Activated leukocyte cell adhesion molecule modulates neurotrophin signaling

Abstract

Development of the nervous system requires differentiating neurons to grow axons over great distances following long-and short-range signals that are controlled by distinct guidance systems. In general, long-distance acting molecules, such as growth factors, are soluble and bind receptors on the neuronal surface. In contrast, short-range membrane-associated molecules interact with surrounding cells and extracellular matrix, allowing for growth through diverse and changing environments. However, the boundaries between long-range diffusible ligands and short-range contact-mediated factors have blurred. Abbreviations used: ALCAM, activated leukocyte cell adhesion molecule; BSA, bovine serum albumin; CAM, cell adhesion molecule; DMEM, Dulbecco's minimum essential medium; EDL, extensor digitorum longus; GDNF, glial cell-derived neurotrophic factor; H C, carboxy-terminal fragment of tetanus toxin; Ig, immunoglobulin; LAL, levator auris longus; MION, monocrystalline iron oxide nanoparticles; NGF, nerve growth factor; NMJ, neuromuscular junction; p75 NTR, p75 neurotrophin receptor; PBS, phosphate-buffered saline; pERK1/2, phosphorylation of ERK1/2; pTrkA, phosphorylation of TrkA; SDS, sodium dodecyl sulfate; Trk, tropomyosin-receptor-kinase. Abstract Cell adhesion molecules of the immunoglobulin superfamily (IgCAMs) have been shown to modulate growth factor signaling and follow complex trafficking pathways in neurons. Similarly, several growth factors, including members of the neurotrophin family, undergo axonal retrograde transport that is required to elicit their full signaling potential in neurons. We sought to determine whether IgCAMs that enter the axonal retrograde transport route co-operate with neurotrophin signaling. We identified activated leukocyte cell adhesion molecule (ALCAM), a protein involved in axon pathfinding and development of the neuromuscular junction, to be associated with an axonal endocytic compartment that contains neurotrophins and their receptors. Although ALCAM enters carriers that are transported bidirectionally in motor neuron axons, it is predominantly co-transported with the neurotrophin receptor p75 NTR toward the cell body. ALCAM was found to specifically potentiate nerve growth factor (NGF)-induced differentiation and signaling. The extracellular domain of ALCAM is both necessary and sufficient to potentiate NGF-induced neurite outgrowth, and its homodimerization is required for this novel role. Our findings indicate that ALCAM synergizes with NGF to induce neuronal differentiation, raising the possibility that it functions not only as an adhesion molecule but also in the modulation of growth factor signaling in the nervous system.

Rabies Virus Envelope Glycoprotein Targets Lentiviral Vectors to the Axonal Retrograde Pathway in Motor Neurons

“…NTR , and NCAM undergo trafficking in pH-neutral endosomes (22,30,46), and we hypothesize that this neutral environment prevents viral fusion and escape. To verify that the endosomal transport compartment used by RV-G pseudotyped HIV-1 vectors is also pH-neutral, motor neurons were incubated with the acidotropic probe LysoTracker DND for 2 h to label the acidic compartments before addition of RV-G pseudotyped HIV-1 DiO-labeled vectors for 2 h. Under these conditions, only 12% of all particles were positive for LysoTracker, and even when restricted to vectors that colocalize with p75 NTR (endosomal), only 20.74% of particles can be found in acidic compartments (Fig.…”

“…6, D and E). It has recently been shown that NCAM undergoes retrograde transport in the same endosomal carriers as p75 NTR (30); however, it is not known whether the nAChR also undergoes retrograde trafficking. Using MFC cultures, retrograde transport of the labeled antagonist ␣-bungarotoxin was clearly observed (supplemental Fig.…”

“…In contrast to the canonical pathway, however, this maturation is not accompanied by a decrease in the luminal pH, which remains neutral. Interestingly, it has been recently demonstrated that two of the reported receptors for RV-G (p75 NTR and NCAM) undergo retrograde axonal transport in these Rab7-positive non-acidified endosomes (23,30), thus suggesting a potential route of delivery for both wild-type rabies virus and RV-G pseudotyped lentiviral vectors that would allow the vector to reach the nucleus without pH-driven endosomal fusion.…”

Uptake and transport of clostridial neurotoxins

“…Impairments in this process are associated with Alzheimer's disease, Huntington's disease, amyotrophic lateral sclerosis (ALS), and Charcot-Marie Tooth disease [151,158]. To investigate the composition of axonal retrogradely transported signaling endosomes, our laboratory developed a method using H C T coupled to magnetic iron oxide nanobeads (MIONs) for their purification and quantitative analysis by mass spectrometry [126,164].…”

Signaling Endosomes and Growth Cone Motility in Axon Regeneration

“…Finally, several CAMs, including activated leukocyte cell adhesion molecule (ALCAM), are transported in signaling endosomes together with neurotrophins like NGF. ALCAM potentiates neurotrophin signaling raising the possibility that ALCAM functions not only as an adhesion molecule but also in modulating the action of NGF (Wade et al, 2012), suggesting a close bidirectional regulation between neurotrophin and adhesion signaling. In summary, signaling from signaling endosomes not only allows for the spatial restriction of plasma membrane initiated signaling but also enables the assembly of distinct signaling complexes regulating actin dynamics through effector localization and adhesion formation/breakdown and signaling.…”