The evolutionarily conserved Semaphorin family proteins are well known axon guidance ligands that mediate both attractive and repulsive responses in invertebrates and vertebrates. In this study, we show that theDrosophilaSemaphorin-1a (Sema1a), a transmembrane Semaphorin, is required cell autonomously in adult photoreceptor (R-cell) axons for the establishment of an appropriate topographic termination pattern in the optic lobe. Loss ofsema1adisrupts the association of neighboring R-cell growth cones leading to defects in local termination pattern, whereas overexpression ofsema1ainduces the hyper-fasciculation of R-cell axons. The function of Sema1a in R-cell axon guidance absolutely requires its cytoplasmic domain. We propose that Sema1a functions as a receptor in regulating R-cell axon guidance in theDrosophilavisual system.