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 the Drosophila Semaphorin-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 of sema1a disrupts the association of neighboring R-cell growth cones leading to defects in local termination pattern, whereas overexpression of sema1a induces 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 the Drosophila visual system.