During the early stages of heart development, progenitors for the heart valves and septa come from endothelial cells via a developmental process known as “epithelial–mesenchymal cell transformation.” This process is restricted to the atrioventricular (AV) canal and outflow tract portions of the embryonic heart. TGFβ signal transduction pathways play critical roles during epithelial–mesenchymal cell transformation in heart development. Previously, we showed that both TGFβ Type II (TβRII) and Type III (TβRIII) receptors are required to mediate epithelial mesenchymal cell transformation in chick heart. Further, distinct TGFβ2 and TGFβ3 activities correspond to separate components of the embryonic cell transformation process. Studies by others of TGFβ‐mediated inhibition of cell proliferation produced a model where TβRIII functions by facilitating TGFβ2 binding to TβRII. In the present study, we provide evidence that TβRIII mediates distinct cellular responses from those of TβRII. Blocking antibody for TβRIII, but not antibody against TβRII, specifically inhibits the endothelial cell–cell separation step. Examination of developmental markers, perturbed by blocking TβRIII antibody, revealed a pattern of expression distinctively different from that of TβRII antibody treatment. These data show that a distinct TβRIII‐mediated process is required for endothelial cell–cell separation during epithelial mesenchymal cell transformation. As TGFβ2 mediates endothelial cell–cell separation, the data point to a specific association of TGFβ2 and TβRIII in the cell separation step of epithelial mesenchymal cell transformation. We conclude that distinct TβRII and TβRIII signal transduction pathways mediate epithelial–mesenchymal cell transformation in the heart. © 2001 Wiley‐Liss, Inc.