Fibrodysplasia ossificans progressiva (FOP), a disorder of ectopic osteogenesis, misregulates cell surface expression and trafficking of BMPRIA.
FOP is a disorder in which skeletal muscle is progressively replaced with bone. FOP lymphocytes, a model system for exploring the BMP pathway in these patients, exhibit a defect in BMPRIA internalization and increased activation of downstream signaling, suggesting that altered BMP receptor trafficking underlies ectopic bone formation in this disease.


BACKGROUND
Fibrodysplasia ossificans progressiva (FOP) is a severely disabling disorder characterized by progressive heterotopic ossification of connective tissues. Whereas the genetic defect and pathophysiology of this condition remain enigmatic, BMP4 mRNA and protein are overexpressed, and mRNAs for a subset of secreted BMP antagonists are not synthesized at appropriate levels in cultured lymphocytes from FOP patients. These data suggest involvement of altered BMP signaling in the disease. In this study, we investigate whether the abnormality is associated with defective BMP receptor function in lymphocytes.


METHODS
Cell surface proteins were quantified by fluorescence-activated cell sorting (FACS). Protein phosphorylation was assayed by immunoprecipitation and immunoblotting. Protein synthesis and degradation were examined by [35S]methionine labeling and pulse-chase assays. mRNA was detected by RT-PCR.


RESULTS
FOP lymphocytes expressed 6-fold higher levels of BMP receptor type IA (BMPRIA) on the cell surface compared with control cells and displayed a marked reduction in ligand-stimulated internalization and degradation of BMPRIA. Moreover, in control cells, BMP4 treatment increased BMPRIA phosphorylation, whereas BMPRIA showed ligand-insensitive constitutive phosphorylation in FOP cells. Our data additionally support that the p38 mitogen-activated protein kinase (MAPK) signaling pathway is a major BMP signaling pathway in these cell lines and that expression of inhibitor of DNA binding and differentiation 1 (ID-1), a transcriptional target of BMP signaling, is enhanced in FOP cells.


CONCLUSIONS
These data extend our previous observations of misregulated BMP4 signaling in FOP lymphocytes and show that cell surface overabundance and constitutive phosphorylation of BMPRIA are associated with a defect in receptor internalization. Altered BMP receptor trafficking may play a significant role in FOP pathogenesis.
de la Peña
Lourdes Serrano
LS
Department of Orthopaedic Surgery, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104-6081, USA.
Billings
Paul C
PC
Fiori
Jennifer L
JL
Ahn
Jaimo
J
Kaplan
Frederick S
FS
Shore
Eileen M
EM
D055415:Bone Morphogenetic Protein 4; D052005:Bone Morphogenetic Protein Receptors, Type I; D019485:Bone Morphogenetic Proteins / Q000378:metabolism*; D002462:Cell Membrane / Q000201:enzymology* / Q000276:immunology; D002478:Cells, Cultured; D006801:Humans; D009221:Myositis Ossificans / Q000201:enzymology* / Q000503:physiopathology; D010012:Osteogenesis; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis / Q000378:metabolism; D017978:Receptors, Growth Factor / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction
C525809:BMP4 protein, human; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins; D012333:RNA, Messenger; D017978:Receptors, Growth Factor; D017346:Protein Serine-Threonine Kinases; C498211:BMPR1A protein, human; D052005:Bone Morphogenetic Protein Receptors, Type I