Abstract
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-HT3 receptors. Therefore, we examined the impact of RIC-3 on differently composed 5-HT3 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. Ca2+ 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-HT3 receptor antagonist [H-3]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-HT3 receptor composition in vivo.
| Original language | English |
|---|---|
| Pages (from-to) | 26956-26965 |
| Number of pages | 10 |
| Journal | Journal of Biological Chemistry |
| Volume | 285 |
| Issue number | 35 |
| DOIs | |
| Publication status | Published - 27 Aug 2010 |
Keywords
- NICOTINIC ACETYLCHOLINE-RECEPTORS
- ENDOPLASMIC-RETICULUM CHAPERONE
- FUNCTIONAL EXPRESSION
- MOLECULAR-CLONING
- SEROTONIN
- PROTEIN
- GENE
- TRAFFICKING
- MATURATION
Cite this
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RIC-3 Exclusively Enhances the Surface Expression of Human Homomeric 5-Hydroxytryptamine Type 3A (5-HT(3)A) Receptors Despite Direct Interactions with 5-HT3A, -C, -D, and -E Subunits. / Walstab, Jutta; Hammer, Christian; Lasitschka, Felix; Möller, Dorothee; Connolly, Christopher N.; Rappold, Gudrun; Bruess, Michael; Boenish, Heinz; Niesler, Beate.
In: Journal of Biological Chemistry, Vol. 285, No. 35, 27.08.2010, p. 26956-26965.Research output: Contribution to journal › Article
TY - JOUR
T1 - RIC-3 Exclusively Enhances the Surface Expression of Human Homomeric 5-Hydroxytryptamine Type 3A (5-HT(3)A) Receptors Despite Direct Interactions with 5-HT3A, -C, -D, and -E Subunits
AU - Walstab, Jutta
AU - Hammer, Christian
AU - Lasitschka, Felix
AU - Möller, Dorothee
AU - Connolly, Christopher N.
AU - Rappold, Gudrun
AU - Bruess, Michael
AU - Boenish, Heinz
AU - Niesler, Beate
PY - 2010/8/27
Y1 - 2010/8/27
N2 - 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-HT3 receptors. Therefore, we examined the impact of RIC-3 on differently composed 5-HT3 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. Ca2+ 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-HT3 receptor antagonist [H-3]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-HT3 receptor composition in vivo.
AB - 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-HT3 receptors. Therefore, we examined the impact of RIC-3 on differently composed 5-HT3 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. Ca2+ 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-HT3 receptor antagonist [H-3]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-HT3 receptor composition in vivo.
KW - NICOTINIC ACETYLCHOLINE-RECEPTORS
KW - ENDOPLASMIC-RETICULUM CHAPERONE
KW - FUNCTIONAL EXPRESSION
KW - MOLECULAR-CLONING
KW - SEROTONIN
KW - PROTEIN
KW - GENE
KW - TRAFFICKING
KW - MATURATION
U2 - 10.1074/jbc.M110.122838
DO - 10.1074/jbc.M110.122838
M3 - Article
C2 - 20522555
VL - 285
SP - 26956
EP - 26965
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 35
ER -