Transforming growth factor- (TGF-) plays a causal role in several human pathologies including fibrotic diseases and metastasis. TGF- signaling is mediated through its interaction with three types of cell surface receptors, RI, RII, and RIII. The soluble ectodomains of RII and RIII bind to TGF-, making them attractive candidates to sequester TGF- and inhibit its activity. To optimize the activity of the ectodomains, we studied the effect of artificially dimerizing them upon their kinetics of binding to TGF- using an optical biosensor and studied their antagonistic potencies using an in vitro signaling assay. We fused the RII ectodomain and the membrane-proximal ligand-binding domain of the RIII ectodomain to de novo designed heterodimerizing coil strands and demonstrated that the coil strands within the fusion proteins were capable of promoting the dimerization of the coil-tagged ectodomains. Our results indicate that coiled coil-induced dimerization of the ectodomains stabilized their interaction with TGF- as compared with the monomeric ectodomains. Also, in contrast to the monomeric ectodomains, which did not block signaling, the coiled coil-induced dimers were characterized by antagonistic potencies in the low nanomolar range.Mammalian transforming growth factor- isoforms (TGF-1, -2, and -3) 1 mediate signaling by binding to and complexing three types of cell surface receptors known as the TGF- type I (RI), type II (RII), and type III (RIII) receptors (1). In the absence of ligand, both RI and RII can form homooligomers (2). TGF-1 and TGF-3 bind simultaneously to two RII ectodomains (RIIEDs) (3). This binding event is thought to reorient the RIIs at the cell surface, allowing for the formation of a signaling-competent RI-RII complex (4). RIII was first thought to be an "accessory" receptor whose role is to present TGF-2 to RI and RII (5). Recent studies suggest, however, that the role of RIII is more complex. Firstly, it was demonstrated that RIII is required for TGF-1-promoted mesenchymal transformation during chick embryonic heart development (6). Furthermore, it was shown that the RIII cytoplasmic domain is important for signaling (7) and that the role of RIII in signaling may be modulated by its glycosylation state (8). Two independent TGF--binding domains have been identified within the RIII ectodomain (9 -11), further emphasizing the complexity of this receptor.TGF- is involved in the regulation of many physiological processes including cell growth and differentiation. TGF- overexpression plays a key role in several human pathologies including fibrotic diseases (12) and metastasis (13). Recently, two reports provided encouraging results showing that a TGF- antagonist, corresponding to the RIIED artificially dimerized through the Fc portion of an antibody (RIIED-Fc), reduces metastasis in vivo without any adverse side effects (14, 15). Other soluble TGF--binding proteins, including the ectodomain of RIII, have also been evaluated for their antagonistic potency (16 -19). We reported th...