Abstract
Transforming growth factor (TGF)-βs are dimeric polypeptides that have vital roles in regulating cell growth and differentiation. They signal by assembling a receptor heterotetramer composed of two TβRI:TβRII heterodimers. To investigate whether the two heterodimers bind and signal autonomously, one of the TGF-β protomers was substituted to block receptor binding. The substituted dimer, TGF-β3 WD, bound the TβRII extracellular domain and recruited the TβRI with affinities indistinguishable from TGF-β3, but with one-half the stoichiometry. TGF-β3 WD was further shown to retain one-quarter to one-half the signalling activity of TGF-β3 in three established assays for TGF-β function. Single-molecule fluorescence imaging with GFP-tagged receptors demonstrated a measurable increase in the proportion of TβRI and TβRII dimers upon treatment with TGF-β3, but not with TGF-β3 WD. These results provide evidence that the two TβRI:TβRII heterodimers bind and signal in an autonomous manner. They further underscore how the TGF-βs diverged from the bone morphogenetic proteins, the ancestral ligands of the TGF-β superfamily that signal through a RI:RII:RII heterotrimer.