The serotonin 5-HT2A and 5-HT2C receptors interact with specific sets of PDZ proteins.
The 5-hydroxytryptamine type 2A (5-HT(2A)) receptor and the 5-HT(2C) receptor are closely related members of the G-protein-coupled receptors activated by serotonin that share very similar pharmacological profiles and cellular signaling pathways. These receptors express a canonical class I PDZ ligand (SXV) at their C-terminal extremity. Here, we have identified proteins that interact with the PDZ ligand of the 5-HT(2A) and 5-HT(2C) receptors by a proteomic approach associating affinity chromatography using immobilized synthetic peptides encompassing the PDZ ligand and mass spectrometry. We report that both receptor C termini interact with specific sets of PDZ proteins in vitro. The 5-HT(2C) receptor but not the 5-HT(2A) receptor binds to the Veli-3.CASK.Mint1 ternary complex and to SAP102. In addition, the 5-HT(2C) receptor binds more strongly to PSD-95 and MPP-3 than the 5-HT(2A) receptor. In contrast, a robust interaction between the 5-HT(2A) receptor and the channel-interacting PDZ protein CIPP was found, whereas CIPP did not significantly associate with the 5-HT(2C) receptor. We also show that residues located at the -1 position and upstream the PDZ ligand in the C terminus of the 5-HT(2A) and 5-HT(2C) receptors are major determinants in their interaction with specific PDZ proteins. Immunofluorescence and electron microscopy studies strongly suggested that these specific interactions also take place in living cells and that the 5-HT(2) receptor-PDZ protein complexes occur in intracellular compartments. The interaction of the 5-HT(2A) and the 5-HT(2C) receptor with specific sets of PDZ proteins may contribute to their different signal transduction properties.
UPR CNRS 2580 and CNRS UMR 5101, 141 rue de la Cardonille, 34094 Montpellier Cedex 5, France.
Carine
C
Bécamel
Sophie
S
Gavarini
Benjamin
B
Chanrion
Gérard
G
Alonso
Nathalie
N
Galéotti
Aline
A
Dumuis
Joël
J
Bockaert
Philippe
P
Marin
D000818:Animals; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism; D019556:COS Cells; D002845:Chromatography; D002846:Chromatography, Affinity; D000076122:Disks Large Homolog 4 Protein; D015180:Electrophoresis, Gel, Two-Dimensional; D051528:Guanylate Kinases; D015151:Immunoblotting; D007150:Immunohistochemistry; D047908:Intracellular Signaling Peptides and Proteins; D008024:Ligands; D013058:Mass Spectrometry; D008565:Membrane Proteins; D051379:Mice; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D009419:Nerve Tissue Proteins / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D020543:Proteome; D044402:Receptor, Serotonin, 5-HT2A / Q000737:chemistry / Q000378:metabolism*; D044405:Receptor, Serotonin, 5-HT2C / Q000737:chemistry / Q000378:metabolism*; D015398:Signal Transduction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D000076122:Disks Large Homolog 4 Protein; C501447:Dlg4 protein, mouse; D047908:Intracellular Signaling Peptides and Proteins; D008024:Ligands; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D010455:Peptides; D020543:Proteome; D044402:Receptor, Serotonin, 5-HT2A; D044405:Receptor, Serotonin, 5-HT2C; C033652:postsynaptic density proteins; D051528:Guanylate Kinases