Ephs and ephrins are cell surface receptors that bind to each other and initiate distinct, bidirectional signalling pathways in processes known as forward (Eph) and reverse (ephrin) signalling. Previous work had shown that the loss of ephrinB1 protein alone or compound loss of EphB2 and EphB3 leads to cleft palate. Because of the bidirectional signalling capability of these molecules, it was not clear whether forward or reverse signalling caused the cleft palate in the ephrinB1 protein null or EphB2 and EphB3 compound null mice. We demonstrate that forward signalling is essential for palatogenesis. Foetuses with a cytoplasmically truncated EphB2 protein, which could initiate reverse but not forward signalling, and were protein null for EphB3 had a cleft palate. This happened because their palatal shelves, which could elevate in vivo and adhere and fuse in culture, were too small to contact one another. Small shelf size was due to reduced proliferation in the palatal mesenchyme. The reduced proliferation was not the result of abnormal vascular development within the palate. In conclusion, strong evidence is provided for specific and co-operative roles of EphB2 and EphB3 in palate development.