Dynamic mass redistribution reveals diverging importance of PDZ-ligands for G protein-coupled receptor pharmacodynamics

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• 作者：Nathan D. Camp^a ; Kyung-Soon Lee^b ; Allison Cherry^b ; Jennifer L. Wacker-Mhyre^b ; Timothy S. Kountz^b ; Ji-Min Park^b ; Dorathy-Ann Harris^b ; Marianne Estrada^b ; Aaron Stewart^b ; Nephi Stella^b ; Alejandro Wolf-Yadlin^a ; Chris Hague^b ; ^{chague@uw.edu" class="auth_mail" title="E-mail the corresponding author}
• 关键词：ADRA1D ; α1Dadrenergic receptor ; ADRA2B ; α2Badrenergic receptor ; ADRB1 ; β1-adrenergic receptor ; ADRB2 ; β2adrenergic receptor ; C3AR1 ; Complement Component 3a Receptor 1 ; CXCR1 ; chemokine receptor 1 ; CXCR2 ; chemokine receptor 2 ; CXCR3 ; chemokine receptor 3 ; CXCR5 ; chemokine receptor 5 ; GALR1 ; galanin receptor 1 ; HRH3 ; histamine receptor H3 ; HTR2A ; 5-hydroxytryptamine (serotonin) receptor 2A ; HTR2B ; 5-hydroxytryptamine (serotonin) receptor 2B ; HTR2C ; 5-hydroxytryptamine (serotonin) receptor 2C ; LPAR
• 刊名：Pharmacological Research
• 出版年：2016
• 出版时间：March 2016
• 年：2016
• 卷：105
• 期：Complete
• 页码：13-21
• 全文大小：2208 K

文摘

G protein-coupled receptors (GPCRs) are essential membrane proteins that facilitate cell-to-cell communication and co-ordinate physiological processes. At least 30 human GPCRs contain a Type I PSD-95/DLG/Zo-1 (PDZ) ligand in their distal C-terminal domain; this four amino acid motif of X-[S/T]-X-[φ] sequence facilitates interactions with PDZ domain-containing proteins. Because PDZ protein interactions have profound effects on GPCR ligand pharmacology, cellular localization, signal-transduction effector coupling and duration of activity, we analyzed the importance of Type I PDZ ligands for the function of 23 full-length and PDZ-ligand truncated (ΔPDZ) human GPCRs in cultured human cells. SNAP-epitope tag polyacrylamide gel electrophoresis revealed most Type I PDZ GPCRs exist as both monomers and multimers; removal of the PDZ ligand played minimal role in multimer formation. Additionally, SNAP-cell surface staining indicated removal of the PDZ ligand had minimal effects on plasma membrane localization for most GPCRs examined. Label-free dynamic mass redistribution functional responses, however, revealed diverging effects of the PDZ ligand. While no clear trend was observed across all GPCRs tested or even within receptor families, a subset of GPCRs displayed diminished agonist efficacy in the absence of a PDZ ligand (i.e. HT2RB, ADRB1), whereas others demonstrated enhanced agonist efficacies (i.e. LPAR2, SSTR5). These results demonstrate the utility of label-free functional assays to tease apart the contributions of conserved protein interaction domains for GPCR signal-transduction coupling in cultured cells.