CONSTITUTIVELY ACTIVE PTH/PTHRP RECEPTORS IN VIVO

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Jansen-type metaphyseal chondrodysplasia (JMC) is characterized by short stature due to impaired growth-plate maturation, and severe hypercalcemia and hypophosphatemia despite normal/undetectable parathyroid hormone (PTH) or PTH-related peptide (PTHrP). In two unrelated JMC patients, different PTH/PTHrP receptor mutations were identified that lead to constitutive receptor activation. While the findings explain the changes in mineral ion metabolism, the mechanisms that lead to the skeletal abnormalities remain obscure. The growth-plate is, however, one of the most prominent sites where PTHrP and the PTH/PTHrP receptor are expressed in a distinct temporal and spatial relationship. Mice which lack' most of the gene encoding PTHrP die shortly after birth from yet unknown causes and display severe abnormalities of endochondral bone formation, and thus provided first direct evidence for the biological importance of PTHrP in normal growth and skeletal development. Mice that lack most of the PTH/PTHrP receptor gene display a similar, but die in utero, indicating that PTHrP actions on growth-plates are mediated through the PTH/PTHrP receptor. Because of these findings, we hypothesize that constitutively active PTH/PTHrP receptors are necessary and sufficient to cause the JMC-specific growth plate abnormalities. We therefore, propose to develop transgenic mice in which the mutant PTH/PTHrP receptors are expressed under the control of either the type Il collagen promotor to direct its expression to "resting", proliferating, and the upper layer of hypertrophic chondrocytes, and the type X collagen promotor to direct receptor expression to hypertrophic chondrocytes. Growth plate analysis of the resulting animals should determine which chondrocytic cells mediate the receptor-dependent growth-plate abnormalities, and would allow the dissection of direct and indirect actions on cartilage development. Transgenic animals that display the JMC phenotype will then be used to rescue, at least partially, PTHrP-less animals. Due to the localized PTHrP-like action of the mutant PTH/PTHrP receptors, such hybrid animals may show a reversal/improvement of skeletal abnormalities, while the impact of generalized PTHrP-deficiency on other organs remains unchanged. To develop an animal model of JMC, portions of the wild-type PTH/PTHrP receptor gene will be replaced with that encoding the mutant receptor. Heterozygous animals will be used to determine how activating receptor mutations cause growth plate and mineral ion abnormalities, and matings between these and PTHrP-less mice will provide tools to determine further whether all actions of PTHrP are likely to be mediated through the common PTH/PTHrP receptor, or whether distinct receptors are involved. The proposed studies will thus take advantage of a rare human disease to define, the cellular and molecular mechanisms of PTHrP action that mediate its role in cartilage maturation and bone elongation. Insights from these studies may lead to treatment protocols of JMC, and more common forms of metabolic bone diseases, e.g. osteoporosis, renal osteodystrophy, and the growth failure in uremic children.