Skeletal‐specific expression of Fgd1 during bone formation and skeletal defects in faciogenital dysplasia (FGDY; Aarskog syndrome)

Abstract

FGD1 encodes a guanine nucleotide exchange factor (GEF) that specifically activates the Rho GTPase Cdc42; FGD1 mutations result in Faciogenital Dysplasia (FGDY, Aarskog syndrome), an X-linked developmental disorder that adversely affects the formation of multiple skeletal structures. To further define the role of FGD1 in skeletal development, we examined its expression in developing mouse embryos and correlated this pattern with FGDY skeletal defects. In this study, we show that Fgd1, the mouse FGD1 ortholog, is initially expressed during the onset of ossification during embryogenesis. Fgd1 is expressed in regions of active bone formation in the trabeculae and diaphyseal cortices of developing long bones. The onset of Fgd1 expression correlates with the expression of bone sialo-protein, a protein specifically expressed in osteoblasts at the onset of matrix mineralization; an analysis of serial sections shows that Fgd1 is expressed in tissues containing calcified and mineralized extracellular matrix. Fgd1 protein is specifically expressed in cultured osteoblast and osteoblast-like cells including MC3T3-E1 cells and human osteosarcoma cells but not in other mesodermal cells; immunohistochemical studies confirm the presence of Fgd1 protein in mouse calvarial cells. Postnatally, Fgd1 is expressed more broadly in skeletal tissue with expression in the perichondrium, resting chondrocytes, and joint capsule fibroblasts. The data indicate that Fgd1 is expressed in a variety of regions of incipient and active endochondral and intramembranous ossification including the craniofacial bones, vertebrae, ribs, long bones and phalanges. The observed pattern of Fgd1 expression correlates with FGDY skeletal manifestations and provides an embryologic basis for the prevalence of observed skeletal defects. The observation that the induction of Fgd1 expression coincides with the initiation of ossification strongly suggests that FGD1 signaling plays a role in ossification and bone formation; it also suggests that FGD1 signaling does not play a role in the earlier phases of skeletogenesis. With the observation that FGD1 mutations result in the skeletal dysplasia FGDY, accumulated data indicate that FGD1 signaling plays a critical role in ossification and skeletal development.

Faciogenital Dysplasia Protein (FGD1) Regulates Export of Cargo Proteins from the Golgi Complex via Cdc42 Activation

“…On the basis of its subcellular localization, FGD1 was suggested to participate in both membrane transport and actin remodeling (Estrada et al, 2001), two processes that have fundamental roles in tissue biogenesis. Moreover, use of in situ hybridization has shown that FGD1 mRNA is up-regulated during bone development (and to a lesser extend in other connective tissues), together with a number of proteins that are involved in bone morphogenesis (Gorski et al, 2000). Here, we have shown that FGD1 is required for effective transport of cargo proteins from the Golgi complex to the plasma membrane in general, whereas in osteoblasts it regulates the secretion of PC-I, osteocalcin, osteonectin, and osteopontin.…”

“…Apparently this can be explained by a consideration of the expression profile of FGD1: although FGD1 can be detected by Western blotting in cell lines of non-bone origin (e.g., HeLa cells), in situ some tissues (and bone especially) show more FGD1 expression than others (Gorski et al, 2000). FGDY manifestations in the heart and in the urogenital system have also been reported (Scott, 1971) and would correspond to the moderate FGD1 expression seen in these tissues during development (Pasteris et al, 1994).…”

“…Postnatally, Fgd1 mRNA has been detected more broadly in skeletal tissue, with a higher signal in the perichondrium, resting chondrocytes and joint capsule fibroblasts. Thus, this pattern of Fgd1 expression correlates with the FGDY skeletal manifestations (Gorski et al, 2000), which themselves define most of the other FGDY-related problems.…”

“…Up-regulation of Fgd1 correlates with an increase in osteopontin, a protein that is specifically expressed in osteoblasts at the onset of matrix mineralization (Gorski et al, 2000).…”

“…Indeed, because polarized secretion of specific proteins is required for correct bone morphogenesis (including collagen-I, osteocalcin, osteopontin, and others; Leblond, 1989) and because the expression of FGD1 strongly correlates with the expression of proteins that are essential for bone development (Gorski et al, 2000), FGD1 appears to have a role in this process Thus, the characterization of FGD1 involvement in the regulation of membrane transport represents an essential task that should have significant importance for our understanding of the pathogenesis of FGDY.…”