Abstract
Potential G protein-coupled receptor kinase (GRK) and protein kinase A (PKA) mediation of homologous desensitization of corticotropin-releasing factor type 1 (CRF1) receptors was investigated in human retinoblastoma Y-79 cells. Inhibition of PKA activity by PKI(5-22) or H-89 failed to attenuate homologous desensitization of CRF1 receptors, and direct activation of PKA by forskolin or dibutyryl cAMP failed to desensitize CRF-induced cAMP accumulation. However, treatment of permeabilized Y-79 cells with heparin, a nonselective GRK inhibitor, reduced homologous desensitization of CRF1 receptors by approximately 35%. Furthermore, Y-79 cell uptake of a GRK3 antisense oligonucleotide (ODN), but not of a random or mismatched ODN, reduced GRK3 mRNA expression by approximately 50% without altering GRK2 mRNA expression and inhibited homologous desensitization of CRF1 receptors by approximately 55%. Finally, Y-79 cells transfected with a GRK3 antisense cDNA construct exhibited an approximately 50% reduction in GRK3 protein expression and an ~65% reduction in homologous desensitization of CRF1 receptors. We conclude that GRK3 contributes importantly to the homologous desensitization of CRF1 receptors in Y-79 cells, a brain-derived cell line.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Colforsin / pharmacology
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Cyclic AMP-Dependent Protein Kinases / antagonists & inhibitors
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Cyclic AMP-Dependent Protein Kinases / genetics
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Cyclic AMP-Dependent Protein Kinases / metabolism
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DNA, Antisense
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Enzyme Inhibitors / pharmacology
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Eye Neoplasms
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G-Protein-Coupled Receptor Kinase 3
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Gene Expression Regulation, Enzymologic / drug effects
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Gene Expression Regulation, Enzymologic / physiology
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Humans
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Isoquinolines / pharmacology
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Oligodeoxyribonucleotides, Antisense / pharmacology
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Peptide Fragments / pharmacology
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Protein Serine-Threonine Kinases / genetics
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Protein Serine-Threonine Kinases / metabolism*
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RNA, Messenger / genetics
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Receptors, Corticotropin-Releasing Hormone / drug effects
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Receptors, Corticotropin-Releasing Hormone / genetics
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Receptors, Corticotropin-Releasing Hormone / physiology*
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Recombinant Proteins / metabolism
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Retinoblastoma
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Sulfonamides*
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Transcription, Genetic / drug effects
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Transfection
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Tumor Cells, Cultured
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beta-Adrenergic Receptor Kinases
Substances
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DNA, Antisense
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Enzyme Inhibitors
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Isoquinolines
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Oligodeoxyribonucleotides, Antisense
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Peptide Fragments
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RNA, Messenger
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Receptors, Corticotropin-Releasing Hormone
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Recombinant Proteins
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Sulfonamides
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Colforsin
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CRF receptor type 1
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Protein Serine-Threonine Kinases
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Cyclic AMP-Dependent Protein Kinases
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G-Protein-Coupled Receptor Kinase 3
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GRK3 protein, human
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beta-Adrenergic Receptor Kinases
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N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide