Slit, the ligand for the Roundabout (Robo) receptors [1-3], is secreted from midline cells of the Drosophila central nervous system (CNS) [4]. It acts as a short-range repellent that controls midline crossing of axons and allows growth cones to select specific pathways along each side of the midline [1, 3]. In addition, Slit directs the migration of muscle precursors and ventral branches of the tracheal system, showing that it provides long-range activity beyond the limit of the developing CNS [2, 5, 6]. Biochemical studies suggest that guidance activity requires cell-surface heparan sulfate to promote binding of mammalian Slit/Robo homologs [7, 8]. Here, we report that the Drosophila homolog of Syndecan (reviewed in [9]), a heparan sulfate proteoglycan (HSPG), is required for proper Slit signaling. We generated syndecan (sdc) mutations and show that they affect all aspects of Slit activity and cause robo-like phenotypes. sdc interacts genetically with robo and slit, and double mutations cause a synergistic strengthening of the single-mutant phenotypes. The results suggest that Syndecan is a necessary component of Slit/Robo signaling and is required in the Slit target cells.