title: A non-signaling role of Robo2 in tendons is essential for Slit processing and muscle patterning.
abstract: Coordinated locomotion of an organism relies on the development of proper musculoskeletal connections. In Drosophila, the Slit-Robo signaling pathway guides muscles to tendons. Here, we show that the Slit receptor Roundabout 2 (Robo2) plays a non-cell-autonomous role in directing muscles to their corresponding tendons. Robo2 is expressed by tendons, and its non-signaling activity in these cells promotes Slit cleavage, producing a cleaved Slit N-terminal guidance signal that provides short-range signaling into muscles. Consistently, robo2 mutant embryos exhibited a muscle phenotype similar to that of slit, which could not be rescued by muscle-specific Robo2 expression but rather by ectodermally derived Robo2. Alternatively, this muscle phenotype could be induced by tendon-specific robo2 RNAi. We further show that membrane immobilization of Slit or its N-terminal cleaved form (Slit-N) on tendons bypasses the functional requirement for Robo2 in tendons, verifying that the major role of Robo2 is to promote the association of Slit with the tendon cell membrane. Slit-N tends to oligomerize whereas full-length uncleavable Slit does not. It is therefore proposed that Slit-N oligomers, produced at the tendon membrane by Robo2, signal to the approaching muscle by combined Robo1 and Robo3 activity. These findings establish a Robo2-mediated mechanism, independent of signaling, that is essential to limiting Slit distribution and which might be relevant to the regulation of Slit-mediated short-range signaling in additional systems.
authors.affiliation: Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Elly
authors.initials: E
authors.lastname: Ordan
authors.affiliation: Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel talila.volk@weizmann.ac.il.
authors.forename: Talila
authors.initials: T
authors.lastname: Volk
mesh_terms: D000818:Animals; D019521:Body Patterning; D029721:Drosophila Proteins / Q000235:genetics / Q000502:physiology*; D004331:Drosophila melanogaster / Q000196:embryology; D018507:Gene Expression Regulation, Developmental; D055879:Gene Knock-In Techniques; D006579:Heterozygote; D006720:Homozygote; D009132:Muscles / Q000196:embryology*; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000502:physiology*; D010641:Phenotype; D017434:Protein Structure, Tertiary; D034622:RNA Interference; D011971:Receptors, Immunologic / Q000235:genetics / Q000502:physiology*; D015398:Signal Transduction; D013710:Tendons / Q000196:embryology*; D000097905:Roundabout Proteins
chemical_list: D029721:Drosophila Proteins; D009419:Nerve Tissue Proteins; D011971:Receptors, Immunologic; D000097905:Roundabout Proteins; C421243:Robo2 protein, Drosophila; C066819:sli protein, Drosophila
keywords: Drosophila embryo; Muscle; Robo; Slit; Slit cleavage