RIC-3 exclusively enhances the surface expression of human homomeric 5-hydroxytryptamine type 3A (5-HT3A) receptors despite direct interactions with 5-HT3A, -C, -D, and -E subunits.
Although five 5-hydroxytryptamine type 3 (5-HT3) subunits (A-E) have been cloned, knowledge on the regulation of their assembly is limited. RIC-3 has been identified as a chaperone specific for the pentameric ligand-gated nicotinic acetylcholine and 5-HT(3) receptors. Therefore, we examined the impact of RIC-3 on differently composed 5-HT(3) receptors with the focus on 5-HT3C, -D, and -E subunits. The influence of RIC-3 on these receptor subtypes is supported by the presence of RIC3 mRNA in tissues expressing at least one of the subunits 5-HT3C, -D, and -E. Furthermore, immunocytochemical studies on transfected mammalian cells revealed co-localization in the endoplasmic reticulum and direct interaction of RIC-3 with 5-HT3A, -C, -D, and -E. Functional and pharmacological characterization was performed using HEK293 cells expressing 5-HT3A or 5-HT3A + 5-HT3B (or -C, -D, or -E) in the presence or absence of RIC-3. Ca(2+) influx analyses revealed that RIC-3 does not influence the 5-HT concentration-response relationship on 5-HT(3)A receptors but leads to differential increases of 5-HT-induced maximum response (E(max)) on cells expressing different subunits. Increases of E(max) were due to analogously enhanced B(max) values for binding of the 5-HT(3) receptor antagonist [(3)H]GR65630. The observed enhanced cell surface expression of the tested 5-HT3 subunit combinations correlated with the increased surface expression of 5-HT3A as determined by flow cytometry. In conclusion, we showed that RIC-3 can interact with 5-HT3A, -C, -D, and -E subunits and predominantly enhances the surface expression of homomeric 5-HT(3)A receptors in HEK293 cells. These data implicate a possible role of RIC-3 in determining 5-HT(3) receptor composition in vivo.
Walstab
Jutta
J
Department of Human Molecular Genetics, University of Heidelberg, Im Neuenheimer Feld 366, 69120 Heidelberg, Germany; Institute of Pharmacology and Toxicology, University of Bonn, Reuterstrasse 2b, 53113 Bonn, Germany.
Hammer
Christian
C
Department of Human Molecular Genetics, University of Heidelberg, Im Neuenheimer Feld 366, 69120 Heidelberg, Germany.
Lasitschka
Felix
F
Institute of Pathology, University of Heidelberg, Im Neuenheimer Feld 220/221, 69120 Heidelberg, Germany.
Möller
Dorothee
D
Department of Human Molecular Genetics, University of Heidelberg, Im Neuenheimer Feld 366, 69120 Heidelberg, Germany.
Connolly
Christopher N
CN
Centre for Neuroscience, Division of Medical Sciences, Ninewells Medical School, University of Dundee, Dundee DD1 9SY, Scotland, United Kingdom.
Rappold
Gudrun
G
Department of Human Molecular Genetics, University of Heidelberg, Im Neuenheimer Feld 366, 69120 Heidelberg, Germany.
Brüss
Michael
M
Institute of Pharmacology and Toxicology, University of Bonn, Reuterstrasse 2b, 53113 Bonn, Germany.
Bönisch
Heinz
H
Institute of Pharmacology and Toxicology, University of Bonn, Reuterstrasse 2b, 53113 Bonn, Germany.
Niesler
Beate
B
Department of Human Molecular Genetics, University of Heidelberg, Im Neuenheimer Feld 366, 69120 Heidelberg, Germany. Electronic address: beate.niesler@med.uni-heidelberg.de.
D000328:Adult; D045744:Cell Line, Tumor; D004721:Endoplasmic Reticulum / Q000235:genetics / Q000378:metabolism; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D008297:Male; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism*; D021122:Protein Subunits / Q000096:biosynthesis / Q000235:genetics; D044406:Receptors, Serotonin, 5-HT3 / Q000096:biosynthesis* / Q000235:genetics; D058831:Serotonin 5-HT3 Receptor Antagonists
C476055:HTR3E protein, human; D007093:Imidazoles; D007211:Indoles; D047908:Intracellular Signaling Peptides and Proteins; D018832:Molecular Chaperones; D021122:Protein Subunits; C496701:RIC3 protein, human; D044406:Receptors, Serotonin, 5-HT3; D058831:Serotonin 5-HT3 Receptor Antagonists; C059068:GR 65630