Nine mutations in the switch I and switch II regions of human ADP-ribosylation factor 3 (ARF3) were isolated from loss-of-interaction screens, using two-hybrid assays with three different effectors. We then analyzed the ability of the recombinant proteins to (i) bind guanine nucleotides, (ii) activate phospholipase D1 (PLD1), (iii) recruit coatomer (COP-I) to Golgi-enriched membranes, and (iv) expand and vesiculate Golgi in intact cells. Correlations of activities in these assays were used as a means of testing specific hypotheses of ARF action, including the role of PLD1 activation in COP-I recruitment, the role of COP-I in Golgi vesiculation caused by expression of the dominant activating mutant [Q71L]ARF3, and the need for PLD1 activation in Golgi vesiculation. Because we were able to find at least one example of a protein that has lost each of these activities with retention of the others, we conclude that activation of PLD1, recruitment of COP-I to Golgi, and vesiculation of Golgi in cells are functionally separable processes. The ability of certain mutants of ARF3 to alter Golgi morphology without changes in PLD1 activity or COP-I binding is interpreted as evidence for at least one additional, currently unidentified, effector for ARF action at the Golgi.