FARP1 Promotes the Dendritic Growth of Spinal Motor Neuron Subtypes through Transmembrane Semaphorin6A and PlexinA4 Signaling

SummaryThe dendritic morphology of neurons dictates their abilities to process and transmit information; however, the signaling pathways that regulate dendritic growth and complexity are poorly understood. Here, we show that retinoids induce the expression of the FERM Rho-GEF protein FARP1 in the developing spinal cord. FARP1 is expressed in subsets of motor neurons and is enriched in dendrites of lateral motor column (LMC) neurons that innervate the limb. FARP1 is necessary and sufficient to promote LMC dendritic growth but does not affect dendrite number or axonal morphology. We show that FARP1 serves as a specific effector of transmembrane Semaphorin6A and PlexinA4 signals to regulate LMC dendritic growth, and that its Rho-GEF domain is necessary for this function. These findings reveal that retinoid and Sema6A/PlexA4 signaling pathways intersect through FARP1 to control dendritic growth, and uncover the existence of subtype-specific signaling networks that control dendritic developmental programs in spinal motor neurons

The Antioxidant Enzyme Prdx1 Controls Neuronal Differentiation by Thiol-Redox-Dependent Activation of GDE2

SummaryThe six-transmembrane protein GDE2 controls the onset and progression of spinal motor neuron differentiation through extracellular glycerophosphodiester phosphodiesterase metabolism. Although this process is likely to be tightly regulated, the relevant mechanisms that modulate its activity are unknown. Here we show that the antioxidant scavenger peroxiredoxin1 (Prdx1) interacts with GDE2, and that loss of Prdx1 causes motor neuron deficits analogous to GDE2 ablation. Prdx1 cooperates with GDE2 to drive motor neuron differentiation, and this synergy requires Prdx1 thiol-dependent catalysis. Prdx1 activates GDE2 through reduction of an intramolecular disulfide bond that bridges its intracellular N- and C-terminal domains. GDE2 variants incapable of disulfide bond formation acquire independence from Prdx1 and are potent inducers of motor neuron differentiation. These findings define Prdx1 as a pivotal regulator of GDE2 activity and suggest roles for coupled thiol-redox-dependent cascades in controlling neuronal differentiation in the spinal cord

Complementary Domains of Retinoic Acid Production and Degradation in the Early Chick Embryo

AbstractExcess retinoids as well as retinoid deprivation cause abnormal development, suggesting that retinoid homeostasis is critical for proper morphogenesis. RALDH-2 and CYP26, two key enzymes that carry out retinoic acid (RA) synthesis and degradation, respectively, were cloned from the chick and show significant homology with their orthologs in other vertebrates. Expression patterns of RALDH-2 and CYP26 genes were determined in the early chick embryo by in situ hybridization. During gastrulation and neurulation RALDH-2 and CYP26 were expressed in nonoverlapping regions, with RALDH-2 transcripts localized to the presumptive presomitic and lateral plate mesoderm and CYP26 mRNA to the presumptive mid- and forebrain. The two domains of expression were separated by an approximately 300-μm-wide gap, encompassing the presumptive hindbrain. In the limb region, a similar spatial segregation of RALDH-2 and CYP26 expression was found at stages 14 and 15. Limb region mesoderm expressed RALDH-2, whereas the overlying limb ectoderm expressed CYP26. RA-synthesizing and -degrading enzymatic activities were measured biochemically in regions expressing RALDH-2 or CYP26. Regions expressing RALDH-2 generated RA efficiently from precursor retinal but degraded RA only inefficiently. Conversely, tissue expressing CYP26 efficiently degraded but did not synthesize RA. Localized regions of RA synthesis and degradation mediated by these two enzymes may therefore provide a mechanism to regulate RA homeostasis spatially in vertebrate embryos

A retinoic acid-dependent checkpoint in the development of CD4+ T cell-mediated immunity

It is known that vitamin A and its metabolite, retinoic acid (RA), are essential for host defense. However, the mechanisms for how RA controls inflammation are incompletely understood. The findings presented in this study show that RA signaling occurs concurrent with the development of inflammation. In models of vaccination and allogeneic graft rejection, whole body imaging reveals that RA signaling is temporally and spatially restricted to the site of inflammation. Conditional ablation of RA signaling in T cells significantly interferes with CD4(+) T cell effector function, migration, and polarity. These findings provide a new perspective of the role of RA as a mediator directly controlling CD4(+) T cell differentiation and immunity

A retinoic acid–dependent checkpoint in the development of CD4+ T cell–mediated immunity

Immune cell activation induces concurrent temporal and spatial retinoic acid signaling, and CD4+ T cell–specific loss of RA signals reduces effector function, migration, and polarity

Nolz1 is induced by retinoid signals and controls motoneuron subtype identity through distinct repressor activities

The acquisition and maintenance of final neuronal identity depends in part upon the implementation of fate-specification programs in postmitotic neurons; however, the mechanisms involved remain unclear. In the developing spinal cord, retinoic acid (RA) signaling pathways specify the columnar and divisional identities of postmitotic motoneurons (MNs). Here we show that RA signals induce expression of the NET transcriptional regulator Nolz1 in differentiated chick MNs, where it regulates the progressive specification of prospective Lim3-negative motor columns. Nolz1 controls the initial formation of forelimb and thoracic Lim3-negative motor columns by downregulating Lim3 expression and maintaining the expression of key homeodomain proteins necessary for MN identity and survival. At forelimb levels, Nolz1 specifies lateral motor column (LMC) identity by inducing the expression of the postmitotic LMC determinant Hoxc6, and implements the partial specification of lateral LMC identity through Lim1 induction. The specificity of Nolz1 function depends upon distinct repressor activities that require, in part, the modulatory activity of Grg5, an atypical member of the Gro-TLE family of co-repressors. Thus, RA signals regulate diverse events in MN subtype specification by inducing the expression of a key transcriptional regulator that controls multiple developmental pathways via functionally distinct repressor complexes

Semaphorin 6B acts as a receptor in post-crossing commissural axon guidance

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 navigation at the midline, whereas a cis-interaction between PlxnA2 and Sema6B on pre-crossing commissural axons may regulate the responsiveness of axons to floorplate-derived cues

Real-Time Imaging Reveals Properties of Glutamate-Induced Arc/Arg 3.1 Translation in Neuronal Dendrites

The immediate early gene Arc (also Arg3.1) produces rapid changes in synaptic properties that are linked to de novo translation. Here we develop a novel translation reporter that exploits the rapid maturation and “flash” kinetics of Gaussia luciferase (Gluc) to visualize Arc translation. Following glutamate stimulation, discrete Arc-Gluc bioluminescent flashes representing sites of de novo translation are detected within 15 s at distributed sites in dendrites, but not spines. Flashes are episodic, lasting ∼20 s, and may be unitary or repeated at ∼minute intervals at the same sites. Analysis of flash amplitudes suggests they represent the quantal product of one or more polyribosomes, while inter-flash intervals appear random, suggesting they arise from a stochastic process. Surprisingly, glutamate-induced translation is dependent on Arc open reading frame. Combined observations support a model in which stalled ribosomes are reactivated to rapidly generate Arc protein. © 2016 Elsevier Inc.161

Additional file 1: Figure S1. of GDE2 is essential for neuronal survival in the postnatal mammalian spinal cord

Sensory neurons exhibit neurodegenerative pathology without impaired nerve conduction in the absence of Gde2. This figure illustrates the presence of neuropathology in the primary sensory neurons of the Gde2 KO, including vacuolization, lipid accrual, and cytoskeletal accumulation; and the maintenance of peripheral sensory nerve conduction. Figure S2-related to Fig. 9. Conditional ablation of Gde2 in the postnatal spinal cord prevents the developmental loss of motor neurons. This figure confirms the effective conditional ablation of Gde2 following neurogenesis. In the constitutive KO, GDE2’s absence during embryonic neurogenesis causes a reduction of alpha motor neurons in the lateral motor column; however, in the Gde2lox/-; ROSA:CreER animals, this loss is avoided by injecting 4-OHT at E17.5. Further, competitive PCR analysis shows a near complete deletion of the conditional Gde2 allele following 4-OHT delivery. Figure S3. Gde2 deletion does not perturb neuromuscular junction morphology. This figure uses wholemount immunohistochemistry to assess the integrity of the neuromuscular junction (NMJ) in aged Gde2 KO hindlimb muscle. At 19 months, no discernible pathology is present in the Gde2 KO NMJ. (DOCX 6586 kb