The role of the Rho GTPases in neuronal development

Our brain serves as a center for cognitive function and neurons within the brain relay and store information about our surroundings and experiences. Modulation of this complex neuronal circuitry allows us to process that information and respond appropriately. Proper development of neurons is therefore vital to the mental health of an individual, and perturbations in their signaling or morphology are likely to result in cognitive impairment. The development of a neuron requires a series of steps that begins with migration from its birth place and initiation of process outgrowth, and ultimately leads to differentiation and the formation of connections that allow it to communicate with appropriate targets. Over the past several years, it has become clear that the Rho family of GTPases and related molecules play an important role in various aspects of neuronal development, including neurite outgrowth and differentiation, axon pathfinding, and dendritic spine formation and maintenance. Given the importance of these molecules in these processes, it is therefore not surprising that mutations in genes encoding a number of regulators and effectors of the Rho GTPases have been associated with human neurological diseases. This review will focus on the role of the Rho GTPases and their associated signaling molecules throughout neuronal development and discuss how perturbations in Rho GTPase signaling may lead to cognitive disorders

Regulators of Rho GTPases in neuronal development

The formation and elaboration of axonal and dendritic morphologies are fundamental aspects of neuronal polarization critical for information processing. In general, developing CNS neurons elaborate one axon and multiple dendrites in response to intracellular and extracellular cues, so as to transmit and receive information, respectively. The molecular mechanisms underlying axon-dendrite polarity are complex and involve the integration of numerous signaling pathways that impinge on the cytoskeleton. One group of proteins, the Rho GTPases, has emerged as key integrators of environmental cues to regulate the underlying axonal and dendritic cytoskeletons. Here, we discuss the role of regulators of the Rac1 GTPase in axon development and highlight the importance of both actin and microtubule remodeling in this process

The junctional multidomain protein AF-6 is a binding partner of the Rap1A GTPase and associates with the actin cytoskeletal regulator profilin

The AF-6 protein is a multidomain protein that contains two potential Ras-binding domains within its N terminus. Because of this feature, AF-6 has been isolated in both two-hybrid and biochemical approaches and is postulated to be a potential Ras-effector protein, Herein, we show that it is specifically the first Ras-binding domain of AF-6 that mediates this interaction and that the Ras-related Rap1A protein can associate with this motif even more efficiently than the oncogenic Ha-, K-, and N-Ras GTPases. We further demonstrate that both Ras and Rap1 interact with full-length AF-6 in vivo in mammalian cells and that a fraction of Rap1 colocalizes with AF-6 at the membrane. Dominant active Rap1A, in contrast to Ras, when introduced into epithelial MDCK and MCF-7 cells, does not perturb AF-6-specific residency in cell-cell adhesion complexes. In a pursuit to gain further understanding of the role of AF-6 in junctions, we identified profilin as an AF-6-binding protein. Profilin activates monomeric actin units for subsequent polymerization steps at barbed ends of actin filaments and has been shown to participate in cortical actin assembly. To our knowledge, AF-6 is the only integral component in cell-cell junctions discovered thus far that interacts with profilin and thus could modulate actin modeling proximal to adhesion complexes

WNT3 inhibits cerebellar granule neuron progenitor proliferation and medulloblastoma formation via MAPK activation

During normal cerebellar development, the remarkable expansion of granule cell progenitors (GCPs) generates a population of granule neurons that outnumbers the total neuronal population of the cerebral cortex, and provides a model for identifying signaling pathways that may be defective in medulloblastoma. While many studies focus on identifying pathways that promote growth of GCPs, a critical unanswered question concerns the identification of signaling pathways that block mitogenic stimulation and induce early steps in differentiation. Here we identify WNT3 as a novel suppressor of GCP proliferation during cerebellar development and an inhibitor of medulloblastoma growth in mice. WNT3, produced in early postnatal cerebellum, inhibits GCP proliferation by down-regulating proproliferative target genes of the mitogen Sonic Hedgehog (SHH) and the bHLH transcription factor Atoh1. WNT3 suppresses GCP growth through a non-canonical Wnt signaling pathway, activating prototypic mitogen-activated protein kinases (MAPKs), the Ras-dependent extracellular-signal-regulated kinases 1/2 (ERK1/2) and ERK5, instead of the classical β-catenin pathway. Inhibition of MAPK activity using a MAPK kinase (MEK) inhibitor reversed the inhibitory effect of WNT3 on GCP proliferation. Importantly, WNT3 inhibits proliferation of medulloblastoma tumor growth in mouse models by a similar mechanism. Thus, the present study suggests a novel role for WNT3 as a regulator of neurogenesis and repressor of neural tumors. © 2013 Anne et al

Myosin II Motors and F-Actin Dynamics Drive the Coordinated Movement of the Centrosome and Soma during CNS Glial-Guided Neuronal Migration

SummaryLamination of cortical regions of the vertebrate brain depends on glial-guided neuronal migration. The conserved polarity protein Par6α localizes to the centrosome and coordinates forward movement of the centrosome and soma in migrating neurons. The cytoskeletal components that produce this unique form of cell polarity and their relationship to polarity signaling cascades are unknown. We show that F-actin and Myosin II motors are enriched in the neuronal leading process and that Myosin II activity is necessary for leading process actin dynamics. Inhibition of Myosin II decreased the speed of centrosome and somal movement, whereas Myosin II activation increased coordinated movement. Ectopic expression or silencing of Par6α inhibited Myosin II motors by decreasing Myosin light-chain phosphorylation. These findings suggest leading-process Myosin II may function to “pull” the centrosome and soma forward during glial-guided migration by a mechanism involving the conserved polarity protein Par6α

Histone demethylase KDM4B regulates otic vesicle invagination via epigenetic control of Dlx3 expression

In vertebrates, the inner ear arises from the otic placode, a thickened swathe of ectoderm that invaginates to form the otic vesicle. We report that histone demethylase KDM4B is dynamically expressed during early stages of chick inner ear formation. A loss of KDM4B results in defective invagination and striking morphological changes in the otic epithelium, characterized by abnormal localization of adhesion and cytoskeletal molecules and reduced expression of several inner ear markers, including Dlx3. In vivo chromatin immunoprecipitation reveals direct and dynamic occupancy of KDM4B and its target, H3K9me3, at regulatory regions of the Dlx3 locus. Accordingly, coelectroporations of DLX3 or KDM4B encoding constructs, but not a catalytically dead mutant of KDM4B, rescue the ear invagination phenotype caused by KDM4B knockdown. Moreover, a loss of DLX3 phenocopies a loss of KDM4B. Collectively, our findings suggest that KDM4B play a critical role during inner ear invagination via modulating histone methylation of the direct target Dlx3

The RHO-1 RhoGTPase Modulates Fertility and Multiple Behaviors in Adult C. elegans

The Rho family of small GTPases are essential during early embryonic development making it difficult to study their functions in adult animals. Using inducible transgenes expressing either a constitutively active version of the single C. elegans Rho ortholog, RHO-1, or an inhibitor of endogenous Rho (C3 transferase), we demonstrate multiple defects caused by altering Rho signaling in adult C. elegans. Changes in RHO-1 signaling in cholinergic neurons affected locomotion, pharyngeal pumping and fecundity. Changes in RHO-1 signaling outside the cholinergic neurons resulted in defective defecation, ovulation, and changes in C. elegans body morphology. Finally both increased and decreased RHO-1 signaling in adults resulted in death within hours. The multiple post-developmental roles for Rho in C. elegans demonstrate that RhoA signaling pathways continue to be used post-developmentally and the resulting phenotypes provide an opportunity to further study post-developmental Rho signaling pathways using genetic screens

Alterations of brain and cerebellar proteomes linked to Aβ and tau pathology in a female triple-transgenic murine model of Alzheimer's disease

The triple-transgenic Alzheimer (3 × Tg-AD) mouse expresses mutant PS1M146V, APPswe, and tauP301L transgenes and progressively develops plaques and neurofibrillary tangles with a temporal- and region-specific profile that resembles the neuropathological progression of Alzheimer's disease (AD). In this study, we used proteomic approaches such as two-dimensional gel electrophoresis and mass spectrometry to investigate the alterations in protein expression occurring in the brain and cerebellum of 3 × Tg-AD and presenilin-1 (PS1) knock-in mice (animals that do not develop Aβ- or tau-dependent pathology nor cognitive decline and were used as control). Finally, using the Ingenuity Pathway Analysis we evaluated novel networks and molecular pathways involved in this AD model. We identified several differentially expressed spots and analysis of 3 × Tg-AD brains showed a significant downregulation of synaptic proteins that are involved in neurotransmitter synthesis, storage and release, as well as a set of proteins that are associated with cytoskeleton assembly and energy metabolism. Interestingly, in the cerebellum, a structure not affected by AD, we found an upregulation of proteins involved in carbohydrate metabolism and protein catabolism. Our findings help to unravel the pathogenic brain mechanisms set in motion by mutant amyloid precursor protein (APP) and hyperphosphorylated tau. These data also reveal cerebellar pathways that may be important to counteract the pathogenic actions of Aβ and tau, and ultimately offer novel targets for therapeutic intervention

RhoGTPase Regulators Orchestrate Distinct Stages of Synaptic Development

Small RhoGTPases regulate changes in post-synaptic spine morphology and density that support learning and memory. They are also major targets of synaptic disorders, including Autism. Here we sought to determine whether upstream RhoGTPase regulators, including GEFs, GAPs, and GDIs, sculpt specific stages of synaptic development. The majority of examined molecules uniquely regulate either early spine precursor formation or later matura- tion. Specifically, an activator of actin polymerization, the Rac1 GEF β-PIX, drives spine pre- cursor formation, whereas both FRABIN, a Cdc42 GEF, and OLIGOPHRENIN-1, a RhoA GAP, regulate spine precursor elongation. However, in later development, a novel Rac1 GAP, ARHGAP23, and RhoGDIs inactivate actomyosin dynamics to stabilize mature synap- ses. Our observations demonstrate that specific combinations of RhoGTPase regulatory pro- teins temporally balance RhoGTPase activity during post-synaptic spine development

RhoE Deficiency Produces Postnatal Lethality, Profound Motor Deficits and Neurodevelopmental Delay in Mice

Rnd proteins are a subfamily of Rho GTPases involved in the control of actin cytoskeleton dynamics and other cell functions such as motility, proliferation and survival. Unlike other members of the Rho family, Rnd proteins lack GTPase activity and therefore remain constitutively active. We have recently described that RhoE/Rnd3 is expressed in the Central Nervous System and that it has a role in promoting neurite formation. Despite their possible relevance during development, the role of Rnd proteins in vivo is not known. To get insight into the in vivo function of RhoE we have generated mice lacking RhoE expression by an exon trapping cassette. RhoE null mice (RhoE gt/gt) are smaller at birth, display growth retardation and early postnatal death since only half of RhoE gt/gt mice survive beyond postnatal day (PD) 15 and 100% are dead by PD 29. RhoE gt/gt mice show an abnormal body position with profound motor impairment and impaired performance in most neurobehavioral tests. Null mutant mice are hypoactive, show an immature locomotor pattern and display a significant delay in the appearance of the hindlimb mature responses. Moreover, they perform worse than the control littermates in the wire suspension, vertical climbing and clinging, righting reflex and negative geotaxis tests. Also, RhoE ablation results in a delay of neuromuscular maturation and in a reduction in the number of spinal motor neurons. Finally, RhoE gt/gt mice lack the common peroneal nerve and, consequently, show a complete atrophy of the target muscles. This is the first model to study the in vivo functions of a member of the Rnd subfamily of proteins, revealing the important role of Rnd3/RhoE in the normal development and suggesting the possible involvement of this protein in neurological disorders