Characterization of the effect of TIMAP phosphorylation on its interaction with protein phosphatase 1

TIMAP, TGF-β inhibited, membrane-associated protein, is highly abundant in endothelial cells (EC). We have shown earlier the involvement of TIMAP in PKA-mediated ERM (ezrin–radixin–moesin) dephosphorylation as part of EC barrier protection by TIMAP (Csortos et al., 2008). Emerging data demonstrate t...

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Bibliographic Details
Published in:Biochimie 2011-07, Vol.93 (7), p.1139-1145
Main Authors: Czikora, István, Kim, Kyung-mi, Kása, Anita, Bécsi, Bálint, Verin, Alexander D., Gergely, Pál, Erdődi, Ferenc, Csortos, Csilla
Format: Article
Language:English
Subjects:
Animals
Blotting, Western
Cell Line
Colforsin - pharmacology
Cyclic AMP-Dependent Protein Kinases - genetics
Cyclic AMP-Dependent Protein Kinases - metabolism
Cytoskeletal Proteins - metabolism
Endothelial Cells - drug effects
Endothelial Cells - metabolism
Glycogen Synthase Kinase 3 - antagonists & inhibitors
Glycogen Synthase Kinase 3 - genetics
Glycogen Synthase Kinase 3 - metabolism
Glycogen Synthase Kinase 3 beta
Humans
Kinetics
Membrane Proteins - genetics
Membrane Proteins - metabolism
Microfilament Proteins - metabolism
Microscopy, Fluorescence
Moesin
Phosphorylation
Protein Binding
Protein phosphatase 1
Protein Phosphatase 1 - genetics
Protein Phosphatase 1 - metabolism
Surface Plasmon Resonance
Thiazoles - pharmacology
TIMAP
Urea - analogs & derivatives
Urea - pharmacology
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Summary:TIMAP, TGF-β inhibited, membrane-associated protein, is highly abundant in endothelial cells (EC). We have shown earlier the involvement of TIMAP in PKA-mediated ERM (ezrin–radixin–moesin) dephosphorylation as part of EC barrier protection by TIMAP (Csortos et al., 2008). Emerging data demonstrate the regulatory role of TIMAP on protein phosphatase 1 (PP1) activity. We provide here evidence for specific interaction ( K a = 1.80 × 10 6 M −1) between non-phosphorylated TIMAP and the catalytic subunit of PP1 (PP1c) by surface plasmon resonance based binding studies. Thiophosphorylation of TIMAP by PKA, or sequential thiophosphorylation by PKA and GSK3β slightly modifies the association constant for the interaction of TIMAP with PP1c and decreases the rate of dissociation. However, dephosphorylation of phospho-moesin substrate by PP1cβ is inhibited to different extent in the presence of non- (∼60% inhibition), mono- (∼50% inhibition) or double-thiophosphorylated (
ISSN:0300-9084
1638-6183
DOI:10.1016/j.biochi.2011.03.011