title: Expression and membrane topology of Anopheles gambiae odorant receptors in lepidopteran insect cells.
abstract: A lepidopteran insect cell-based expression system has been employed to express three Anopheles gambiae odorant receptors (ORs), OR1 and OR2, which respond to components of human sweat, and OR7, the ortholog of Drosophila's OR83b, the heteromerization partner of all functional ORs in that system. With the aid of epitope tagging and specific antibodies, efficient expression of all ORs was demonstrated and intrinsic properties of the proteins were revealed. Moreover, analysis of the orientation of OR1 and OR2 on the cellular plasma membrane through the use of a novel 'topology screen' assay and FACS analysis demonstrates that, as was recently reported for the ORs in Drosophila melanogaster, mosquito ORs also have a topology different than their mammalian counterparts with their N-terminal ends located in the cytoplasm and their C-terminal ends facing outside the cell. These results set the stage for the production of mosquito ORs in quantities that should permit their detailed biochemical and structural characterization and the exploration of their functional properties.
authors.affiliation: Insect Molecular Genetics and Biotechnology Group, Institute of Biology, National Centre for Scientific Research "Demokritos", Athens, Greece.
authors.forename: Panagiota
authors.initials: P
authors.lastname: Tsitoura
authors.forename: Evi
authors.initials: E
authors.lastname: Andronopoulou
authors.forename: Daniela
authors.initials: D
authors.lastname: Tsikou
authors.forename: Adamantia
authors.initials: A
authors.lastname: Agalou
authors.forename: Maria P
authors.initials: MP
authors.lastname: Papakonstantinou
authors.forename: Georgia A
authors.initials: GA
authors.lastname: Kotzia
authors.forename: Vassiliki
authors.initials: V
authors.lastname: Labropoulou
authors.forename: Luc
authors.initials: L
authors.lastname: Swevers
authors.forename: Zafiroula
authors.initials: Z
authors.lastname: Georgoussi
authors.forename: Kostas
authors.initials: K
authors.lastname: Iatrou
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000852:Anopheles / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D005434:Flow Cytometry; D015870:Gene Expression; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D019476:Insect Proteins / Q000235:genetics / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D011485:Protein Binding; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D018035:Receptors, Odorant / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
chemical_list: D019476:Insect Proteins; D020033:Protein Isoforms; D018035:Receptors, Odorant; D049452:Green Fluorescent Proteins