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Protein kinase-zeta interacts with munc18c: role in GLUT4 trafficking
C P Hodgkinson1, A Mander, G J Sale
1School of Biological Sciences, University of Southampton, Bassett Crescent East, Southampton SO16 7PX, UK.
Diabetologia
|June 30, 2005
Summary
Researchers identified a new link in insulin signaling: protein kinase Czeta (PKCzeta) interacts with munc18c. This interaction is crucial for insulin to stimulate glucose uptake and GLUT4 vesicle movement.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Insulin stimulates glucose transport via a signaling cascade involving protein kinase Czeta/lambda (PKCzeta/lambda) and protein kinase B (PKB).
- This cascade culminates in the translocation of GLUT4 vesicles to the plasma membrane, facilitating glucose uptake.
- The precise molecular links between upstream kinases and the GLUT4 vesicle trafficking machinery remain incompletely understood.
Purpose of the Study:
- To identify novel components that bridge the gap between insulin-regulated kinases and the GLUT4 vesicle trafficking system.
- To elucidate the molecular mechanisms underlying insulin-stimulated glucose transport.
Main Methods:
- A yeast two-hybrid screen was employed using full-length mouse munc18c as bait.
- Glutathione S-transferase (GST) pull-down assays were performed to confirm and characterize the interaction between PKCzeta and munc18c.
- Immunoprecipitation and Western blotting were used to assess the association of endogenous PKCzeta and munc18c in vivo.
Main Results:
- The yeast two-hybrid screen identified PKCzeta as a novel binding partner for munc18c.
- GST pull-downs confirmed the interaction, mapping the binding site to residues 295-338 of munc18c and the N-terminal region of PKCzeta.
- Endogenous PKCzeta and munc18c were found to associate in various cell types, with insulin stimulation increasing this association approximately threefold.
- Disruption of the PKCzeta-munc18c interaction significantly impaired insulin-stimulated glucose uptake and GLUT4 translocation.
Conclusions:
- A novel, insulin-regulated physiological interaction between munc18c and PKCzeta has been identified.
- This interaction establishes a direct link between the insulin signaling kinase PKCzeta and the GLUT4 vesicle trafficking component munc18c.
- The findings suggest a model where insulin promotes PKCzeta docking to munc18c, thereby enhancing GLUT4 vesicle translocation and glucose uptake.