Rapid Flux in Transforming Growth Factor-β Receptors on Bone Cells

Abstract

The proportion of transforming growth factor-␤ (TGF-␤) binding among conventional membrane receptors on bone cells can vary with hormone or growth factor treatment or with the state of osteoblast-like activity and appears to determine the nature of its biological effects. Therefore, functional TGF-␤ receptor stability could be an important aspect of regulation. Suppression of protein synthesis reduced TGF-␤ binding to types I and II receptors with t 1/2 of 2 h and to betaglycan with t 1/2 of 6 h. In contrast, suppression of mRNA transcription reduced TGF-␤ binding at least 3-fold more slowly at each receptor site. Preexposure to TGF-␤ decreased its binding at all three sites within 4 h in osteoblast-enriched cultures. This effect was transient with lower TGF-␤ concentrations, where the receptor profile was nearly fully restored within 24 -48 h. In contrast, less differentiated bone cells were less sensitive to ligand-dependent receptor down-regulation. Agents that alter protein kinase and phosphatase activity also modified the TGF-␤ binding profile in specific ways. Together, these results indicate that cell surface TGF-␤ receptors turn over rapidly by ligand-independent and ligand-dependent mechanisms, demonstrate that the binding capacity of TGF-␤ receptors is less stable than their mRNAs, and that functional receptor levels may be determined in part by post-transcriptional events.Transforming growth factor type ␤s (TGF-␤s) 1 are potent, ubiquitous growth regulators that are abundant in skeletal tissue. Their expression is controlled in temporal and spatial ways during skeletal tissue development, and they produce complex stimulatory and inhibitory changes in osteoblast function in vitro. The presence of TGF-␤s in bone and their importance as local skeletal growth regulators were first described in fetal rat calvariae, and cells derived from various bone tissues have provided much information about TGF-␤ activity and receptor expression during osteogenesis (reviewed in Ref. 1).Similar to cells found in many tissues, fetal rat bone cells express type I (53 kDa) and type II (73 kDa) glycoprotein receptors, as well as type III proteoglycans (Ͼ250 betaglycans) that could directly or indirectly influence TGF-␤ binding to the signaling types I and II sites (2-5). In this model, the association of TGF-␤ with each receptor varies with the basal state of bone cell differentiation, and relative mRNA abundance levels generally reflect the cell surface binding profiles found in untreated cells (5). Rapid changes in the proportion of TGF-␤ binding among receptors, induced by certain growth factors and hormones, are consistent with specific changes in its biological effects (5-8). For example, glucocorticoid decreases TGF-␤ binding to types I and II receptors, increases its binding to type III sites, and reduces its overall effectiveness (6). In contrast, bone morphogenetic protein (BMP)-2 reduces TGF-␤ binding to type II receptors and betaglycan, whereas it enhances binding to type I receptors. After BMP-2 treatment, the m...

Rescue of coronal suture fusion using transforming growth factor‐beta 3 (Tgf‐β3) in rabbits with delayed‐onset craniosynostosis

“…Opperman and colleagues recently showed that small doses of Tgf-␤3 (3-30 ng) were associated with a reduced number of suture fibroblasts that were immunoreactive for T␤r-I in rat IF sutures both in vivo and in vitro . Since Tgf-␤2 and Tgf-␤3 share the same set of transmembrane receptors and the same intracellular Smads (Centrella et al, 1996), downregulation of T␤r-I expression by Tgf-␤3 may limit receptor access, thereby reducing the mitogenic and osteogenic effects of Tgf-␤2. This may explain, in part, the antagonistic effects of these two isoforms in regulating suture morphogenesis and fusion (Opperman et al, 2000;Opperman and Ogle, 2002).…”

Endocytic regulation of TGF-β signaling

“…For instance, H-8, an inhibitor of cAMP-dependent protein kinase and cGMP-dependent protein kinase, and staurosporine, a potent protein kinase C inhibitor, have been shown to enhance TGF-b binding to its receptors, whereas phosphatase inhibitors okadaic acid, vanadate and fluoride decrease the binding [83], indicating that the turnover of TGF-b receptors is modulated by other signaling events.…”

“…Several studies have examined the half-life of TGF-b receptors and yielded different results. Using the 125 I-TGF-b1-binding assay, Centrella et al [83] found that the half-life of cell surface TbRI and TbRII is about 2 h in primary osteoblasts. However, the half-life of the newly synthesized mature (endoglycosidase H-resistant) form of TbRII in MvlLu lung epithelial cells, as analyzed by 35 S-Met metabolic labeling, varied from about 1 h [84] to 2.5 h [85].…”

Ligand-induced downregulation of receptors for TGF-beta in human osteoblast-like cells from adult donors

“…Accordingly, there is circumstantial evidence that betaglycan in its soluble form might act as a potential antagonist of TGF-, whereas in a membraneassociated form it may enhance the binding of TGF-to the signalling receptors I and II (López-Casillas et al 1994. Other regulatory mechanisms are suggested by a set of experiments which show that a rapid turnover of T Rs -I, -II and -III by ligand-independent and ligand-dependent mechanisms are observed in nonhuman, fetal cell culture systems (Centrella et al 1996). The present study was undertaken to investigate the influence of TGF-1 on the cell-surface expression of its own receptors in cultured human osteoblast-like cells from adult donors.…”