Lipp

doi:10.1128/MCB.00040-14

Targeted activation of conventional and novel protein kinases C through differential translocation patterns

Mol Cell Biol. 2014 Jul;34(13):2370-81. doi: 10.1128/MCB.00040-14. Epub 2014 Apr 14.

Authors

Xin Hui¹, Gregor Reither¹, Lars Kaestner¹, Peter Lipp²

Affiliations

¹ Molecular Cell Biology, Research Center for Molecular Imaging and Screening, Medical Faculty, Saarland University, Homburg/Saar, Germany.
² Molecular Cell Biology, Research Center for Molecular Imaging and Screening, Medical Faculty, Saarland University, Homburg/Saar, Germany [email protected].

Abstract

Activation of the two ubiquitous families of protein kinases, protein kinase A (PKA) and protein kinase C (PKC), is thought to be independently coupled to stimulation of Gαs and Gαq, respectively. Live-cell confocal imaging of protein kinase C fluorescent protein fusion constructs revealed that simultaneous activation of Gαs and Gαq resulted in a differential translocation of the conventional PKCα to the plasma membrane while the novel PKCδ was recruited to the membrane of the endoplasmic reticulum (ER). We demonstrate that the PKCδ translocation was driven by a novel Gαs-cyclic AMP-EPAC-RAP-PLCε pathway resulting in specific diacylglycerol production at the membrane of the ER. Membrane-specific phosphorylation sensors revealed that directed translocation resulted in phosphorylation activity confined to the target membrane. Specific stimulation of PKCδ caused phosphorylation of the inositol-1,4,5-trisphosphate receptor and dampening of global Ca(2+) signaling revealed by graded flash photolysis of caged inositol-1,4,5-trisphosphate. Our data demonstrate a novel signaling pathway enabling differential decoding of incoming stimuli into PKC isoform-specific membrane targeting, significantly enhancing the versatility of cyclic AMP signaling, thus demonstrating the possible interconnection between the PKA and PKC pathways traditionally treated independently. We thus provide novel and elementary understanding and insights into intracellular signaling events.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Calcium / metabolism
Calcium Signaling / physiology*
Cell Line
Cell Membrane / metabolism
Cyclic AMP / metabolism*
Cyclic AMP-Dependent Protein Kinases / metabolism
Diglycerides / biosynthesis
Endoplasmic Reticulum / metabolism
Enzyme Activation
GTP-Binding Protein alpha Subunits, Gq-G11 / metabolism
GTP-Binding Protein alpha Subunits, Gs / metabolism
Guanine Nucleotide Exchange Factors / genetics
HEK293 Cells
Humans
Indoles / pharmacology
Inositol 1,4,5-Trisphosphate / chemistry
Inositol 1,4,5-Trisphosphate Receptors / metabolism
Maleimides / pharmacology
Phosphoinositide Phospholipase C / genetics
Phosphorylation / drug effects
Protein Kinase C-alpha / antagonists & inhibitors
Protein Kinase C-alpha / genetics
Protein Kinase C-alpha / metabolism*
Protein Kinase C-delta / antagonists & inhibitors
Protein Kinase C-delta / genetics
Protein Kinase C-delta / metabolism*
Protein Transport / physiology
RNA Interference
RNA, Small Interfering
Tetradecanoylphorbol Acetate / pharmacology

Substances

2-(1-(3-dimethylaminopropyl)-5-methoxyindol-3-yl)-3-(1H-indol-3-yl)maleimide
Diglycerides
Guanine Nucleotide Exchange Factors
Indoles
Inositol 1,4,5-Trisphosphate Receptors
Maleimides
RAPGEF3 protein, human
RNA, Small Interfering
Inositol 1,4,5-Trisphosphate
Cyclic AMP
Cyclic AMP-Dependent Protein Kinases
Protein Kinase C-alpha
Protein Kinase C-delta
Phosphoinositide Phospholipase C
phospholipase C epsilon
GTP-Binding Protein alpha Subunits, Gq-G11
GTP-Binding Protein alpha Subunits, Gs
Tetradecanoylphorbol Acetate
Calcium