Expression cloning of a rat B2 bradykinin receptor.
A cDNA encoding a functional bradykinin receptor was isolated from a rat uterus library by a clonal selection strategy using Xenopus laevis oocytes to assay for expression of bradykinin responses. The predicted protein is homologous to the seven transmembrane G protein-coupled superfamily of receptors. Bradykinin and its analogs stimulate a Cl- current oocytes expressing the receptor with the rank order of potency: bradykinin approximately Lys-bradykinin greater than [Tyr8]-bradykinin much greater than [Phe6]bradykinin. This is the rank order of potency observed for these compounds in competitive binding assays on soluble receptor from rat uterus. Des-Arg9-bradykinin (10 microM) elicits no response when applied to oocytes expressing the receptor; thus, the cDNA encodes a B2 type bradykinin receptor. [Thi5,8,DPhe7]bradykinin, where Thi is beta-(2-thienyl)-alanine, is a very weak partial agonist and inhibits the bradykinin-mediated ion flux, suggesting the cDNA encodes a smooth muscle, rather than a neuronal, B2 receptor subtype. Receptor message has a distribution consistent with previous reports of bradykinin function and/or binding in several tissues and is found in rat uterus, vas deferens, kidney, lung, heart, ileum, testis, and brain. Receptor subtypes are a possibility because several tissues contain two or three message species (4.0, 5.7, and 6.5 kilobases). Southern blot high-stringency analysis demonstrated that the rat, guinea pig, and human genomes contain a single gene. As bradykinin is a key mediator of pain, knowledge of the primary structure of this receptor will allow a molecular understanding of the receptor and aid the design of antagonists for pain relief.
McEachern
A E
AE
Howard Hughes Medical Institute, Stanford University, CA 94305.
Shelton
E R
ER
Bhakta
S
S
Obernolte
R
R
Bach
C
C
Zuppan
P
P
Fujisaki
J
J
Aldrich
R W
RW
Jarnagin
K
K
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001920:Bradykinin / Q000494:pharmacology* / Q000502:physiology; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D005260:Female; D015723:Gene Library; D007700:Kinetics; D008564:Membrane Potentials / Q000187:drug effects; D008969:Molecular Sequence Data; D009865:Oocytes / Q000187:drug effects / Q000502:physiology; D011061:Poly A / Q000235:genetics / Q000302:isolation & purification; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D018002:Receptors, Bradykinin; D017981:Receptors, Neurotransmitter / Q000235:genetics* / Q000502:physiology; D012689:Sequence Homology, Nucleic Acid; D014599:Uterus / Q000502:physiology; D014982:Xenopus laevis
D012333:RNA, Messenger; D018002:Receptors, Bradykinin; D017981:Receptors, Neurotransmitter; D011061:Poly A; D012313:RNA; D004247:DNA; D001920:Bradykinin