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PMID: 22523077 Published · ppublish English Journal Article Research Support, N.I.H., Extramural

Silent scaffolds: inhibition OF c-Jun N-terminal kinase 3 activity in cell by dominant-negative arrestin-3 mutant.

The Journal of biological chemistry ·Vol. 287 ·No. 23 ·2012-06-01 ·Pages 19653-64

Breitman M, Kook S, Gimenez LE, Lizama BN, Palazzo MC, Gurevich EV, Gurevich VV

Abstract

We established a new in vivo arrestin-3-JNK3 interaction assay based on bioluminescence resonance energy transfer (BRET) between JNK3-luciferase and Venus-arrestins. We tested the ability of WT arrestin-3 and its 3A mutant that readily binds β2-adrenergic receptors as well as two mutants impaired in receptor binding, Δ7 and KNC, to directly bind JNK3 and to promote JNK3 phosphorylation in cells. Both receptor binding-deficient mutants interact with JNK3 significantly better than WT and 3A arrestin-3. WT arrestin-3 and Δ7 mutant robustly promoted JNK3 activation, whereas 3A and KNC mutants did not. Thus, receptor binding, JNK3 interaction, and JNK3 activation are three distinct arrestin functions. We found that the KNC mutant, which tightly binds ASK1, MKK4, and JNK3 without facilitating JNK3 phosphorylation, has a dominant-negative effect, competitively decreasing JNK activation by WT arrestin-3. Thus, KNC is a silent scaffold, a novel type of molecular tool for the suppression of MAPK signaling in living cells.

MeSH Terms
Animals Arrestins/genetics,metabolism Biological Assay/methods COS Cells Chlorocebus aethiops Enzyme Activation/genetics Humans MAP Kinase Kinase 4/genetics,metabolism MAP Kinase Kinase Kinase 5/genetics,metabolism MAP Kinase Signaling System Mitogen-Activated Protein Kinase 10/genetics,metabolism Mutation Phosphorylation/genetics Protein Binding/genetics
Chemicals
Arrestins arrestin3 Mitogen-Activated Protein Kinase 10 MAP Kinase Kinase Kinase 5 MAP Kinase Kinase 4 MAP2K4 protein, human
Authors & Affiliations
7 authors, click to expand affiliations / ORCID
Breitman Maya
Department of Pharmacology, Vanderbilt University, Nashville, Tennessee 37232, USA.
Kook Seunghyi
Gimenez Luis E
Lizama Britney N
Palazzo Maria C
Gurevich Eugenia V
Gurevich Vsevolod V
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Article Info
Journal
The Journal of biological chemistry
Abbr.
J Biol Chem
ISSN
1083-351X
DOI
10.1074/jbc.M112.358192
Published
2012-06-01
Epub
2012-00-20
Pages
19653-64
Language
English
Region
United States
NLM ID
2985121R
PMCID
PMC3366000
Subset
IM
Grants
NIGMS NIH HHS · GM081756 · United States
NIGMS NIH HHS · R01 GM077561 · United States
NIGMS NIH HHS · T34 GM008718 · United States
NIGMS NIH HHS · R01 GM081756 · United States
NINDS NIH HHS · NS065868 · United States
NEI NIH HHS · EY011500 · United States
NIGMS NIH HHS · R25 GM062459 · United States
NIGMS NIH HHS · GM077561 · United States
NEI NIH HHS · R01 EY011500 · United States
NINDS NIH HHS · R01 NS065868 · United States
External Links
PubMed source DOI: 10.1074/jbc.M112.358192 PMC Full Text View journal details
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