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Axonal survival and growth requires signalling from tropomyosin receptor kinases (Trks). To transmit their signals, receptor–ligand complexes are endocytosed and undergo retrograde trafficking to the soma, where downstream signalling occurs. Vesicles transporting neurotrophic receptors to the soma are reported to be Rab7-positive late endosomes and/or multivesicular bodies (MVBs), where receptors localize within so-called intraluminal vesicles (herein Rab7 corresponds to Rab7A unless specified otherwise). Therefore, one challenging question is how downstream signalling is possible given the insulating properties of intraluminal vesicles. In this study, we report that Rab7-positive endosomes and MVBs retrieve TrkA (also known as NTRK1) through tubular microdomains. Interestingly, this phenotype is absent for the EGF receptor. Furthermore, we found that endophilinA1, endophilinA2 and endophilinA3, together with WASH1 (also known as WASHC1), are involved in the tubulation process. In Charcot–Marie–Tooth disease 2B (CMT2B), a neuropathy of the peripheral nervous system, this tubulating mechanism is disrupted. In addition, the ability to tubulate correlates with the phosphorylation levels of TrkA as well as with neurite length in neuronal cultures from dorsal root ganglia. In all, we report a new retrieval mechanism of late Rab7-positive endosomes, which enables TrkA signalling and sheds new light onto how neurotrophic signalling is disrupted in CMT2B. This article has an associated First Person interview with the first author of the paper.

Neural circuit development involves the coordinated growth and guidance of axons. During this process, axons encounter many different cues, but how these cues are integrated and translated into growth is poorly understood. In this study, we report that receptor signaling does not follow a linear path but changes dependent on developmental stage and coreceptors involved. Using developing chicken embryos of both sexes, our data show that calcium-sensing receptor (CaSR), a G-protein-coupled receptor important for regulating calcium homeostasis, regulates neurite growth in two distinct ways. First, when signaling in isolation, CaSR promotes growth through the PI3-kinase-Akt pathway. At later developmental stages, CaSR enhances tropomyosin receptor kinase B (TrkB)/BDNF-mediated neurite growth. This enhancement is facilitated through a switch in the signaling cascade downstream of CaSR (i.e., from the PI3-kinase-Akt pathway to activation of GSK3α Tyr279). TrkB and CaSR colocalize within late endosomes, cotraffic and coactivate GSK3, which serves as a shared signaling node for both receptors. Our study provides evidence that two unrelated receptors can integrate their individual signaling cascades toward a nonadditive effect and thus control neurite growth during development. SIGNIFICANCE STATEMENT This work highlights the effect of receptor coactivation and signal integration in a developmental setting. During embryonic development, neurites grow toward their targets guided by cues in the extracellular environment. These cues are sensed by receptors at the surface that trigger intracellular signaling events modulating the cytoskeleton. Emerging evidence suggests that the effects of guidance cues are diversified, therefore expanding the number of responses. Here, we show that two unrelated receptors can change the downstream signaling cascade and regulate neuronal growth through a shared signaling node. In addition to unraveling a novel signaling pathway in neurite growth, this research stresses the importance of receptor coactivation and signal integration during development of the nervous system.

Amyotrophic lateral sclerosis (ALS) is a progressive, adult-onset neurodegenerative disease caused by degeneration of motor neurons in the brain and spinal cord leading to muscle weakness. Median survival after symptom onset in patients is 3-5 years and no effective therapies are available to treat or cure ALS. Therefore, further insight is needed into the molecular and cellular mechanisms that cause motor neuron degeneration and ALS. Different ALS disease mechanisms have been identified and recent evidence supports a prominent role for defects in intracellular transport. Several different ALS-causing gene mutations (e.g., in FUS, TDP-43, or C9ORF72) have been linked to defects in neuronal trafficking and a picture is emerging on how these defects may trigger disease. This review summarizes and discusses these recent findings. An overview of how endosomal and receptor trafficking are affected in ALS is followed by a description on dysregulated autophagy and ER/Golgi trafficking. Finally, changes in axonal transport and nucleocytoplasmic transport are discussed. Further insight into intracellular trafficking defects in ALS will deepen our understanding of ALS pathogenesis and will provide novel avenues for therapeutic intervention. peerReviewed

Forgetting and receptor removal The trafficking of AMPA receptors to and from the surface of postsynaptic membranes regulates synaptic strength and underlies learning and memory. Awasthi et al. found that the integral membrane protein synaptotagmin-3 (Syt3) is predominantly found on postsynaptic endocytic zones of neurons, where it promotes AMPA receptor internalization (see the Perspective by Mandelberg and Tsien). In Syt3 overexpressing or knockdown neurons, synaptic transmission and short-term plasticity were unchanged. However, in neurons from Syt3 knockout mice, synaptic long-term depression was abolished and decaying long-term potentiation endured. In Syt3 knockout mice, spatial learning was unaltered; however, these animals showed signs of impaired forgetting and relearning during the water maze spatial memory task. Science , this issue p. eaav1483 ; see also p. 31

During the development of the nervous system, axons extend through complex environments. Growth cones at the axon tip allow axons to find and innervate their appropriate targets and form functional synapses. Axon pathfinding requires axons to respond to guidance signals and these cues need to be detected by specialized receptors followed by intracellular signal integration and translation. Several downstream signaling pathways have been identified for axon guidance receptors and it has become evident that these pathways are often initiated from intracellular vesicles called endosomes. Endosomes allow receptors to traffic intracellularly, re-locating receptors from one cellular region to another. The localization of axon guidance receptors to endosomal compartments is crucial for their function, signaling output and expression levels. For example, active receptors within endosomes can recruit downstream proteins to the endosomal membrane and facilitate signaling. Also, endosomal trafficking can re-locate receptors back to the plasma membrane to allow re-activation or mediate downregulation of receptor signaling via degradation. Accumulating evidence suggests that axon guidance receptors do not follow a pre-set default trafficking route but may change their localization within endosomes. This re-routing appears to be spatially and temporally regulated, either by expression of adaptor proteins or co-receptors. These findings shed light on how signaling in axon guidance is regulated and diversified - a mechanism which explains how a limited set of guidance cues can help to establish billions of neuronal connections. In this review, we summarize and discuss our current knowledge of axon guidance receptor trafficking and provide directions for future research.

The sorting of activated receptors into distinct endosomal compartments is essential to activate specific signaling cascades and cellular events including growth and survival. However, the proteins involved in this sorting are not well understood. We discovered a novel role of EndophilinAs in sorting of activated BDNF-TrkB receptors into late endosomal compartments. Mice lacking all three EndophilinAs accumulate Rab7-positive late endosomes. Moreover, EndophilinAs are differentially localized to, co-traffic with, and tubulate, distinct endosomal compartments: In response to BDNF, EndophilinA2 is recruited to both early and late endosomes, EndophilinA3 is recruited to Lamp1-positive late endosomes, and co-trafficks with Rab5 and Rab7 in both the presence and absence of BDNF, while EndophilinA1 colocalizes at lower levels with endosomes. The absence of all three EndophilinAs caused TrkB to accumulate in EEA1 and Rab7-positive endosomes, and impaired BDNF-TrkB-dependent survival signaling cascades. In addition, EndophilinA triple knockout neurons exhibited increased cell death which could not be rescued by exogenous BDNF, in a neurotrophin-dependent survival assay. Thus, EndophilinAs differentially regulate activated receptor sorting via distinct endosomal compartments to promote BDNF-dependent cell survival.

International audience; Many organs, such as lungs, nerves, blood and lymphatic vessels, consist of complex networks that carry flows of information, gases, and nutrients within the body. The morphogenetic patterning that generates these organs involves the coordinated action of developmental signaling cues that guide migration of specialized cells. Precision guidance of endothelial tip cells by vascular endothelial growth factors (VEGFs) is well established, and several families of neural guidance molecules have been identified to exert guidance function in both the nervous and the vascular systems. This review discusses recent advances in VEGF research, focusing on the emerging role of neural guidance molecules as key regulators of VEGF function during vascular development and on the novel role of VEGFs in neural cell migration and nerve wiring.

Dendritic spines compartmentalize information in the brain, and their morphological characteristics are thought to underly synaptic plasticity. Here we identify copine-6 as a novel modulator of dendritic spine morphology. We found that brain-derived neurotrophic factor (BDNF) - a molecule essential for long-term potentiation of synaptic strength - upregulated and recruited copine-6 to dendritic spines in hippocampal neurons. Overexpression of copine-6 increased mushroom spine number and decreased filopodia number, while copine-6 knockdown had the opposite effect and dramatically increased the number of filopodia, which lacked PSD95. Functionally, manipulation of post-synaptic copine-6 levels affected miniature excitatory post-synaptic current (mEPSC) kinetics and evoked synaptic vesicle recycling in contacting boutons, and post-synaptic knockdown of copine-6 reduced hippocampal LTP and increased LTD. Mechanistically, copine-6 promotes BDNF-TrkB signaling and recycling of activated TrkB receptors back to the plasma membrane surface, and is necessary for BDNF-induced increases in mushroom spines in hippocampal neurons. Thus copine-6 regulates BDNF-dependent changes in dendritic spine morphology to promote synaptic plasticity.

In the adult forebrain, new interneurons are continuously generated and integrated into the existing circuitry of the olfactory bulb (OB). In an attempt to identify signals that regulate this synaptic integration process, we found strong expression of agrin in adult generated neuronal precursors that arrive in the olfactory bulb after their generation in the subventricular zone. While the agrin receptor components MuSK and Lrp4 were below detection level in neuron populations that represent synaptic targets for the new interneurons, the alternative receptor α3-Na+K+-ATPase was strongly expressed in mitral cells. Using a transplantation approach, we demonstrate that agrin-deficient interneuron precursors migrate correctly into the OB. However, in contrast to wild-type neurons, which form synapses and survive for prolonged periods, mutant neurons do not mature and are rapidly eliminated. Usingin vivobrain electroporation of the olfactory system, we show that the transmembrane form of agrin alone is sufficient to mediate integration and demonstrate that excess transmembrane agrin increases the number of dendritic spines. Last, we providein vivoevidence that an interaction between agrin and α3-Na+K+-ATPase is of functional importance in this system.

It has long been known that the secreted factor Reelin is crucial for proper neuronal migration during the development of the cortex. However, the mechanistic actions leading to a Reelin response are less well known. Acker-Palmer and colleagues reveal that the axonal guidance cue ephrin B2 acts as a co-factor with Reelin receptors to initiate signalling cascades important to produce the proper lamination of the cortex. This additional role for ephrin B2 expands the functions for this developmental protein beyond proper axonal targeting to cell migration. Coordinated migration of neurons in the developing and adult brain is essential for its proper function. The secreted glycoprotein Reelin (also known as RELN) guides migration of neurons by binding to two lipoprotein receptors, the very-low-density lipoprotein receptor (VLDLR) and apolipoprotein E receptor 2 (ApoER2, also known as LRP8)1. Loss of Reelin function in humans results in the severe developmental disorder lissencephaly2 and it has also been associated with other neurological disorders such as epilepsy, schizophrenia and Alzheimer’s disease3. The molecular mechanisms by which Reelin activates its receptors and controls cellular functions are largely unknown. Here we show that the neuronal guidance cues ephrin B proteins are essential for Reelin signalling during the development of laminated structures in the brain. We show that ephrin Bs genetically interact with Reelin. Notably, compound mouse mutants (Reln+/−; Efnb3−/− or Reln+/−; Efnb2−/−) and triple ephrin B1, B2, B3 knockouts show neuronal migration defects that recapitulate the ones observed in the neocortex, hippocampus and cerebellum of the reeler mouse. Mechanistically, we show that Reelin binds to the extracellular domain of ephrin Bs, which associate at the membrane with VLDLR and ApoER2 in neurons. Clustering of ephrin Bs leads to the recruitment and phosphorylation of Dab1 which is necessary for Reelin signalling. Conversely, loss of function of ephrin Bs severely impairs Reelin-induced Dab1 phosphorylation. Importantly, activation of ephrin Bs can rescue the reeler neuronal migration defects in the absence of Reelin protein. Together, our results identify ephrin Bs as essential components of the Reelin receptor/signalling pathway to control neuronal migration during the development of the nervous system.