Axon guidance receptors: Endocytosis, trafficking and downstream signaling from endosomes

During neural development, growing axons express specific surface receptors in response to various environmental guidance cues. These axon guidance receptors are regulated through intracellular trafficking and degradation to enable navigating axons to reach their targets. In Caenorhabditis elegans, the UNC-5 receptor is necessary for dorsal migration of developing motor axons. We previously found that MAX-1 is required for UNC-5–mediated axon repulsion, but its mechanism of action remained unclear. Here, we demonstrate that UNC-5–mediated axon repulsion in C. elegans motor axons requires both max-1 SUMOylation and the AP-3 complex β subunit gene, apb-3. Genetic interaction studies show that max-1 is SUMOylated by gei-17/PIAS1 and acts upstream of apb-3. Biochemical analysis suggests that constitutive interaction of MAX-1 and UNC-5 receptor is weakened by MAX-1 SUMOylation and by the presence of APB-3, a competitive interactor with UNC-5. Overexpression of APB-3 reroutes the trafficking of UNC-5 receptor into the lysosome for protein degradation. In vivo fluorescence recovery after photobleaching experiments shows that MAX-1 SUMOylation and APB-3 are required for proper trafficking of UNC-5 receptor in the axon. Our results demonstrate that SUMOylation of MAX-1 plays an important role in regulating AP-3–mediated trafficking and degradation of UNC-5 receptors during axon guidance.

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Genetic analyses ofRoundabout(ROBO) axon guidance receptors in autism

American Journal of Medical Genetics Part B Neuropsychiatric Genetics ◽

10.1002/ajmg.b.30697 ◽

2008 ◽

Vol 147B (7) ◽

pp. 1019-1027 ◽

Cited By ~ 54

Author(s):

A. Anitha ◽

Kazuhiko Nakamura ◽

Kazuo Yamada ◽

Shiro Suda ◽

Ismail Thanseem ◽

...

Keyword(s):

Axon Guidance ◽

Genetic Analyses ◽

Guidance Receptors

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A role for axon guidance receptors and ligands in blood vessel development and tumor angiogenesis

Cytokine & Growth Factor Reviews ◽

10.1016/j.cytogfr.2005.05.002 ◽

2005 ◽

Vol 16 (4-5) ◽

pp. 535-548 ◽

Cited By ~ 162

Author(s):

Michael Klagsbrun ◽

Anne Eichmann

Keyword(s):

Blood Vessel ◽

Axon Guidance ◽

Tumor Angiogenesis ◽

Blood Vessel Development ◽

Guidance Receptors ◽

Vessel Development

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Minimal structural elements required for midline repulsive signaling and regulation of Drosophila Robo1

10.1101/2020.03.30.016766 ◽

2020 ◽

Cited By ~ 1

Author(s):

Haley E. Brown ◽

Timothy A. Evans

Keyword(s):

Axon Guidance ◽

Biochemical Properties ◽

Structural Elements ◽

Wild Type ◽

Domain Interaction ◽

Robo Receptors ◽

Guidance Receptors ◽

Repulsive Signaling

AbstractThe Roundabout (Robo) family of axon guidance receptors has a conserved ectodomain arrangement of five immunoglobulin-like (Ig) domains plus three fibronectin (Fn) repeats. Based on the strong evolutionary conservation of this domain structure among Robo receptors, as well as in vitro structural and domain-domain interaction studies of Robo family members, this ectodomain arrangement is predicted to be important for Robo receptor signaling in response to Slit ligands. Here, we define the minimal ectodomain structure required for Slit binding and midline repulsive signaling in vivo by Drosophila Robo1. We find that the majority of the Robo1 ectodomain is dispensable for both Slit binding and repulsive signaling. We show that a significant level of midline repulsive signaling activity is retained when all Robo1 ectodomain elements apart from Ig1 are deleted, and that the combination of Ig1 plus one additional ectodomain element (Ig2, Ig5, or Fn3) is sufficient to restore midline repulsion to wild type levels. Further, we find that deleting four out of five Robo1 Ig domains (ΔIg2-5) does not affect negative regulation of Robo1 by Commissureless (Comm) or Robo2, while variants lacking all three fibronectin repeats (ΔFn1-3 and ΔIg2-Fn3) are insensitive to regulation by both Comm and Robo2, signifying a novel regulatory role for Robo1’s Fn repeats. Our results provide an in vivo perspective on the importance of the conserved 5+3 ectodomain structure of Robo receptors, and suggest that specific biochemical properties and/or ectodomain structural conformations observed in vitro for domains other than Ig1 may have limited significance for in vivo signaling in the context of midline repulsion.

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Where does axon guidance lead us?

F1000Research ◽

10.12688/f1000research.10126.1 ◽

2017 ◽

Vol 6 ◽

pp. 78 ◽

Cited By ~ 11

Author(s):

Esther Stoeckli

Keyword(s):

Axon Guidance ◽

Protein Interactions ◽

Neural Circuit ◽

Surface Expression ◽

Target Cells ◽

Protein Protein Interactions ◽

Guidance Receptors ◽

Recent Developments ◽

Genomic Studies ◽

Circuit Formation

During neural circuit formation, axons need to navigate to their target cells in a complex, constantly changing environment. Although we most likely have identified most axon guidance cues and their receptors, we still cannot explain the molecular background of pathfinding for any subpopulation of axons. We lack mechanistic insight into the regulation of interactions between guidance receptors and their ligands. Recent developments in the field of axon guidance suggest that the regulation of surface expression of guidance receptors comprises transcriptional, translational, and post-translational mechanisms, such as trafficking of vesicles with specific cargos, protein-protein interactions, and specific proteolysis of guidance receptors. Not only axon guidance molecules but also the regulatory mechanisms that control their spatial and temporal expression are involved in synaptogenesis and synaptic plasticity. Therefore, it is not surprising that genes associated with axon guidance are frequently found in genetic and genomic studies of neurodevelopmental disorders.

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The Slit-binding Ig1 domain is required for multiple axon guidance activities of Drosophila Robo2

10.1101/2021.05.07.443153 ◽

2021 ◽

Author(s):

LaFreda J. Howard ◽

Marie C. Reichert ◽

Timothy A. Evans

Keyword(s):

Axon Guidance ◽

Lateral Position ◽

Commissural Axons ◽

Lateral Zone ◽

Guidance Receptors ◽

Longitudinal Axon ◽

A Cell ◽

Pathway Formation ◽

Embryonic Neurons

Drosophila Robo2 is a member of the evolutionarily conserved Roundabout (Robo) family of axon guidance receptors. The canonical role of Robo receptors is to signal midline repulsion in response to their cognate Slit ligands, which bind to the N-terminal Ig1 domain in most Robo family members. In the Drosophila embryonic ventral nerve cord, Robo1 and Robo2 cooperate to signal Slit-dependent midline repulsion, while Robo2 also regulates the medial-lateral position of longitudinal axon pathways and acts non-autonomously to promote midline crossing of commissural axons. Although it is clear that Robo2 signals midline repulsion in response to Slit, it is less clear whether Robo2's other activities are also Slit-dependent. To determine which of Robo2's axon guidance roles depend on its Slit-binding Ig1 domain, we have used a CRISPR/Cas9-based strategy replace the endogenous robo2 gene with a robo2 variant from which the Ig1 domain has been deleted (robo2ΔIg1). We compare the expression and localization of Robo2ΔIg1 protein with that of full-length Robo2 in embryonic neurons in vivo, and examine its ability to substitute for Robo2 to mediate midline repulsion and lateral axon pathway formation. We find that removal of the Ig1 domain from Robo2ΔIg1 disrupts both of these axon guidance activities. In addition, we find that the Ig1 domain of Robo2 is required for its proper subcellular localization in embryonic neurons, a role that is not shared by the Ig1 domain of Robo1. Finally, we report that although FasII-positive lateral axons are misguided in embryos expressing Robo2ΔIg1, the axons that normally express Robo2 are correctly guided to the lateral zone, suggesting that Robo2 may guide lateral longitudinal axons through a cell non-autonomous mechanism.

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