title: The N-terminal region of the human 5-HT₂C receptor has as a cleavable signal peptide.
abstract: The 5-hydroxytryptamine 2C (5-HT(2C)) receptor has a single nucleotide polymorphism (SNP) site at amino acid position 23 in its N-terminal tail. The polymorphism involves conversion of a cysteine to serine. The site, designated C23S, is located within a 32 amino acid long predicted signal peptide. The aim of the present study was to investigate whether the 5-HT(2C) receptor indeed has a functional cleavable signal peptide. For this purpose, ten N-terminally modified 5-HT(2C) receptors were constructed. Modifications included addition of the influenza virus hemagglutinin signal peptide, addition of a FLAG epitope, truncation of the N-terminal tail, and combinations of these changes. The receptors were transiently expressed in COS-7 cells. The relative amounts of receptors expressed at the membranes were quantified by [(3)H]-mesulergine radioligand binding. In one of the receptor constructs the FLAG epitope was inserted just after the endogenous putative signal peptide. Immunostaining with the M1 antibody, which recognizes the FLAG epitope only as free N-terminal entity, was used to detect whether the putative signal peptide preceding the FLAG epitope was cleaved off. The results suggest the following conclusions. The predicted signal peptide in the N-terminal tail of the 5-HT(2C) receptor acts as a cleavable signal peptide. Cleaving of the signal peptide is important for translocation of the wild type receptor to the plasma membrane. The two amino acids differentially encoded by the C23S SNP are likely absent from the mature 5-HT(2C) receptor.
authors.affiliation: Section of Pharmacology, Institute of Medicine, Faculty of Medicine and Dentistry, University of Bergen, Norway. jan.jahnsen@med.uib.no
authors.forename: Jan Anker
authors.initials: JA
authors.lastname: Jahnsen
authors.forename: Staffan
authors.initials: S
authors.lastname: Uhlén
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002462:Cell Membrane / Q000378:metabolism; D002522:Chlorocebus aethiops; D004873:Ergolines / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D008969:Molecular Sequence Data; D020641:Polymorphism, Single Nucleotide; D021382:Protein Sorting Signals; D017433:Protein Structure, Secondary; D059748:Proteolysis; D044405:Receptor, Serotonin, 5-HT2C / Q000737:chemistry* / Q000378:metabolism*; D014162:Transfection
chemical_list: D004873:Ergolines; D021382:Protein Sorting Signals; D044405:Receptor, Serotonin, 5-HT2C; C031649:mesulergine