title: Molecular cloning of two isoforms of the guinea pig C3a anaphylatoxin receptor: alternative splicing in the large extracellular loop.
abstract: The anaphylatoxin C3a is released from C3 during complement activation. C3a is a potent spasmogen and has recently been described as an eosinophil and mast cell chemotactic factor that mediates a number of inflammatory reactions. Previously, we demonstrated the presence of a specific C3a receptor (C3aR) on guinea pig platelets. We report here the isolation of cDNA clones encoding for two isoforms of guinea pig C3aR (gpC3aR). Hydropathy analysis of the deduced amino acid sequence of both gpC3aR clones indicated seven transmembrane domains with a large extracellular (EC) loop between the fourth and fifth transmembrane domains, which is a known characteristic of the human C3aR. Northern blot analysis revealed that the gpC3aR was abundantly expressed on macrophages and in the spleen. A comparison of the deduced amino acid sequence of the larger gpC3aR (gpC3aR-L) with the recently cloned human C3aR indicated a 59.5% identity. The deduced amino acid sequence of the second, smaller cDNA clone was identical with gpC3aR-L, except that it lacked 35 amino acids in the large EC loop. Our evidence indicates that alternative splicing occurred in the large EC loop that accounts for these two isoforms. L cells separately expressing one of these two isoforms of the gpC3aR showed similar high-affinity C3a binding. An RT-PCR analysis documented that both forms of the C3aR were expressed in a variety of guinea pig tissues. The cloning and expression of these two natural forms of gpC3aR cDNA indicated that the deletion of the 35-residue portion of the large EC loop of gpC3aR-L did not alter C3a binding.
authors.affiliation: Department of Immunology, The Scripps Research Institute, La Jolla, CA 92037, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Fukuoka
authors.forename: J A
authors.initials: JA
authors.lastname: Ember
authors.forename: T E
authors.initials: TE
authors.lastname: Hugli
mesh_terms: D017398:Alternative Splicing / Q000276:immunology*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001792:Blood Platelets / Q000378:metabolism; D015152:Blotting, Northern; D003001:Cloning, Molecular; D015926:Complement C3a / Q000378:metabolism*; D018076:DNA, Complementary / Q000032:analysis / Q000235:genetics; D005434:Flow Cytometry; D006168:Guinea Pigs; D006801:Humans; D007536:Isomerism; D007739:L Cells; D008565:Membrane Proteins; D051379:Mice; D008969:Molecular Sequence Data; D009928:Organ Specificity / Q000235:genetics / Q000276:immunology; D016133:Polymerase Chain Reaction; D011485:Protein Binding / Q000235:genetics / Q000276:immunology; D017434:Protein Structure, Tertiary; D011951:Receptors, Complement / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014162:Transfection / Q000276:immunology
chemical_list: D018076:DNA, Complementary; D008565:Membrane Proteins; D011951:Receptors, Complement; C085567:complement C3a receptor; D015926:Complement C3a