Source:http://linkedlifedata.com/resource/pubmed/id/12815037
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| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:issue |
36
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| pubmed:dateCreated |
2003-9-1
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| pubmed:abstractText |
G-protein receptor kinase and beta-arrestin mediated desensitization of the rat kappa-opioid receptor (KOR) was previously shown using Xenopus oocyte expression to require serine 369 within the C terminus of KOR. To define the effects of phosphorylation of this residue in desensitization and internalization processes in mammalian expression systems, wild-type KOR-green fluorescent protein (KOR-GFP) and KOR(S369A)-GFP were stably expressed in AtT-20 and HEK293 cells. Using whole-cell patch clamp recording in transfected AtT-20 cells, agonist activation of either kappa receptor form produced equivalent activation of the intrinsic G-protein-gated inwardly rectifying potassium channel. Incubation for 60 min with the kappa agonist U50,488 (100 nm) desensitized the response in cells expressing wild-type KOR-GFP by 86% but had no effect on KOR(S369A)-GFP-expressing cells. Phosphorylation of serine 369 was detected using a phosphospecific antibody (KOR-P) able to distinguish the phosphorylated form of the receptor. The agonist-induced increase in KOR-P labeling was dose-dependent, blocked by co-treatment with the kappa antagonist norbinaltorphimine, and prevented by co-expression of the dominant negative form of the G-protein receptor kinase, GRK2(K220R). In contrast, agonist-induced increase in KOR-P labeling was not evident in KOR(S369A) expressing cells. Prolonged activation resulted in receptor internalization that was also blocked by KOR(S369A) substitution, but interestingly, KOR-P labeling was evident at lower agonist concentrations than required to induce internalization. Following the removal of agonist, receptor dephosphorylation detected by loss of KOR-P labeling was complete within 60 min, could be blocked by okadaic acid, and was not blocked by sucrose inhibition of receptor internalization. These results demonstrate that GRK-mediated phosphorylation of serine 369 mediates rat KOR desensitization and internalization.
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| pubmed:grant | |
| pubmed:language |
eng
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| pubmed:journal | |
| pubmed:citationSubset |
IM
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| pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/DNA, Complementary,
http://linkedlifedata.com/resource/pubmed/chemical/Green Fluorescent Proteins,
http://linkedlifedata.com/resource/pubmed/chemical/Luminescent Proteins,
http://linkedlifedata.com/resource/pubmed/chemical/Peptides,
http://linkedlifedata.com/resource/pubmed/chemical/Potassium Channels,
http://linkedlifedata.com/resource/pubmed/chemical/Receptors, Opioid, kappa,
http://linkedlifedata.com/resource/pubmed/chemical/Recombinant Fusion Proteins,
http://linkedlifedata.com/resource/pubmed/chemical/Serine
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| pubmed:status |
MEDLINE
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| pubmed:month |
Sep
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| pubmed:issn |
0021-9258
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| pubmed:author | |
| pubmed:issnType |
Print
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| pubmed:day |
5
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| pubmed:volume |
278
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| pubmed:owner |
NLM
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| pubmed:authorsComplete |
Y
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| pubmed:pagination |
34631-40
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| pubmed:dateRevised |
2011-1-19
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| pubmed:meshHeading |
pubmed-meshheading:12815037-Amino Acid Sequence,
pubmed-meshheading:12815037-Animals,
pubmed-meshheading:12815037-Cell Line,
pubmed-meshheading:12815037-DNA, Complementary,
pubmed-meshheading:12815037-Dose-Response Relationship, Drug,
pubmed-meshheading:12815037-Electrophysiology,
pubmed-meshheading:12815037-Enzyme-Linked Immunosorbent Assay,
pubmed-meshheading:12815037-Green Fluorescent Proteins,
pubmed-meshheading:12815037-Humans,
pubmed-meshheading:12815037-Immunohistochemistry,
pubmed-meshheading:12815037-Luminescent Proteins,
pubmed-meshheading:12815037-Microscopy, Confocal,
pubmed-meshheading:12815037-Microscopy, Fluorescence,
pubmed-meshheading:12815037-Models, Biological,
pubmed-meshheading:12815037-Molecular Sequence Data,
pubmed-meshheading:12815037-Mutagenesis, Site-Directed,
pubmed-meshheading:12815037-Patch-Clamp Techniques,
pubmed-meshheading:12815037-Peptides,
pubmed-meshheading:12815037-Phosphorylation,
pubmed-meshheading:12815037-Potassium Channels,
pubmed-meshheading:12815037-Protein Structure, Tertiary,
pubmed-meshheading:12815037-Rats,
pubmed-meshheading:12815037-Receptors, Opioid, kappa,
pubmed-meshheading:12815037-Recombinant Fusion Proteins,
pubmed-meshheading:12815037-Serine,
pubmed-meshheading:12815037-Temperature,
pubmed-meshheading:12815037-Time Factors,
pubmed-meshheading:12815037-Transfection
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| pubmed:year |
2003
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| pubmed:articleTitle |
Phosphorylation of a carboxyl-terminal serine within the kappa-opioid receptor produces desensitization and internalization.
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| pubmed:affiliation |
Department of Pharmacology, University of Washington School of Medicine, Seattle, Washington 98195-7280, USA.
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| pubmed:publicationType |
Journal Article,
Research Support, U.S. Gov't, P.H.S.
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