Selective and nonselective inverse agonists for constitutively active type-1 parathyroid hormone receptors: evidence for altered receptor conformations.
The spontaneous signaling activity of some G protein-coupled receptors and the capacity of certain ligands (inverse agonists) to inhibit such constitutive activity are poorly understood phenomena. We investigated these processes for several analogs of PTH-related peptide (PTHrP) and the constitutively active human PTH/PTHrP receptors (hP1Rcs) hP1Rc-H223R and hP1Rc-T410P. The N-terminally truncated antagonist PTHrP(5-36) functioned as a weak partial/neutral agonist with both mutant receptors but was converted to an inverse agonist for both receptors by the combined substitution of Leu(11) and D-Trp(12). The N-terminally intact analog [Bpa(2)]PTHrP(1-36)-a partial agonist with the wild-type hP1Rc-was a selective inverse agonist, in that it depressed basal cAMP signaling by hP1Rc-H223R but enhanced signaling by hP1Rc-T410P. The ability of [Bpa(2)]PTHrP(1-36) to discriminate between the two receptor mutants suggested that H223R and T410P confer constitutive receptor activity by inducing distinct conformational changes. This hypothesis was confirmed by the observations that: 1) the double mutant receptor hP1Rc-H223R/T410P exhibited basal cAMP levels that were 2-fold higher than those of either single mutant; and 2) hP1Rc-H223R and hP1Rc-T410P internalized (125)I-PTHrP(5-36) to markedly different extents. The overall results thus reveal that two different types of inverse agonists are possible for PTHrP ligands (nonselective and selective) and that constitutively active PTH-1 receptors can access different conformational states.
Endocrine Unit, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114, USA.
P H
PH
Carter
B D
BD
Petroni
R C
RC
Gensure
E
E
Schipani
J T
JT
Potts
T J
TJ
Gardella
D000818:Animals; D001667:Binding, Competitive / Q000187:drug effects; D019556:COS Cells; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D004247:DNA / Q000235:genetics; D006801:Humans; D009154:Mutation; D010281:Parathyroid Hormone / Q000494:pharmacology; D010777:Photochemistry; D011487:Protein Conformation; D018016:Receptors, Parathyroid Hormone / Q000819:agonists* / Q000737:chemistry / Q000235:genetics; D015398:Signal Transduction; D014162:Transfection
D010281:Parathyroid Hormone; D018016:Receptors, Parathyroid Hormone; D004247:DNA; D000242:Cyclic AMP