Structure and functional expression of a complementary DNA for porcine parathyroid hormone/parathyroid hormone-related peptide receptor.
A complementary DNA (cDNA) encoding the receptor for porcine parathyroid hormone/parathyroid hormone-related peptide (PTH/PTHrP) was isolated from a porcine kidney cDNA library. The porcine PTH/PTHrP receptor is a 585 amino acid protein containing seven putative membrane-spanning domains. The porcine PTH/PTHrP receptor has amino acid identity of 95.6%, 80.4%, and 88.7% with human, opossum, and rat PTH/PTHrP receptors, respectively and 53.4% identity to the recently cloned human PTH2 receptor. The receptor cDNA was subsequently cloned into a mammalian cell expression vector (pRC/CMV) which contains a human cytomegalovirus promoter. A human kidney cell line (293), stably transfected with this vector, expressed the receptor at a high level and, when challenged with human PTH(1-34), increased cytoplasmic cAMP and inositol triphosphate production. Radioligand binding studies revealed that the receptor bound both human PTH(1-34), and PTHrP(1-36). Scatchard analyses of three clones showed that the cells harbor a single class of high affinity receptor (Kd = 1-4 nM for human PTH(1-34)) but had varying receptor numbers (10(5)-10(6) receptors/cell). In contrast to PTH(1-34), the [Arg2]PTH(1-34) analog bound to the porcine PTH/PTHrP receptor with low affinity and was a weak agonist for cAMP stimulation with the cloned receptor. These response characteristics differentiate the porcine receptor from the previously cloned rat and opossum PTH/PTHrP receptors.
Smith
D P
DP
Endocrine Division, Lilly Research Laboratories, Eli Lilly and Company, Lilly Corporate Center, Indianapolis, IN 46285, USA.
Zhang
X Y
XY
Frolik
C A
CA
Harvey
A
A
Chandrasekhar
S
S
Black
E C
EC
Hsiung
H M
HM
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D000242:Cyclic AMP / Q000096:biosynthesis; D018076:DNA, Complementary / Q000737:chemistry* / Q000302:isolation & purification; D006801:Humans; D008969:Molecular Sequence Data; D051381:Rats; D044168:Receptor, Parathyroid Hormone, Type 1; D018016:Receptors, Parathyroid Hormone / Q000737:chemistry / Q000235:genetics* / Q000502:physiology; D013552:Swine
D018076:DNA, Complementary; D044168:Receptor, Parathyroid Hormone, Type 1; D018016:Receptors, Parathyroid Hormone; D000242:Cyclic AMP