Previous studies showed that grasshopper semaphorin I, a transmembrane semaphorin, functions in vivo to steer a pair of growth cones, prevent defasciculation, and inhibit branching; and that chick collapsin, a secreted semaphorin, can function in vitro to cause growth cone collapse. Semaphorin II, a secreted semaphorin in Drosophila, is transiently expressed by a single large muscle during motoneuron outgrowth and synapse formation. To test the in vivo function of semaphorin II, we created transgenic Drosophila that generate ectopic semaphorin II expression by muscles that normally do not express it. The results show that semaphorin II can function in vivo as a selective target-derived signal that inhibits the formation of specific synaptic terminal arbors.