Trafficking guidance receptors

Abstract

Wiring of the brain relies initially on the correct outgrowth of axons to reach the appropriate target area for innervation. A large number of guidance receptors present in the plasma membrane of axonal growth cones and elsewhere on the neuron read and execute directional cues present in the extracellular environment of the navigating growth cone. The exact timing, levels, and localization of expression of the guidance receptors in the plasma membrane therefore determine the outcome of guidance decisions. Many guidance receptors are localized in exquisitely precise spatial and temporal patterns. The cellular mechanisms ensuring these localization patterns include spatially accurate sorting after synthesis in the secretory pathway, retrieval of inappropriately expressed receptors by endocytosis followed by degradation or recycling, and restriction of diffusion. This article will discuss the machinery and regulation underlying the restricted distribution of membrane receptors, focusing on the currently best-studied example, the L1 cell adhesion molecule. In addition to the long-range mechanisms ensuring appropriate localization, the same mechanisms can act locally to adjust levels and localization of receptors. These local mechanisms are regulated by ligand binding and subsequent activation of local signaling cascades. It is likely that the localization of all guidance receptors is regulated by a combination of sorting, retrieval, recycling and retention, similar to the ones we discuss here for L1.

Figure 1.

Endosomal compartments involved in NgCAM transcytosis to the axon. NgCAM is endocytosed into EEA1-containing early endosomes with transferrin. Transferrin (Tf) recycles from the early endosome whereas NgCAM traverses NEEP21-positive early endosomes and reaches recycling endosomes. Transferrin is also found in recycling endosomes. In recycling endosomes, transferrin and NgCAM occupy partially overlapping domains and are ultimately sorted away from each other. NgCAM then enters axons in small motile endosomally derived carriers and is transported anterogradely via fast axonal transport. Stationary endosomes containing NgCAM along axons provide stations from which small carriers can likely bud or fuse.

Figure 2.

Regulation of NgCAM transcytotic routing by spatial regulation of phosphorylation of the adaptor binding site (based on Anderson et al. 2005). NgCAM is sorted to the basolateral domain based on a tyrosine-based motif (YRSLE) in its cytoplasmic tail. This signal is recognized by the basolateral sorting adaptor AP1B. The site of action of AP1B-based sorting is either the TGN or the recycling endosome. NgCAM is then exocytosed on the basolateral plasma membrane. The clathrin adaptor AP2 binds to the YRSLE motif and mediates endocytosis into endosomes. At some point after endocytosis, the YRSLE is phosphorylated by a src family kinase. This phosphorylation event prevents binding of AP1B and thereby redirects the endocytosed NgCAM away from basolateral recycling into an apical-directed transcytotic route. In the presence of the kinase inhibitor herbimycin A, NgCAM accumulates on the basolateral, instead of the apical, domain. An analogous mechanism likely operates for NgCAM in neurons.

Figure 3.

Three axonal transport pathways. Transmembrane receptors can accumulate on the axonal surface by three mechanisms:

1. They can be directly sorted to the axon from the TGN based on sorting signals and association with axonally directed motor proteins (orange pathway: direct sorting).

2. They can be sorted in the TGN to the somatodendritic domain and be redirected to the axon after endocytosis (purple pathway: transcytosis).

3. They can be uniformly inserted into the plasma membrane in the axonal and somatodendritic domains and then be removed preferentially from the somatodendritic domain and stabilized in the axon (green pathway: domain-specific retrieval/retention).

Figure 4.

Local regulation of L1 surface levels at the growth cone. L1 ligation via L1 homophilic binding leads to diffusional trapping of surface L1 at the contact site and mobilization and exocytosis of L1 from vesicular pools. Endocytic retrieval from the rear of the growth cone maintains a gradient of L1 across the growth cone and provides a pool of vesicular L1 available to be exocytosed locally.