BoneKEy-Osteovision | Commentary

A novel PTH/PTHrP receptor (PPRc) mutation: The ongoing tale of PPRc and the growth plate becomes more complex

DOI:10.1138/2002036

The parathyroid hormone/parathyroid hormone-related protein receptor (PPRc) is a 7-transmembrane domain G-protein coupled receptor that mediates both the endocrine actions of parathyroid hormone (PTH) and the auto/paracrine actions of parathyroid hormone-related protein (PTHrP) (). PPRc binds PTH and PTHrP with equal affinity, and upon challenge with either PTH or PTHrP, is able to stimulate the accumulation of at least two second messengers, cAMP and phosphoinositol, through the coupling with Gs and Gq/G11, respectively (). Numerous in vitro and in vivo findings place PPRc as a central regulator of both mineral ion homeostasis and endochondral bone development ().

Endochondral bone development () involves a two-stage mechanism whereby chondrocytes form a matrix template in which osteoblasts differentiate and initiate the ossification process. The well-ordered and controlled proliferation, differentiation, and apoptotic death of these chondrocytes is crucial for proper control of bone elongation, since it sets the stage for the timing of the replacement of the cartilage matrix with a trabecular bone matrix. The analysis of both murine models (,,,) and two human chondrodysplasias () have clearly demonstrated that PTHrP and PPRc play a critical role in this process, by keeping chondrocytes in the proliferative pool and delaying their hypertrophy. Furthermore, it is now established that Indian Hedgehog (Ihh) is an upstream regulator of PTHrP production in the chondrocytic growth plate (). Ihh is a member of a family of proteins that are important for embryonic patterning, and Ihh is highly expressed in the transition zone between proliferating and hypertrophic cells (). It appears that Ihh stimulates chondrocyte proliferation mainly through a PTHrP-independent mechanism (), and, concomitantly, delays their hypertrophy by increasing PTHrP synthesis by the periarticular chrondrocytes (). Conversely, PTHrP has been reported to be a negative regulator of Ihh expression in chondrocytes in vitro ().

In a fascinating and challenging study recently published by Nature Genetics (), Hopyan et al. now add another interesting piece to the puzzle by identifying and characterizing a novel PPRc mutation (R150C) as the likely cause of some cases of human enchondromatosis. Enchondromas are common benign cartilage tumors of bone that can occur as solitary lesions or as multiple lesions in enchondromatosis. Hopyan et al. have analyzed the PPRc in six cases of enchondromatosis, and in two of them have identified the heterozygous missense mutation R150C. One patient had inherited the mutation from the father, who had only a very mild chondrodysplasia; in the second case the mutation was limited to the tumor tissue. When tested in vitro, the mutant PPRc appeared to be very poorly expressed, and displayed a severe impairment of both binding to PTH and cAMP production upon challenge with the agonist, very likely as a result of impaired cell surface expression. Interestingly, cells transfected with the mutant PPRc appeared to have higher basal cAMP levels than controls, when basal cAMP values were corrected for level of receptor cell surface expression. Based on these findings, the authors conclude that R150C mutant is indeed a constitutively active PPRc. Heterozygous missense mutations that cause ligand-independent constitutive cAMP accumulation have been identified in the PPRc of patients with Jansen's metaphyseal chondrodysplasia, a rare form of short limbed dwarfism associated with hypercalcemia and low or undetectable levels of circulating PTH (). Interestingly, the patient heterozygous for the R150C mutation, unlike the patients with Jansen's disease, did not display any sign of chondrodysplasia besides the enchondromas, and presumably was not hypercalcemic. Of course, a variety of considerations could explain the obvious differences. The R150C mutation could affect receptor cell surface expression in a more severe manner than the Jansen mutations, or the Jansen mutations so far identified could be intrinsically more potent in terms of constitutive activity. Alternatively, it is possible that constitutive activity is not the only critical feature of the R150C mutant PPRc, and that other PPRc signaling properties not yet explored could also be affected by the R150C mutation, in addition to the increase of basal cAMP levels.

Hopyan et al. then generated transgenic mice expressing the R150C mutant PPRc in the chondrocytic growth plate under the control of the collagen type II promoter. Consistent with the findings in humans, transgenic long bones were not shorter than controls even if the hypertrophic layer appeared to be reduced in size. More importantly, in adulthood they showed persistence of cartilage islands in the bony diaphyses that were phenotypically similar to the human “enchondromas”. In the authors’ view this result would be consistent with the role of PPRc in delaying the process of endochondral bone development. Interestingly, both Jansen patients and Jansen mice in which a Jansen mutant PPRc was expressed in the growth plate by using a collagen type II promoter similar to the one adopted by Hopyan et al. are dwarfed, and no enchondromas have been reported in either one. Is the R150C-dependent increase in basal cAMP accumulation the cause of the “enchondroma” lesions both in humans and mice?

Furthermore, which are the biological mechanisms that link the R150C mutant PPRc to the enchondroma formation? In order to address this question, Hopyan and colleagues investigate the effect in vitro of the R150C mutation on Ihh activity. They transfected HeLa cells with a Hedgehog-responsive Gli-2 luciferase reporter gene. Co-transfection of cells with the R150C mutant PPRc, but not with the wild-type PPRc, resulted in a constitutive activation of the reporter. Furthermore, transgenic overexpression in vivo in the growth plate of Gli-2 caused “enchondroma” lesions similar to the ones observed in the R150C-PPRc transgenic animals. Based on these findings, Hopyan et al. conclude that the R150C mutation over-activates the Ihh signaling pathway. This is a fascinating hypothesis, but in real life the picture could be a bit more complicated. Increased activity of the Gli-2 reporter is not per se a direct demonstration of increased Hedgehog activity; furthermore, the relation between cAMP and the Hedgehog system is a complex one, in the sense that both inhibitory and stimulatory effects of cAMP on the Hedgehog signaling pathway have been reported ().

More studies will be necessary in order to unveil the pathway that links the R150C mutant PPRc to the enchondroma phenotype. It is not difficult to predict that very likely other fascinating chapters will be soon added to the ongoing tale of PPRC and the growth plate.

This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 3.0 United States License.