title: Expression of the rat muscarinic receptor gene m3 in Dictyostelium discoideum.
abstract: We functionally expressed the rat muscarinic m3 receptor (rm3) in the cellular slime mold Dictyostelium discoideum under the control of the homologous discoidin I gamma promoter. Cells transfected with the authentic rm3 receptor gene expressed about 100 functional receptor molecules per cell, corresponding to a Bmax for [3H]-NMS of 36 +/- 9 fmol/mg of protein in isolated membranes. Genetic fusion of the Dictyostelium contact site A (csA) leader peptide to the amino terminus of rm3 increased the receptor expression by about 17-fold. Remarkable, in [3H]-NMS ligand binding experiments performed with whole cells no characteristic saturable binding was observed and there was no significant difference in [3H]-NMS binding to whole cells of rm3 and csA/rm3 transformants. The recombinant rm3 receptor showed an about 10-fold higher affinity to the M3-selective antagonist p-F-HHSiD compared to the M2-selective antagonist AQ-RA 741, suggesting that membranes derived from transgenic D. discoideum cells may be useful for the search of new subtype-specific muscarinic receptor ligands.
authors.affiliation: Institute for Pharmaceutical Biology, Johann Wolfgang Goethe-University, Frankfurt/Main, Germany.
authors.forename: G
authors.initials: G
authors.lastname: Voith
authors.forename: H
authors.initials: H
authors.lastname: Kramm
authors.forename: I
authors.initials: I
authors.lastname: Zündorf
authors.forename: T
authors.initials: T
authors.lastname: Winkler
authors.forename: T
authors.initials: T
authors.lastname: Dingermann
mesh_terms: D000818:Animals; D015152:Blotting, Northern; D004023:Dictyostelium / Q000378:metabolism*; D005822:Genetic Vectors; D007700:Kinetics; D008024:Ligands; D009154:Mutation; D051381:Rats; D011976:Receptors, Muscarinic / Q000096:biosynthesis* / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism
chemical_list: D008024:Ligands; D011976:Receptors, Muscarinic; D011994:Recombinant Proteins