Characterization of transforming growth factor-beta (TGF-beta) receptors on BeWo choriocarcinoma cells including the identification of a novel 38-kDa TGF-beta binding glycoprotein.
Transforming growth factor-beta (TGF-beta) is a potential mediator of placental trophoblast functions, including differentiation, hormone production, endometrial invasion, and immunosuppression. Equilibrium binding and affinity-labeling assays were used to investigate the binding characteristics of TGF-beta 1 and TGF-beta 2 on an established human choriocarcinoma trophoblastic cell line (BeWo). The equilibrium binding experiments indicated that the BeWo cells exhibited similar average affinities and total number of binding sites for TGF-beta 1 and TGF-beta 2. The Kd values obtained from Scatchard analyses were approximately 65 pM for 125I-TGF-beta 1 and approximately 40 pM for 125I-TGF-beta 2, with 70,000 and 85,000 sites per cell, respectively. Competitive equilibrium binding experiments indicated that TGF-beta 1 and TGF-beta 2 were equipotent (apparent half maximal inhibition [IC50] approximately 70 pM) and that all binding sites were capable of recognizing both isoforms. Affinity-labeling studies with 125I-TGF-beta 1 and 125I-TGF-beta 2 and the chemical cross-linking agent bis(sulfosuccinimidyl)suberate (BS3) revealed a predominant type III/betaglycan receptor, a low level of apparently heterogeneous type I and II receptors and an additional novel 38-kDa TGF-beta binding glycoprotein that was present both under reducing and nonreducing conditions on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Affinity-labeling saturation and competition studies indicated that the type III/betaglycan component appears to have a 7-fold higher capacity for TGF-beta 1 than for -beta 2 yet exhibits a 5- to 10-fold higher affinity for TGF-beta 2 than for -beta 1. The 38-kDa TGF-beta binding component, an N-linked glycoprotein, exhibits a higher affinity for TGF-beta 2 than for -beta 1 that is strikingly similar to that of the type III/betaglycan receptor. This 38-kDa binding protein appears to be upregulated after methotrexate-induced differentiation of the BeWo cells.
Mitchell
E J
EJ
Biotechnology Research Institute, National Research Council, Canada, Montreal, Quebec.
Lee
K
K
O'Connor-McCourt
M D
MD
D000345:Affinity Labels; D001665:Binding Sites; D001667:Binding, Competitive; D002454:Cell Differentiation; D002822:Choriocarcinoma / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008727:Methotrexate / Q000494:pharmacology; D008970:Molecular Weight; D011956:Receptors, Cell Surface / Q000378:metabolism*; D018125:Receptors, Transforming Growth Factor beta; D013237:Stereoisomerism; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D014327:Trophoblasts / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D000345:Affinity Labels; D011956:Receptors, Cell Surface; D018125:Receptors, Transforming Growth Factor beta; D016212:Transforming Growth Factor beta; D008727:Methotrexate