isoforms, called 2a and 2b, show only 10% identity in
Semaphorins are a large family of axon guidance molecules that are known primarily as ligands for plexins and neuropilins. Although class-6 semaphorins are transmembrane proteins, they have been implicated as ligands in different aspects... more
Semaphorins are a large family of axon guidance molecules that are known primarily as ligands for plexins and neuropilins. Although class-6 semaphorins are transmembrane proteins, they have been implicated as ligands in different aspects of neural development, including neural crest cell migration, axon guidance and cerebellar development. However, the specific spatial and temporal expression of semaphorin 6B (Sema6B) in chick commissural neurons suggested a receptor role in axon guidance at the spinal cord midline. Indeed, in the absence of Sema6B, post-crossing commissural axons lacked an instructive signal directing them rostrally along the contralateral floorplate border, resulting in stalling at the exit site or even caudal turns. Truncated Sema6B lacking the intracellular domain was unable to rescue the loss-of-function phenotype, confirming a receptor function of Sema6B. In support of this, we demonstrate that Sema6B binds to floorplate-derived plexin A2 (PlxnA2) for navigati...
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Loss-of-function approaches are important tools for functional gene analysis. Due to the availability of sophisticated methods to manipulate gene expression in embryonic stem cells that can be used to generate mutant mice, the mouse is by... more
Loss-of-function approaches are important tools for functional gene analysis. Due to the availability of sophisticated methods to manipulate gene expression in embryonic stem cells that can be used to generate mutant mice, the mouse is by far the most important vertebrate model organism for basic and applied biomedical research. Unfortunately, the available methods do not allow for precise temporal and spatial control of gene silencing during embryonic development limiting the usefulness of the mouse for developmental studies. Due to their easy accessibility chicken embryos have been one of the preferred model organisms for developmental studies. Their disadvantage, the lack of genetic tools, could be overcome by the development of in ovo RNAi (in ovo RNA interference), a method that allows for temporal and spatial control of gene silencing in vivo.
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Axon navigation depends on the interactions between guidance molecules along the trajectory and specific receptors on the growth cone. However, our in vitro and in vivo studies on the role of Endoglycan demonstrate that in addition to... more
Axon navigation depends on the interactions between guidance molecules along the trajectory and specific receptors on the growth cone. However, our in vitro and in vivo studies on the role of Endoglycan demonstrate that in addition to specific guidance cue – receptor interactions, axon guidance depends on fine-tuning of cell-cell adhesion. Endoglycan, a sialomucin, plays a role in axon guidance in the central nervous system of chicken embryos, but it is neither an axon guidance cue nor a receptor. Rather, Endoglycan acts as a negative regulator of molecular interactions based on evidence from in vitro experiments demonstrating reduced adhesion of growth cones. In the absence of Endoglycan, commissural axons fail to properly navigate the midline of the spinal cord. Taken together, our in vivo and in vitro results support the hypothesis that Endoglycan acts as a negative regulator of cell-cell adhesion in commissural axon guidance.
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ABSTRACTDuring neural circuit formation, axons navigate several choice points to reach their final target. At each one of these intermediate targets, growth cones need to switch responsiveness from attraction to repulsion in order to move... more
ABSTRACTDuring neural circuit formation, axons navigate several choice points to reach their final target. At each one of these intermediate targets, growth cones need to switch responsiveness from attraction to repulsion in order to move on. Molecular mechanisms that allow for the precise timing of surface expression of a new set of receptors that support the switch in responsiveness are difficult to study in vivo. Mostly, mechanisms are inferred from the observation of snapshots of many different growth cones analyzed in different preparations of tissue harvested at distinct time points. However, to really understand the behavior of growth cones at choice points, a single growth cone should be followed arriving at and leaving the intermediate target.Here, we describe a spinal cord preparation that allows for live imaging of individual axons during navigation in their intact environment. The possibility to observe single growth cones navigating their intermediate target allows for ...
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Axonal growth and guidance rely on correct growth cone responses to guidance cues. Unlike the signaling cascades that link axonal growth to cytoskeletal dynamics, little is known about the crosstalk mechanisms between guidance and... more
Axonal growth and guidance rely on correct growth cone responses to guidance cues. Unlike the signaling cascades that link axonal growth to cytoskeletal dynamics, little is known about the crosstalk mechanisms between guidance and membrane dynamics and turnover. Recent studies indicate that whereas axonal attraction requires exocytosis, chemorepulsion relies on endocytosis. Indeed, our own studies have shown that Netrin-1/Deleted in Colorectal Cancer (DCC) signaling triggers exocytosis through the SNARE Syntaxin-1 (STX1). However, limited in vivo evidence is available about the role of SNARE proteins in axonal guidance. To address this issue, here we systematically deleted SNARE genes in three species. We show that loss-of-function of STX1 results in pre- and post-commissural axonal guidance defects in the midline of fly, chick, and mouse embryos. Inactivation of VAMP2, Ti-VAMP, and SNAP25 led to additional abnormalities in axonal guidance. We also confirmed that STX1 loss-of-functi...
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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... more
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 ar...
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Axon guidance at choice points depends on the precise regulation of guidance receptors on the growth cone surface. Upon arrival at the intermediate target or choice point, a switch from attraction to repulsion is required for the axon to... more
Axon guidance at choice points depends on the precise regulation of guidance receptors on the growth cone surface. Upon arrival at the intermediate target or choice point, a switch from attraction to repulsion is required for the axon to move on. Dorsal commissural (dI1) axons crossing the ventral midline of the spinal cord in the floor plate represent a convenient model for the analysis of the molecular mechanism underlying the switch in axonal behavior. We identified a role of Calsyntenin1 in the regulation of vesicular trafficking of guidance receptors in dI1 axons at choice points. In cooperation with RabGDI, Calsyntenin1 shuttles Rab11-positive vesicles containing Robo1 to the growth cone surface in a precisely regulated manner. In contrast, Calsyntenin1-mediated trafficking of Frizzled3, a guidance receptor in the Wnt pathway, is independent of RabGDI. Thus, tightly regulated insertion of guidance receptors, which is required for midline crossing and the subsequent turn into t...
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<b>Copyright information:</b>Taken from "Semaphorin6A acts as a gate keeper between the central and the peripheral nervous system"http://www.neuraldevelopment.com/content/2/1/28Neural Development... more
<b>Copyright information:</b>Taken from "Semaphorin6A acts as a gate keeper between the central and the peripheral nervous system"http://www.neuraldevelopment.com/content/2/1/28Neural Development 2007;2():28-28.Published online 18 Dec 2007PMCID:PMC2238753. In the absence of Sema6A from BCCs, motoneurons stream out of the ventral spinal cord and migrate along the ventral roots (arrows). The open arrow points to a motoneuron that is located in the ventral funiculus. In control-treated embryos motoneurons along ventral roots or in the ventral funiculus were rarely seen. Motoneurons were identified by Isl-1 (red). An EGFP plasmid was co-injected with the dsRNA derived from SEMA6A. Axons were stained with an antibody against neurofilament (blue). Note that sensory neurons in the DRG (asterisk in (a, b)) are also stained by Isl-1. Perturbation of Sema6B or Sema6D did not enhance the number of motoneurons in the periphery compared to control-treated embryos injected only with the plasmid encoding EGFP. Three asterisks indicate &lt; 0.0001 for the comparison between dsS6A and all other treatment groups. Values are given as mean ± standard error of mean. Bar: 50 μm.
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As indicated by their name, morphogens were first identified for their role in the formation of tissues early in development. Secreted from a source, they spread through the tissue to form gradients by which they affect the... more
As indicated by their name, morphogens were first identified for their role in the formation of tissues early in development. Secreted from a source, they spread through the tissue to form gradients by which they affect the differentiation of precursor cells in a concentration-dependent manner. In this context, the antagonistic roles of the morphogens of the Wnt family and Sonic hedgehog (Shh) in the specification of cell types along the dorso-ventral axis of the neural tube have been studied in detail. However, more recently, morphogens have been demonstrated to act well beyond the early stages of nervous system development, as additional roles of morphogen gradients in vertebrate neural circuit formation have been identified. Both Wnt and Shh affect neural circuit formation at several stages by their influence on neurite extension, axon pathfinding and synapse formation. In this review, we will summarize the mechanisms of morphogen function during axon guidance in the vertebrate n...
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Semaphorins are a large family of axon guidance molecules that are known primarily as ligands for plexins and neuropilins. Although class-6 semaphorins are transmembrane proteins, they have been implicated as ligands in different aspects... more
Semaphorins are a large family of axon guidance molecules that are known primarily as ligands for plexins and neuropilins. Although class-6 semaphorins are transmembrane proteins, they have been implicated as ligands in different aspects of neural development, including neural crest cell migration, axon guidance and cerebellar development. However, the specific spatial and temporal expression of semaphorin 6B (Sema6B) in chick commissural neurons suggested a receptor role in axon guidance at the spinal cord midline. Indeed, in the absence of Sema6B, post-crossing commissural axons lacked an instructive signal directing them rostrally along the contralateral floorplate border, resulting in stalling at the exit site or even caudal turns. Truncated Sema6B lacking the intracellular domain was unable to rescue the loss-of-function phenotype, confirming a receptor function of Sema6B. In support of this, we demonstrate that Sema6B binds to floorplate-derived plexin A2 (PlxnA2) for navigati...
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Research Interests: Genetics, Cell Migration, Apoptosis, Gene Silencing, Neural Development, and 15 moreBiological Sciences, RNA interference, Cerebellum, Electroporation, Animals, Axon Guidance, Chickens, Nervous System, Embryos, Axons, Immunoglobulin, Neurite Outgrowth, Chick embryo, Medical and Health Sciences, and Expression pattern
Background Axons navigate to their future synaptic targets with the help of choice points, intermediate targets that express axon guidance cues. Once they reach a choice point, axons need to switch their response from attraction to... more
Background Axons navigate to their future synaptic targets with the help of choice points, intermediate targets that express axon guidance cues. Once they reach a choice point, axons need to switch their response from attraction to repulsion in order to move on with the next stage of their journey. The mechanisms underlying the change in axonal responsiveness are poorly understood. Commissural axons become sensitive to the repulsive activity of Slits when they cross the ventral midline of the CNS. Responsiveness to Slits depends on surface expression of Robo receptors. In Drosophila, Commissureless (Comm) plays a crucial regulatory role in midline crossing by keeping Robo levels low on precommissural axons. Interestingly, to date no vertebrate homolog of comm has been identified. Robo3/Rig1 has been shown to control Slit sensitivity before the midline, but without affecting Robo1 surface expression. Results We had identified RabGDI, a gene linked to human mental retardation and an e...
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The Necl/SynCAM subgroup of immunoglobulin superfamily cell adhesion molecules has been implicated in late stages of neural circuit formation. They were shown to be sufficient for synaptogenesis by their trans-synaptic interactions.... more
The Necl/SynCAM subgroup of immunoglobulin superfamily cell adhesion molecules has been implicated in late stages of neural circuit formation. They were shown to be sufficient for synaptogenesis by their trans-synaptic interactions. Additionally, they are involved in myelination, both in the central and the peripheral nervous system, by mediating adhesion between glia cells and axons. Here, we show that Necls/SynCAMs are also required for early stages of neural circuit formation. We demonstrate a role for Necls/SynCAMs in post-crossing commissural axon guidance in the developing spinal cord in vivo. Necl3/SynCAM2, the family member that has not been characterized functionally so far, plays a crucial role in this process. It is expressed by floorplate cells and interacts with Necls/SynCAMs expressed by commissural axons to mediate a turning response in post-crossing commissural axons.
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The Journal of Neuroscience, QUICK SEARCH: [advanced], Author: Keyword(s): Year: Vol: Page: ...
Research Interests: Neuroscience, Biology, Medicine, Western blotting, In Situ Hybridization, and 13 moreDrosophila, Electroporation, Animals, Spinal Cord, Phenotype, Amino Acids, Protein Tyrosine Phosphatases, Axons, Motor Neurons, Psychology and Cognitive Sciences, reverse transcriptase polymerase chain reaction, Chick embryo, and Medical and Health Sciences
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Research Interests: Cognitive Science, Cell Adhesion, Immunohistochemistry, Extracellular Matrix, Western blotting, and 13 moreAnimals, Spinal Cord, Peptides, Molecular cloning, Rats, Neuron, Embryos, Axons, Neurosciences, Neutralizing antibodies, Molecular Sequence Data, Neural Cell Adhesion Molecules, and Chick embryo
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<b>Copyright information:</b>Taken from "Semaphorin6A acts as a gate keeper between the central and the peripheral nervous system"http://www.neuraldevelopment.com/content/2/1/28Neural Development... more
<b>Copyright information:</b>Taken from "Semaphorin6A acts as a gate keeper between the central and the peripheral nervous system"http://www.neuraldevelopment.com/content/2/1/28Neural Development 2007;2():28-28.Published online 18 Dec 2007PMCID:PMC2238753. In control embryos axon bundles from each dorsal root ganglion extend to the DREZ in a well organized manner. Roots from adjacent DRGs are segregated and they are all of the same length (dashed bars). In contrast, in embryos lacking Sema6A, roots from adjacent DRGs are no longer segregated (arrowheads). The arrangement of roots arising from individual DRGs is strongly disorganized and roots are often formed by fibers from two adjacent DRGs (arrowheads in (b)). Similarly, roots are disorganized in embryos lacking Sema6D (arrowheads). In addition the length of the roots varied more in the absence of Sema6D (compare dashed bars in (c)). Strong phenotypes were seen in 71% of the embryos lacking Sema6A and in 68% of the embryos lacking Sema6D. Only 13% of the embryos injected with an EGFP plasmid had a comparable phenotype. Downregulation of Sema6B resulted in aberrant DRG shapes and root arrangement in 30% of the embryos. The shapes of DRGs were classified as arc-like when the distance between the most anterior and the most posterior fiber emanating from the DRG was the same as the anteroposterior diameter of the DRG; as bell-shaped when the fibers spread an anteroposterior length that was bigger than the diameter of the DRG; and as mushroom-like when the fibers entered the dorsal spinal cord in a segment that was shorter than the diameter of the DRG. Note that the diameter of the mushroom-like DRGs was smaller than the diameter of arc-like or bell-shaped DRGs. Bar: 200 μm.
Axonal growth and guidance rely on correct growth cone responses to guidance cues, both in the central nervous system (CNS) and in the periphery. Unlike the signaling cascades that link axonal growth to cytoskeletal dynamics, little is... more
Axonal growth and guidance rely on correct growth cone responses to guidance cues, both in the central nervous system (CNS) and in the periphery. Unlike the signaling cascades that link axonal growth to cytoskeletal dynamics, little is known about the crosstalk mechanisms between guidance and membrane dynamics and turnover in the axon. Our studies have shown that Netrin-1/Deleted in Colorectal Cancer (DCC) signaling triggers exocytosis through the SNARE Syntaxin-1 (STX-1) during the formation of commissural pathways. However, limited in vivo evidence is available about the role of SNARE proteins in motor axonal guidance. Here we show that loss-of-function of SNARE complex members results in motor axon guidance defects in fly and chick embryos. Knock-down of Syntaxin-1, VAMP-2, and SNAP-25 leads to abnormalities in the motor axon routes out of the CNS. Our data point to an evolutionarily conserved role of the SNARE complex proteins in motor axon guidance, thereby pinpointing an impor...
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Morphogens have been identified as guidance cues for postcrossing commissural axons in the spinal cord. Shh has a dual effect on postcrossing commissural axons: a direct repellent effect mediated by Hhip as a receptor, and an indirect... more
Morphogens have been identified as guidance cues for postcrossing commissural axons in the spinal cord. Shh has a dual effect on postcrossing commissural axons: a direct repellent effect mediated by Hhip as a receptor, and an indirect effect by shaping a Wnt activity gradient. Wnts were shown to be attractants for postcrossing commissural axons in both chicken and mouse embryos. In mouse, the effects of Wnts on axon guidance were concluded to depend on the planar cell polarity (PCP) pathway. Canonical Wnt signaling was excluded based on the absence of axon guidance defects in mice lacking Lrp6 which is an obligatory coreceptor for Fzd in canonical Wnt signaling. In the loss-of-function studies reported here, we confirmed a role for the PCP pathway in postcrossing commissural axon guidance also in the chicken embryo. However, taking advantage of the precise temporal control of gene silencing provided by in ovo RNAi, we demonstrate that canonical Wnt signaling is also required for pro...
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Plexins are a family of transmembrane proteins that were shown to act as receptors for Semaphorins either alone or in a complex together with Neuropilins. Based on structural criteria Plexins were subdivided into 4 classes, A through D.... more
Plexins are a family of transmembrane proteins that were shown to act as receptors for Semaphorins either alone or in a complex together with Neuropilins. Based on structural criteria Plexins were subdivided into 4 classes, A through D. PlexinAs are mainly thought to act as mediators of repulsive signals in cell migration and axon guidance. Their functional role in vertebrates has been studied almost exclusively in the context of Semaphorin signaling, i.e. as co-receptors for class 3 Semaphorins. Much less is known about Plexins of the other three classes. Despite the fact that Plexins are involved in the formation of neuronal circuits, the temporal changes of their expression patterns during development of the nervous system have not been analyzed in detail. Only seven plexins are found in the chicken genome in contrast to mammals, where nine plexins have been identified. Here, we describe the dynamic expression patterns of all known plexin family members in comparison to the neuro...
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An interaction of growth cone axonin-1 with the floor-plate NgCAM-related cell adhesion molecule (NrCAM) was shown to play a crucial role in commissural axon guidance across the midline of the spinal cord. We now provide evidence that... more
An interaction of growth cone axonin-1 with the floor-plate NgCAM-related cell adhesion molecule (NrCAM) was shown to play a crucial role in commissural axon guidance across the midline of the spinal cord. We now provide evidence that axonin-1 mediates a guidance signal without promoting axon elongation. In an in vitro assay, commissural axons grew preferentially on stripes coated with a mixture of NrCAM and NgCAM. This preference was abolished in the presence of anti-axonin-1 antibodies without a decrease in neurite length. Consistent with these findings, commissural axons in vivo only fail to extend along the longitudinal axis when both NrCAM and NgCAM interactions, but not when axonin-1 and NrCAM or axonin-1 and NgCAM interactions, are perturbed. Thus, we conclude that axonin-1 is involved in guidance of commissural axons without promoting their growth.
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Research Interests: Membrane Proteins, Brain, Humans, Sequence alignment, Animals, and 13 moreSpinal Cord, Rats, Transfection, Protein Conformation, Nerve Growth Factor, Amino Acid Sequence, Membrane Protein, Growth Factor, Recombinant Protein, Recombinant Proteins, Embryos, Biochemistry and cell biology, and Cell Adhesion Molecules
We have identified and chromatographically purified an axonally secreted glycoprotein of CNS and PNS neurons. Several peptides derived from it were microsequenced. Based on these sequences, a fragment of the corresponding cDNA was... more
We have identified and chromatographically purified an axonally secreted glycoprotein of CNS and PNS neurons. Several peptides derived from it were microsequenced. Based on these sequences, a fragment of the corresponding cDNA was amplified and used as a probe to isolate a full length cDNA from a chicken brain cDNA library. Because the deduced amino acid sequence qualified the protein as a novel member of the serpin family of serine protease inhibitors, we called it neuroserpin. Analysis of the primary structural features further characterized neuroserpin as a heparin-independent, functional inhibitor of a trypsin-like serine protease. In situ hybridization revealed a predominantly neuronal expression during the late stages of neurogenesis and in the adult brain in regions which exhibit synaptic plasticity. Thus, neuroserpin might function as an axonally secreted regulator of the local extracellular proteolysis involved in the reorganization of the synaptic connectivity during devel...
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After midline crossing, axons of dorsolateral commissural neurons turn rostrally into the longitudinal axis of the spinal cord. In mouse, the graded distribution of Wnt4 attracts post-crossing axons rostrally. In contrast, in the chicken... more
After midline crossing, axons of dorsolateral commissural neurons turn rostrally into the longitudinal axis of the spinal cord. In mouse, the graded distribution of Wnt4 attracts post-crossing axons rostrally. In contrast, in the chicken embryo, the graded distribution of Sonic hedgehog (Shh) guides post-crossing axons by a repulsive mechanism mediated by hedgehog-interacting protein. Based on these observations, we tested for a possible cooperation between the two types of morphogens. Indeed, we found that Wnts also act as axon guidance cues in the chicken spinal cord. However, in contrast to the mouse, Wnt transcription did not differ along the anteroposterior axis of the spinal cord. Rather, Wnt function was regulated by a gradient of the Wnt antagonist Sfrp1 (Secreted frizzled-related protein 1) that in turn was shaped by the Shh gradient. Thus, Shh affects post-crossing axon guidance both directly and indirectly by regulating Wnt function.
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During development of the nervous system, growing axons must navigate through their environment to find their correct target. Accurate pathfinding of axons is essential for the establishment of functional neuronal circuits. How do axons... more
During development of the nervous system, growing axons must navigate through their environment to find their correct target. Accurate pathfinding of axons is essential for the establishment of functional neuronal circuits. How do axons know where to go? What provides them with the necessary information? Which part of the axon actually makes the directional decision? These are questions scientists have
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Semaphorins and their receptors, plexins, have emerged as important cellular cues regulating key developmental processes. B-type plexins directly regulate the actin cytoskeleton in a variety of cell types. Recently, B-type plexins have... more
Semaphorins and their receptors, plexins, have emerged as important cellular cues regulating key developmental processes. B-type plexins directly regulate the actin cytoskeleton in a variety of cell types. Recently, B-type plexins have been shown to be expressed in striking patterns in the nervous system over critical developmental windows. However, in contrast to the well characterized plexin-A family, the functional role of plexin-B proteins in neural development and organogenesis in vertebrates in vivo is not known. Here, we have elucidated the functional contribution of the two neuronally expressed plexin-B proteins, Plexin-B1 or Plexin-B2, toward the development of the peripheral nervous system and the CNS by generating and analyzing constitutive knock-out mice. The development of the nervous system was found to be normal in mice lacking Plexin-B1, whereas mice lacking Plexin-B2 demonstrated defects in closure of the neural tube and a conspicuous disorganization of the embryonic brain. After analyzing mutant mice, which bypassed neural tube defects, we observed a key requirement for Plexin-B2 in proliferation and migration of granule cell precursors in the developing dentate gyrus, olfactory bulb, and cerebellum. Furthermore, we identified semaphorin 4C as a high-affinity ligand for Plexin-B2 in binding and functional assays. Semaphorin 4C stimulated activation of ErbB-2 and RhoA via Plexin-B2 and enhanced proliferation and migration of granule cell precursors. Semaphorin 4C-induced proliferation of ventricular zone neuroblasts was abrogated in mice lacking Plexin-B2. These genetic and functional analyses reveal a key requirement for Plexin-B2, but not Plexin-B1, in patterning of the vertebrate nervous system in vivo.