Axon Guidance at the Midline: From Mutants to Mechanisms

How axons in the developing nervous system successfully navigate to their correct targets is a fundamental problem in neurobiology. Understanding the mechanisms that mediate axon guidance will give important insight into how the nervous system is correctly wired during development and may have implications for therapeutic approaches to developmental brain disorders and nerve regeneration. Achieving this understanding will require unraveling the molecular logic that ensures the proper expression and localization of axon guidance cues and receptors, and elucidating the signaling events that regulate the growth cone cytoskeleton in response to guidance receptor activation. Studies of axon guidance at the midline of many experimental systems, from the ventral midline of Drosophila to the vertebrate spinal cord, have led to important mechanistic insights into the complex problem of wiring the nervous system. Here we review recent advances in understanding the regulation of midline axon guidance, with a particular emphasis on the contributions made from molecular genetic studies of invertebrate model systems.     

Slit antagonizes netrin-1 attractive effects during the migration of inferior olivary neurons

Inferior olivary neurons (ION) migrate circumferentially around the caudal rhombencephalon starting from the alar plate to locate ventrally close to the floor-plate, ipsilaterally to their site of proliferation. The floor-plate constitutes a source of diffusible factors. Among them, netrin-1 is implied in the survival and attraction of migrating ION in vivo and in vitro. We have looked for a possible involvement of slit-1/2 during ION migration. We report that: (1) slit-1 and slit-2 are coexpressed in the floor-plate of the rhombencephalon throughout ION development; (2) robo-2, a slit receptor, is expressed in migrating ION, in particular when they reach the vicinity of the floor-plate; (3) using in vitro assays in collagen matrix, netrin-1 exerts an attractive effect on ION leading processes and nuclei; (4) slit has a weak repulsive effect on ION axon outgrowth and no effect on migration by itself, but (5) when combined with netrin-1, it antagonizes part of or all of the effects of netrin-1 in a dose-dependent manner, inhibiting the attraction of axons and the migration of cell nuclei. Our results indicate that slit silences the attractive effects of netrin-1 and could participate in the correct ventral positioning of ION, stopping the migration when cell bodies reach the floor-plate.     

Robo1 Forms a Compact Dimer-of-Dimers Assembly

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Roundabout gene family functions during sensory axon guidance in the drosophila embryo are mediated by both Slit-dependent and Slit-independent mechanisms

roundabout (robo) family genes play key roles in axon guidance in a wide variety of animals. We have investigated the roles of the robo family members, robo, robo2, and robo3, in the guidance of sensory axons in the Drosophila embryo. In robo(-/-), slit(-/-), and robo(-/+) slit(-/+) mutants, lateral cluster sensory neurons misproject to cells and axons in the nearby ventral' (v') cluster. These phenotypes, together with the normal expression pattern of Slit and Robo, suggest that Slit ligand secreted from the epidermis interacts with Robo receptors on lateral cluster sensory growth cones to limit their exploration of nearby attractive substrates. The most common sensory axon phenotype seen in robo2(-/-) mutants was misprojection of dorsal cluster sensory axons away from their normal growth substrate, the transverse connective of the trachea. slit appears to play no role in this aspect of sensory axon growth. Robo2 is expressed, not on the dorsal sensory axons, but on the transverse connective. These results suggest a novel, non-cell-autonomous mechanism for axon guidance by robo family genes: Robo2 expressed on the trachea acts as an attractant for the dorsal sensory growth cones.     

Unidirectional startle responses and disrupted left–right co-ordination of motor behaviors in robo3 mutant zebrafish

The Roundabout (Robo) family of receptors and their Slit ligands play well-established roles in axonal guidance, including in humans where horizontal gaze palsy with progressive scoliosis (HGPPS) is caused by mutations in the robo3 gene. Although significant progress has been made toward understanding the mechanism by which Robo receptors establish commissural projections in the central nervous system, less is known about how these projections contribute to neural circuits mediating behavior. In this study, we report cloning of the zebrafish behavioral mutant twitch twice and show that twitch twice encodes robo3 . We show that in mutant hindbrains the axons of an identified pair of neurons, the Mauthner cells, fail to cross the midline. The Mauthner neurons are essential for the startle response, and in twitch twice / robo3 mutants misguidance of the Mauthner axons results in a unidirectional startle response. Moreover, we show that twitch twice mutants exhibit normal visual acuity but display defects in horizontal eye movements, suggesting a specific and critical role for twitch twice / robo3 in sensory-guided behavior.