Cloning and functional expression of the bovine natriuretic peptide receptor-B (natriuretic factor R1c subtype)

We describe the isolation of a 3,276 base pair cDNA for the bovine natriuretic peptide receptor-B (NPR-B). Expression of this clone in Cos-P cells demonstrates that it encodes an agonist-dependent guanylyl cyclase. Porcine CNP stimulates the activity of this receptor up to 200-fold with an ED₅₀ of 12±2 nM, whereas brain natriuretic peptide C-type natriuretic peptide (CNP) and atrial natriuretic factor (ANF) are less efficacious. In addition, ligand binding studies indicate that this receptor exhibits the pharmacology appropriate for the bovine NPR-B. CNP binds to Cos-P cell membranes expressing this clone with a K_d of 13±1 pM, and natriuretic peptides compete for ¹²⁵I]-CNP binding with a rank order of pCNP>pBNP>rANF. Thus, the expressed receptor-guanylyl cyclase exhibits the expected pharmacological profile for ligand binding and cyclase activation of the bovine NPR-B receptor.Abbreviations BSA bovine serum albumin - dNTP deoxynucleotide triphosphate - SDS sodium dodecyl sulfate - DEAE-dextran diethylaminoethyl-dextran - EDTA ethylenediamine tetraacetic acid - Tris Tris(hydroxymethyl)aminomethane - DMSO dimethyl sulfoxide - RP-HPLC reverse phase-high performance liquid chromatography - AMV avian myeloblastosis virus - Arg arginine - Lys lysine