Significance: Ypt1 and Ypt31/32 RAB GTPases regulate traffic across the Golgi. Both are activated by TRAPP, an oligomeric GEF. Three TRAPP versions share the same core subunits. TRAPPI and TRAPPIII activate Ypt1. The third, TRAPPII, composed of TRAPPI plus specific subunits, appears to act specifically on Ypt31, but this role has been disputed. By combining the resolving power of fungal genetics with biochemical assays, we establish that the physiological target of TRAPPII is RabE, the Aspergillus Ypt31 ortholog. However, our data suggest that TRAPPII contains independent binding sites for RabE and RabO (Ypt1), possibly explaining its relative lack of discrimination in vitro. TRAPPII arrives at Golgi cisternae preceding their dissipation into carriers, determining the Golgi–to–post-Golgi transition through RabE recruitment.