Expression of mRNA of beta 1- and beta 2-adrenergic receptors in Xenopus oocytes results from structurally distinct receptor mRNAs.
Poly(A)+-selected RNA prepared from cells or tissues that express a homogeneous population of either beta 1- or beta 2-adrenergic receptors was isolated and then microinjected into Xenopus laevis oocytes. Following microinjection, the expression of beta-adrenergic receptors was assessed by equilibrium radioligand binding analysis using the antagonist ligand [3H]dihydroalprenolol. The pharmacology of the newly- expressed beta-adrenergic receptors in oocyte membranes was the same as that of the original tissue used as a source of RNA. Hybridization of nick-translated cDNA of hamster beta 2-adrenergic receptor to poly(A)+-selected RNA from tissues containing beta 2-adrenergic receptors was to a mRNA species of 2.2 kilobases. In contrast, hybridization of the cDNA probe to poly(A)+-selected RNA from tissues containing beta 1-adrenergic receptors was to a mRNA species of 2.0 kilobases. A single-stranded fragment of hamster beta 2-adrenergic receptor cDNA corresponding to nucleotides 730-886 was isolated and uniformly radiolabeled. This region of the gene is predicted to encode for the entire second exofacial loop (L4-5), the entire fifth transmembrane-spanning region, and the first 5 amino acid residues of the third cytoplasmic loop (L5-6) of the beta 2-adrenergic receptor. Hybridization at 48 and 56 degrees C of poly(A)+-selected RNA prepared from sources that express either beta 1 or beta 2-adrenergic receptors to the antisense orientation strand of this region of the beta 2-adrenergic receptor cDNA was followed by S1 endonuclease digestion of nonhybridized sequences. At 48 degrees C, S1-resistant hybrids from both sources of RNA protected the probe from S1 endonuclease digestion. At 56 degrees C, however, only the RNA prepared from the source of beta 2-adrenergic receptors protected the probe from S1 endonuclease digestion. These results demonstrate that the mRNAs encoding for the structurally homologous beta 1- and beta 2-adrenergic receptors are distinct in the pharmacological specificity of their translation products and in their size and structure.
Bahouth
S W
SW
Department of Pharmacological Sciences, School of Medicine, State University of New York, Stony Brook 11794-8651.
Hadcock
J R
JR
Malbon
C C
CC
D000818:Animals; D006224:Cricetinae; D004247:DNA / Q000235:genetics; D005260:Female; D005796:Genes; D006168:Guinea Pigs; D008297:Male; D051379:Mice; D009865:Oocytes / Q000378:metabolism*; D011061:Poly A / Q000235:genetics / Q000302:isolation & purification; D014176:Protein Biosynthesis; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D012333:RNA, Messenger / Q000235:genetics*; D051381:Rats; D011919:Rats, Inbred Strains; D011943:Receptors, Adrenergic, beta / Q000235:genetics* / Q000378:metabolism; D012110:Reserpine / Q000494:pharmacology; D013045:Species Specificity; D013363:Submandibular Gland / Q000187:drug effects / Q000378:metabolism; D014982:Xenopus laevis
D012333:RNA, Messenger; D011943:Receptors, Adrenergic, beta; D011061:Poly A; D012313:RNA; D012110:Reserpine; D004247:DNA