New sorting nexin (SNX27) and NHERF specifically interact with the 5-HT4(a) receptor splice variant: roles in receptor targeting

Abstract

). Most of them are scaffolding proteins that contain several structural interaction domains such as Src homology 2 (SH2) or 3 (SH3) domains, post-synapticdensity-95/disc-large/zonula-occludens-1 (PDZ) domains and Drosophila enabled and vasodilator-stimulated phosphoprotein homologous (EVH) domains. These proteins participate in the building of large submembrane protein signaling networks. We have recently isolated a complex of at least 15 proteins interacting with the C-terminal tail of the 5-HT2C receptor, using a proteomic approach combining peptide-affinity chromatography and mass spectrometry. This further supports the interaction of the GPCR C-terminus with large protein networks. Moreover, functional studies have established that the proteins that interact with the GPCR C-terminus are implicated in various GPCR functions that do not involve Gproteins. These functions include trafficking, targeting to specific subcellular compartments, clustering with effectors, fine tuning of G-protein activation and desensitization.

Uncoupling and Endocytosis of 5-Hydroxytryptamine 4 Receptors

“…Indeed, a 15-min stimulation of the receptor was sufficient to induce receptor trafficking to a perinuclear compartment, where the receptor bound by arrestin was retained for Ͼ6 h. This behavior clearly classifies the 5-HT 4 Rs into class B GPCRs (33). We have recently reported that 5-HT 4 Rs are associated with a series of GPCR-interacting proteins, including SNX-27 (sorting nexin-27) and the Na ϩ /H ϩ exchanger regulatory factor (61). Although the role of GPCR-interacting proteins in 5-HT 4 R endocytosis and recycling remains to be analyzed, they have been shown to be clearly implicated in other GPCR systems (62,63).…”

Constitutive dimerization of human serotonin 5‐HT4 receptors in living cells

“…The low BRET signal observed with the h5-HT 4(g) R-RLuc construct suggests that the C-terminus of this splice variant may influence the spatial organization of the Rluc and therefore the intensity of the energy transfer between the donor and the acceptor. It is interesting to note that the mouse 5-HT 4(e) R splice variant specifically binds with its C-terminus to several PDZ-domain containing proteins [25]. Since the h5-HT 4(g) R is very similar to the mouse 5-HT 4(e) R and also contains a canonical recognition motif for PDZ domains at their extreme C-termini [25], it is therefore likely that the presence of these PDZdomain containing proteins in the h5-HT 4(g) R modifies the relative distance and orientation of Rluc and YFP within dimers in such a way that no significant energy transfer is detectable.…”

“…It is interesting to note that the mouse 5-HT 4(e) R splice variant specifically binds with its C-terminus to several PDZ-domain containing proteins [25]. Since the h5-HT 4(g) R is very similar to the mouse 5-HT 4(e) R and also contains a canonical recognition motif for PDZ domains at their extreme C-termini [25], it is therefore likely that the presence of these PDZdomain containing proteins in the h5-HT 4(g) R modifies the relative distance and orientation of Rluc and YFP within dimers in such a way that no significant energy transfer is detectable. Results obtained in membranes prepared from these cells support this interpretation since specific BRET signals have been observed in these preparations most likely due to the lost of the PDZ domain containing proteins.…”

“…Some differences have been noted in 5-HT 4 R isoform intrinsic properties such as their constitutive activity [30], desensitisation process [12], and second messenger production in response to 5-HT 4 ligands [11,13,31]. In addition, as mentioned above, specific proteins to some 5-HT 4 R splice variants associate to their intracellular C-terminal tail [25] and may contribute to the functional specificity of the isoforms. Therefore, it is tempting to speculate that depending on the nature of the 5-HT 4 R heterodimer, 5-HT 4 R functional response may greatly differ from the coexpressed and isolated receptors.…”

Pathophysiological consequences of receptor mistraffic: Tales from the platelet P2Y 12 receptor

“…12 cargo proteins including a number of GPCRs such as the β2-adrenoceptor, 5HT 2 receptor and parathyroid hormone receptor (78)(79)(80). Detailed proteomic analysis conducted in mammalian cell lines has revealed that a loss of SNX27 expression results in the downregulation of multiple membrane proteins (81).…”