Specific Neuronal Functions Research Articles

Transfer RNA levels are tuned to support differentiation during Drosophila neurogenesis.

Neural differentiation requires a multifaceted program to alter gene expression along the proliferation to differentiation axis. While critical changes occur at the level of transcription, post-transcriptional mechanisms allow fine-tuning of protein output. We investigated the role of tRNAs in regulating gene expression during neural differentiation by quantifying tRNA abundance in neural progenitor-biased and neuron-biased Drosophila larval brains. We found that tRNA profiles are largely consistent between progenitor-biased and neuron-biased brains but significant variation occurs for 10 cytoplasmic isodecoders (individual tRNA genes) and this establishes differential tRNA levels for 8 anticodon groups. We used these tRNA data to investigate relationships between tRNA abundance, codon optimality-mediated mRNA decay, and translation efficiency in progenitors and neurons. Our data reveal that tRNA levels strongly correlate with codon optimality-mediated mRNA decay within each cell type but generally do not explain differences in stabilizing versus destabilizing codons between cell types. Regarding translation efficiency, we found that tRNA expression in neural progenitors preferentially supports translation of mRNAs whose products are in high demand in progenitors, such as those associated with protein synthesis. In neurons, tRNA expression shifts to disfavor translation of proliferation-related transcripts and preferentially support translation of transcripts tied to neuron-specific functions like axon pathfinding and synapse formation. Overall, our analyses reveal that changes in tRNA levels along the neural differentiation axis support optimal gene expression in progenitors and neurons.

Cooperative nuclear action of RNA-binding proteins PSF and G3BP2 to sustain neuronal cell viability is decreased in aging and dementia.

Dysfunctional RNA-binding proteins (RBPs) have been implicated in several geriatric diseases, including Alzheimer's disease (AD). However, little is known about the nuclear molecular actions and cooperative functions mediated by RBPs that affect gene regulation in sporadic AD or aging. In the present study, we investigated aging- and AD-associated changes in the expression of PSF and G3BP2, which are representative RBPs associated with sex hormone activity. We determined that both PSF and G3BP2 levels were decreased in aged brains compared to young brains of mice. RNA sequencing (RNA-seq) analysis of human neuronal cells has shown that PSF is responsible for neuron-specific functions and sustains cell viability. In addition, we showed that PSF interacted with G3BP2 in the nucleus and stress granules (SGs) at the protein level. Moreover, PSF-mediated gene regulation at the RNA level correlated with G3BP2. Interestingly, PSF and G3BP2 target genes are associated with AD development. Mechanistically, quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis demonstrated that the interaction of RBPs with the pre-mRNA of target genes enhanced post-transcriptional mRNA stability, suggesting a possible role for these RBPs in preserving neuronal cell viability. Notably, in the brains of patients with sporadic AD, decreased expression of PSF and G3BP2 in neurons was observed compared to non-AD patients. Overall, our findings suggest that the cooperative action of PSF and G3BP2 in the nucleus is important for preventing aging and AD development.

DNA Damage and Parkinson's Disease.

The etiology underlying most sporadic Parkinson's' disease (PD) cases is unknown. Environmental exposures have been suggested as putative causes of the disease. In cell models and in animal studies, certain chemicals can destroy dopaminergic neurons. However, the mechanisms of how these chemicals cause the death of neurons is not understood. Several of these agents are mitochondrial toxins that inhibit the mitochondrial complex I of the electron transport chain. Familial PD genes also encode proteins with important functions in mitochondria. Mitochondrial dysfunction of the respiratory chain, in combination with the presence of redox active dopamine molecules in these cells, will lead to the accumulation of reactive oxygen species (ROS) in dopaminergic neurons. Here, I propose a mechanism regarding how ROS may lead to cell killing with a specificity for neurons. One rarely considered hypothesis is that ROS produced by defective mitochondria will lead to the formation of oxidative DNA damage in nuclear DNA. Many genes that encode proteins with neuron-specific functions are extraordinary long, ranging in size from several hundred kilobases to well over a megabase. It is predictable that such long genes will contain large numbers of damaged DNA bases, for example in the form of 8-oxoguanine (8-oxoG), which is a major DNA damage type produced by ROS. These DNA lesions will slow down or stall the progression of RNA polymerase II, which is a term referred to as transcription stress. Furthermore, ROS-induced DNA damage may cause mutations, even in postmitotic cells such as neurons. I propose that the impaired transcription and mutagenesis of long, neuron-specific genes will lead to a loss of neuronal integrity, eventually leading to the death of these cells during a human lifetime.

A distinct circular DNA profile intersects with proteome changes in the genotoxic stress-related hSOD1G93A model of ALS

BackgroundNumerous genes, including SOD1, mutated in familial and sporadic amyotrophic lateral sclerosis (f/sALS) share a role in DNA damage and repair, emphasizing genome disintegration in ALS. One possible outcome of chromosomal instability and repair processes is extrachromosomal circular DNA (eccDNA) formation. Therefore, eccDNA might accumulate in f/sALS with yet unknown function.MethodsWe combined rolling circle amplification with linear DNA digestion to purify eccDNA from the cervical spinal cord of 9 co-isogenic symptomatic hSOD1G93A mutants and 10 controls, followed by deep short-read sequencing. We mapped the eccDNAs and performed differential analysis based on the split read signal of the eccDNAs, referred as DifCir, between the ALS and control specimens, to find differentially produced per gene circles (DPpGC) in the two groups. Compared were eccDNA abundances, length distributions and genic profiles. We further assessed proteome alterations in ALS by mass spectrometry, and matched the DPpGCs with differentially expressed proteins (DEPs) in ALS. Additionally, we aligned the ALS-specific DPpGCs to ALS risk gene databases.ResultsWe found a six-fold enrichment in the number of unique eccDNAs in the genotoxic ALS-model relative to controls. We uncovered a distinct genic circulome profile characterized by 225 up-DPpGCs, i.e., genes that produced more eccDNAs from distinct gene sequences in ALS than under control conditions. The inter-sample recurrence rate was at least 89% for the top 6 up-DPpGCs. ALS proteome analyses revealed 42 corresponding DEPs, of which 19 underlying genes were itemized for an ALS risk in GWAS databases. The up-DPpGCs and their DEP tandems mainly impart neuron-specific functions, and gene set enrichment analyses indicated an overrepresentation of the adenylate cyclase modulating G protein pathway.ConclusionsWe prove, for the first time, a significant enrichment of eccDNA in the ALS-affected spinal cord. Our triple circulome, proteome and genome approach provide indication for a potential importance of certain eccDNAs in ALS neurodegeneration and a yet unconsidered role as ALS biomarkers. The related functional pathways might open up new targets for therapeutic intervention.Graphical

Dissecting neuron-specific functions of circadian genes using modified cell-specific CRISPR approaches

Circadian behavioral rhythms in Drosophila melanogaster are regulated by about 75 pairs of brain neurons. They all express the core clock genes but have distinct functions and gene expression profiles. To understand the importance of these distinct molecular programs, neuron-specific gene manipulations are essential. Although RNAi based methods are standard to manipulate gene expression in a cell-specific manner, they are often ineffective, especially in assays involving smaller numbers of neurons or weaker Gal4 drivers. We and others recently exploited a neuron-specific CRISPR-based method to mutagenize genes within circadian neurons. Here, we further explore this approach to mutagenize three well-studied clock genes: the transcription factor gene vrille, the photoreceptor gene Cryptochrome (cry), and the neuropeptide gene Pdf (pigment dispersing factor). The CRISPR-based strategy not only reproduced their known phenotypes but also assigned cry function for different light-mediated phenotypes to discrete, different subsets of clock neurons. We further tested two recently published methods for temporal regulation in adult neurons, inducible Cas9 and the auxin-inducible gene expression system. The results were not identical, but both approaches successfully showed that the adult-specific knockout of the neuropeptide Pdf reproduces the canonical loss-of-function mutant phenotypes. In summary, a CRISPR-based strategy is a highly effective, reliable, and general method to temporally manipulate gene function in specific adult neurons.

Circadian protein TIMELESS regulates synaptic function and memory by modulating cAMP signaling

The regulation of neurons by circadian clock genes is thought to contribute to the maintenance of neuronal functions that ultimately underlie animal behavior. However, the impact of specific circadian genes on cellular and molecular mechanisms controlling synaptic plasticity and cognitive function remains elusive. Here, we show that the expression of the circadian protein TIMELESS displays circadian rhythmicity in the mammalian hippocampus. We identify TIMELESS as a chromatin-bound protein that targets synaptic-plasticity-related genes such as phosphodiesterase 4B (Pde4b). By promoting Pde4b transcription, TIMELESS negatively regulates cAMP signaling to modulate AMPA receptor GluA1 function and influence synaptic plasticity. Conditional deletion of Timeless in the adult forebrain impairs working and contextual fear memory in mice. These cognitive phenotypes were accompanied by attenuation of hippocampal Schaffer-collateral synapse long-term potentiation. Together, these data establish a neuron-specific function of mammalian TIMELESS by defining a mechanism that regulates synaptic plasticity and cognitive function.

Phosphatidylinositol (3,5)-bisphosphate machinery regulates neurite thickness through neuron-specific endosomal protein NSG1/NEEP21

Phosphatidylinositol (3,5)-bisphosphate [PtdIns(3,5)P2] is a critical signaling phospholipid involved in endolysosome homeostasis. It is synthesized by a protein complex composed of PIKfyve, Vac14, and Fig4. Defects in PtdIns(3,5)P2 synthesis underlie a number of human neurological disorders, including Charcot-Marie-Tooth disease, child onset progressive dystonia, and others. However, neuron-specific functions of PtdIns(3,5)P2 remain less understood. Here, we show that PtdIns(3,5)P2 pathway is required to maintain neurite thickness. Suppression of PIKfyve activities using either pharmacological inhibitors or RNA silencing resulted in decreased neurite thickness. We further find that the regulation of neurite thickness by PtdIns(3,5)P2 is mediated by NSG1/NEEP21, a neuron-specific endosomal protein. Knockdown of NSG1 expression also led to thinner neurites. mCherry-tagged NSG1 colocalized and interacted with proteins in the PtdIns(3,5)P2 machinery. Perturbation of PtdIns(3,5)P2 dynamics by overexpressing Fig4 or a PtdIns(3,5)P2-binding domain resulted in mislocalization of NSG1 to nonendosomal locations, and suppressing PtdIns(3,5)P2 synthesis resulted in an accumulation of NSG1 in EEA1-positive early endosomes. Importantly, overexpression of NSG1 rescued neurite thinning in PtdIns(3,5)P2-deficient CAD neurons and primary cortical neurons. Our study uncovered the role of PtdIns(3,5)P2 in the morphogenesis of neurons, which revealed a novel aspect of the pathogenesis of PtdIns(3,5)P2-related neuropathies. We also identified NSG1 as an important downstream protein of PtdIns(3,5)P2, which may provide a novel therapeutic target in neurological diseases.

Insulin/IGF signaling regulates presynaptic glutamate release in aversive olfactory learning.

Insulin/insulin-like growth factor (IGF) receptor signaling (IIS) supports context-dependent learning in vertebrates and invertebrates. Here, we identify cell-specific mechanisms of IIS that integrate sensory information with food context to drive synaptic plasticity and learning. In the nematode Caenorhabditis elegans, pairing food deprivation with an odor such as butanone suppresses attraction to that odor. We find that aversive olfactory learning requires the insulin receptor substrate (IRS) protein IST-1 and atypical signaling through the insulin/IGF-1 receptor DAF-2. Cell-specific knockout and rescue demonstrate that DAF-2 acts in the AWCON sensory neuron, which detects butanone, and that learning preferentially depends upon the axonally localized DAF-2c isoform. Acute food deprivation increases DAF-2 levels in AWCON post-transcriptionally through an insulin- and insulin receptor substrate-1 (ist-1)-dependent process. Aversive learning alters the synaptic output of AWCON by suppressing odor-regulated glutamate release in wild-type animals, but not in ist-1 mutants, suggesting that axonal insulin signaling regulates synaptic transmission to support aversive memory.

The role of neurotransmitter systems in mediating deep brain stimulation effects in Parkinson's disease.

Deep brain stimulation (DBS) is among the most successful paradigms in both translational and reverse translational neuroscience. DBS has developed into a standard treatment for movement disorders such as Parkinson’s disease (PD) in recent decades, however, specific mechanisms behind DBS’s efficacy and side effects remain unrevealed. Several hypotheses have been proposed, including neuronal firing rate and pattern theories that emphasize the impact of DBS on local circuitry but detail distant electrophysiological readouts to a lesser extent. Furthermore, ample preclinical and clinical evidence indicates that DBS influences neurotransmitter dynamics in PD, particularly the effects of subthalamic nucleus (STN) DBS on striatal dopaminergic and glutamatergic systems; pallidum DBS on striatal dopaminergic and GABAergic systems; pedunculopontine nucleus DBS on cholinergic systems; and STN-DBS on locus coeruleus (LC) noradrenergic system. DBS has additionally been associated with mood-related side effects within brainstem serotoninergic systems in response to STN-DBS. Still, addressing the mechanisms of DBS on neurotransmitters’ dynamics is commonly overlooked due to its practical difficulties in monitoring real-time changes in remote areas. Given that electrical stimulation alters neurotransmitter release in local and remote regions, it eventually exhibits changes in specific neuronal functions. Consequently, such changes lead to further modulation, synthesis, and release of neurotransmitters. This narrative review discusses the main neurotransmitter dynamics in PD and their role in mediating DBS effects from preclinical and clinical data.

L-Serine, an Endogenous Amino Acid, Is a Potential Neuroprotective Agent for Neurological Disease and Injury.

Central nervous system (CNS) lesions are major causes of human death and disability worldwide, and they cause different extents of motor and sensory dysfunction in patients. Thus, it is crucial to develop new effective neuroprotective drugs and approaches targeted to the heterogeneous nature of CNS injury and disease. L-serine is an indispensable neurotrophic factor and a precursor for neurotransmitters. Although L-serine is a native amino acid supplement, its metabolic products have been shown to be essential not only for cell proliferation but also for neuronal development and specific functions in the brain. Growing evidence has suggested that L-serine regulates the release of several cytokines in the brain under some neuropathological conditions to recover cognitive function, improve cerebral blood flow, inhibit inflammation, promote remyelination and exert other neuroprotective effects on neurological injury. L-serine has also been used to treat epilepsy, schizophrenia, psychosis, and Alzheimer’s Disease as well as other neurological diseases. Furthermore, the dosing of animals with L-serine and human clinical trials investigating the therapeutic effects of L-serine generally support the safety of L-serine. The high significance of this review lies in its emphasis on the therapeutic potential of using L-serine as a general treatment for numerous CNS diseases and injuries. Because L-serine performs a broad spectrum of functions, it may be clinically used as an effective neuroprotective agent.

Reciprocal regulation of spontaneous synaptic vesicle fusion by Fragile X mental retardation protein and group I metabotropic glutamate receptors.

Fragile X mental retardation protein (FMRP) is a neuronal protein mediating multiple functions, with its absence resulting in one of the most common monogenic causes of autism, Fragile X syndrome (FXS). Analyses of FXS pathophysiology have identified a range of aberrations in synaptic signaling pathways and plasticity associated with group I metabotropic glutamate (mGlu) receptors. These studies, however, have mostly focused on the post-synaptic functions of FMRP and mGlu receptor activation, and relatively little is known about their presynaptic effects. Neurotransmitter release is mediated via multiple forms of synaptic vesicle (SV) fusion, each of which contributes to specific neuronal functions. The impacts of mGlu receptor activation and loss of FMRP on these SV fusion events remain unexplored. Here we combined electrophysiological and fluorescence imaging analyses on primary hippocampal cultures prepared from an Fmr1 knockout (KO) rat model. Compared to wild-type (WT) hippocampal neurons, KO neurons displayed an increase in the frequency of spontaneous excitatory post-synaptic currents (sEPSCs), as well as spontaneous SV fusion events. Pharmacological activation of mGlu receptors in WT neurons caused a similar increase in spontaneous SV fusion and sEPSC frequency. Notably, this increase in SV fusion was not observed when spontaneous activity was blocked using the sodium channel antagonist tetrodotoxin. Importantly, the effect of mGlu receptor activation on spontaneous SV fusion was occluded in Fmr1 KO neurons. Together, our results reveal that FMRP represses spontaneous presynaptic SV fusion, whereas mGlu receptor activation increases this event. This reciprocal control appears to be mediated via their regulation of intrinsic neuronal excitability.

Fluorescence Correlation Spectroscopy Reveals Survival Motor Neuron Oligomerization but No Active Transport in Motor Axons of a Zebrafish Model for Spinal Muscular Atrophy.

Spinal Muscular Atrophy (SMA) is a progressive neurodegenerative disease affecting lower motor neurons that is caused by a deficiency in ubiquitously expressed Survival Motor Neuron (SMN) protein. Two mutually exclusive hypotheses have been discussed to explain increased motor neuron vulnerability in SMA. Reduced SMN levels have been proposed to lead to defective snRNP assembly and aberrant splicing of transcripts that are essential for motor neuron maintenance. An alternative hypothesis proposes a motor neuron-specific function for SMN in axonal transport of mRNAs and/or RNPs. To address these possibilities, we used a novel in vivo approach with fluorescence correlation spectroscopy (FCS) in transgenic zebrafish embryos to assess the subcellular dynamics of Smn in motor neuron cell bodies and axons. Using fluorescently tagged Smn we show that it exists as two freely diffusing components, a monomeric, and a complex-bound, likely oligomeric, component. This oligomer hypothesis was supported by the disappearance of the complex-bound form for a truncated Smn variant that is deficient in oligomerization and a change in its dynamics under endogenous Smn deficient conditions. Surprisingly, our FCS measurements did not provide any evidence for an active transport of Smn in axons. Instead, our in vivo observations are consistent with previous findings that SMN acts as a chaperone for the assembly of snRNP and mRNP complexes.

Physiology and Therapeutic Potential of SK, H, and M Medium AfterHyperPolarization Ion Channels.

SK, HCN, and M channels are medium afterhyperpolarization (mAHP)-mediating ion channels. The three channels co-express in various brain regions, and their collective action strongly influences cellular excitability. However, significant diversity exists in the expression of channel isoforms in distinct brain regions and various subcellular compartments, which contributes to an equally diverse set of specific neuronal functions. The current review emphasizes the collective behavior of the three classes of mAHP channels and discusses how these channels function together although they play specialized roles. We discuss the biophysical properties of these channels, signaling pathways that influence the activity of the three mAHP channels, various chemical modulators that alter channel activity and their therapeutic potential in treating various neurological anomalies. Additionally, we discuss the role of mAHP channels in the pathophysiology of various neurological diseases and how their modulation can alleviate some of the symptoms.

Phenotypic Characterization and Brain Structure Analysis of Calcium Channel Subunit α2δ-2 Mutant (Ducky) and α2δ Double Knockout Mice.

Auxiliary α2δ subunits of voltage-gated calcium channels modulate channel trafficking, current properties, and synapse formation. Three of the four isoforms (α2δ-1, α2δ-2, and α2δ-3) are abundantly expressed in the brain; however, of the available knockout models, only α2δ-2 knockout or mutant mice display an obvious abnormal neurological phenotype. Thus, we hypothesize that the neuronal α2δ isoforms may have partially specific as well as redundant functions. To address this, we generated three distinct α2δ double knockout mouse models by crossbreeding single knockout (α2δ-1 and -3) or mutant (α2δ-2/ducky) mice. Here, we provide a first phenotypic description and brain structure analysis. We found that genotypic distribution of neonatal litters in distinct α2δ-1/-2, α2δ-1/-3, and α2δ-2/-3 breeding combinations did not conform to Mendel’s law, suggesting premature lethality of single and double knockout mice. Notably, high occurrences of infant mortality correlated with the absence of specific α2δ isoforms (α2Δ-2 > α2δ-1 > α2δ-3), and was particularly observed in cages with behaviorally abnormal parenting animals of α2δ-2/-3 cross-breedings. Juvenile α2δ-1/-2 and α2δ-2/-3 double knockout mice displayed a waddling gate similar to ducky mice. However, in contrast to ducky and α2δ-1/-3 double knockout animals, α2δ-1/-2 and α2δ-2/-3 double knockout mice showed a more severe disease progression and highly impaired development. The observed phenotypes within the individual mouse lines may be linked to differences in the volume of specific brain regions. Reduced cortical volume in ducky mice, for example, was associated with a progressively decreased space between neurons, suggesting a reduction of total synaptic connections. Taken together, our findings show that α2δ subunits differentially regulate premature survival, postnatal growth, brain development, and behavior, suggesting specific neuronal functions in health and disease.

Synaptic processes and immune-related pathways implicated in Tourette syndrome

Tourette syndrome (TS) is a neuropsychiatric disorder of complex genetic architecture involving multiple interacting genes. Here, we sought to elucidate the pathways that underlie the neurobiology of the disorder through genome-wide analysis. We analyzed genome-wide genotypic data of 3581 individuals with TS and 7682 ancestry-matched controls and investigated associations of TS with sets of genes that are expressed in particular cell types and operate in specific neuronal and glial functions. We employed a self-contained, set-based association method (SBA) as well as a competitive gene set method (MAGMA) using individual-level genotype data to perform a comprehensive investigation of the biological background of TS. Our SBA analysis identified three significant gene sets after Bonferroni correction, implicating ligand-gated ion channel signaling, lymphocytic, and cell adhesion and transsynaptic signaling processes. MAGMA analysis further supported the involvement of the cell adhesion and trans-synaptic signaling gene set. The lymphocytic gene set was driven by variants in FLT3, raising an intriguing hypothesis for the involvement of a neuroinflammatory element in TS pathogenesis. The indications of involvement of ligand-gated ion channel signaling reinforce the role of GABA in TS, while the association of cell adhesion and trans-synaptic signaling gene set provides additional support for the role of adhesion molecules in neuropsychiatric disorders. This study reinforces previous findings but also provides new insights into the neurobiology of TS.

Neuron-Specific FMRP Roles in Experience-Dependent Remodeling of Olfactory Brain Innervation during an Early-Life Critical Period.

Critical periods are developmental windows during which neural circuits effectively adapt to the new sensory environment. Animal models of fragile X syndrome (FXS), a common monogenic autism spectrum disorder (ASD), exhibit profound impairments of sensory experience-driven critical periods. However, it is not known whether the causative fragile X mental retardation protein (FMRP) acts uniformly across neurons, or instead manifests neuron-specific functions. Here, we use the genetically-tractable Drosophila brain antennal lobe (AL) olfactory circuit of both sexes to investigate neuron-specific FMRP roles in the odorant experience-dependent remodeling of the olfactory sensory neuron (OSN) innervation during an early-life critical period. We find targeted OSN class-specific FMRP RNAi impairs innervation remodeling within AL synaptic glomeruli, whereas global dfmr1 null mutants display relatively normal odorant-driven refinement. We find both OSN cell autonomous and cell non-autonomous FMRP functions mediate odorant experience-dependent remodeling, with AL circuit FMRP imbalance causing defects in overall glomerulus innervation refinement. We find OSN class-specific FMRP levels bidirectionally regulate critical period remodeling, with odorant experience selectively controlling OSN synaptic terminals in AL glomeruli. We find OSN class-specific FMRP loss impairs critical period remodeling by disrupting responses to lateral modulation from other odorant-responsive OSNs mediating overall AL gain control. We find that silencing glutamatergic AL interneurons reduces OSN remodeling, while conversely, interfering with the OSN class-specific GABAA signaling enhances remodeling. These findings reveal control of OSN synaptic remodeling by FMRP with neuron-specific circuit functions, and indicate how neural circuitry can compensate for global FMRP loss to reinstate normal critical period brain circuit remodeling.SIGNIFICANCE STATEMENT Fragile X syndrome (FXS), the leading monogenic cause of intellectual disability and autism spectrum disorder (ASD), manifests severe neurodevelopmental delays. Likewise, FXS disease models display disrupted neurodevelopmental critical periods. In the well-mapped Drosophila olfactory circuit model, perturbing the causative fragile X mental retardation protein (FMRP) within a single olfactory sensory neuron (OSN) class impairs odorant-dependent remodeling during an early-life critical period. Importantly, this impairment requires activation of other OSNs, and the olfactory circuit can compensate when FMRP is removed from all OSNs. Understanding the neuron-specific FMRP requirements within a developing neural circuit, as well as the FMRP loss compensation mechanisms, should help us engineer FXS treatments. This work suggests FXS treatments could use homeostatic mechanisms to alleviate circuit-level deficits.

Rab43 GTPase directs postsynaptic trafficking and neuron-specific sorting of G protein–coupled receptors

G protein–coupled receptors (GPCRs) are important modulators of synaptic functions. A fundamental but poorly addressed question in neurobiology is how targeted GPCR trafficking is achieved. Rab GTPases are the master regulators of vesicle-mediated membrane trafficking, but their functions in the synaptic presentation of newly synthesized GPCRs are virtually unknown. Here, we investigate the role of Rab43, via dominant-negative inhibition and CRISPR–Cas9–mediated KO, in the export trafficking of α2-adrenergic receptor (α2-AR) and muscarinic acetylcholine receptor (mAChR) in primary neurons and cells. We demonstrate that Rab43 differentially regulates the overall surface expression of endogenous α2-AR and mAChR, as well as their signaling, in primary neurons. In parallel, Rab43 exerts distinct effects on the dendritic and postsynaptic transport of specific α2B-AR and M3 mAChR subtypes. More interestingly, the selective actions of Rab43 toward α2B-AR and M3 mAChR are neuronal cell specific and dictated by direct interaction. These data reveal novel, neuron-specific functions for Rab43 in the dendritic and postsynaptic targeting and sorting of GPCRs and imply multiple forward delivery routes for different GPCRs in neurons. Overall, this study provides important insights into regulatory mechanisms of GPCR anterograde traffic to the functional destination in neurons.

Associations between Neurological Diseases and Mutations in the Human Glycyl-tRNA Synthetase

Aminoacyl-RNA synthetases (aaRSs) are among the key enzymes of protein biosynthesis. They are responsible for conducting the first step in the protein biosynthesis, namely attaching amino acids to the corresponding tRNA molecules both in cytoplasm and mitochondria. More and more research demonstrates that mutations in the genes encoding aaRSs lead to the development of various neurodegenerative diseases, such as incurable Charcot–Marie–Tooth disease (CMT) and distal spinal muscular atrophy. Some mutations result in the loss of tRNA aminoacylation activity, while other mutants retain their classical enzyme activity. In the latter case, disease manifestations are associated with additional neuron-specific functions of aaRSs. At present, seven aaRSs (GlyRS, TyrRS, AlaRS, HisRS, TrpRS, MetRS, and LysRS) are known to be involved in the CMT etiology with glycyl-tRNA synthetase (GlyRS) being the most studied of them.

Repeated low-dose exposures to sarin disrupted the homeostasis of phospholipid and sphingolipid metabolism in guinea pig hippocampus

Repeated low-level exposure to sarin results to hippocampus dysfunction. Metabonomics involves a holistic analysis of a set of metabolites in an organism in the search for a relationship between these metabolites and physiological or pathological changes. The objective of the present study was to evaluate the effects of repeated exposure to low-level sarin on the metabonomics in hippocampus of a guinea pig model. Guinea pigs were divided randomly into control and sarin treated groups (n = 14). Guinea pigs in the control group received saline; while the sarin-treated group received 0.4×LD50 (16.8 μg/kg) sarin. Daily injections (a total of 14 days) were administered sc between the shoulder blades in a volume of 1.0 ml/kg body weight. At the end of the final injection, 6 animals in each group were chosen for Morris water maze test. The rest guinea pigs (n = 8 for each group) were sacrificed by decapitation, and hippocampus were dissected for analysis. Compared with the control-group, the escape latency in sarin-group was significantly (p < 0.05) longer while the crossing times were significantly decreased in the Morris water task (p < 0.05). Sarin inhibited activities of acetylcholinesterase (AChE) and neuropathy target esterase (NTE) in hippocampus. The AChE activity of hippocampus from sarin-treated groups is equivalent to 59.9 ± 6.4 %, and the NTE activity of hippocampus from sarin-groups is equivalent to 78.1 ± 8.3 % of that from control-group. Metabolites were identified and validated. A total of 14 variables were selected as potential biomarkers. Phospholipids [phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidic acid (PA), phosphatidylglycerol (PG), phosphatidylinositol (PI), Lysophosphatidylethanolamine (LysoPE or LPE)] and sphingolipids (SPs) [sphinganine (SA), phytosphingosine (PSO) and sphinganine-1-phosphate (SA1P)] were clearly modified. In conclusion, repeated low-dose exposures to sarin disrupted the homeostasis of phospholipid and sphingolipid metabolism in guinea pig hippocampus and may lead to a neuronal-specific function disorders. Identified metabolites such as SA1P need to be studied more deeply on their biological function that against sarin lesions. In future research, we should pay more attention to characterize the physiological roles of lipid metabolism enzymes as well as their involvement in pathologies induced by repeated low-level sarin exposure.

MicroRNAs Regulating Cytoskeleton Dynamics, Endocytosis, and Cell Motility-A Link Between Neurodegeneration and Cancer?

The cytoskeleton is one of the most mobile and complex cell structures. It is involved in cellular transport, cell division, cell shape formation and adaptation in response to extra- and intracellular stimuli, endo- and exocytosis, migration, and invasion. These processes are crucial for normal cellular physiology and are affected in several pathological processes, including neurodegenerative diseases, and cancer. Some proteins, participating in clathrin-mediated endocytosis (CME), play an important role in actin cytoskeleton reorganization, and formation of invadopodia in cancer cells and are also deregulated in neurodegenerative disorders. However, there is still limited information about the factors contributing to the regulation of their expression. MicroRNAs are potent negative regulators of gene expression mediating crosstalk between different cellular pathways in cellular homeostasis and stress responses. These molecules regulate numerous genes involved in neuronal differentiation, plasticity, and degeneration. Growing evidence suggests the role of microRNAs in the regulation of endocytosis, cell motility, and invasiveness. By modulating the levels of such microRNAs, it may be possible to interfere with CME or other processes to normalize their function. In malignancy, the role of microRNAs is undoubtful, and therefore changing their levels can attenuate the carcinogenic process. Here we review the current advances in our understanding of microRNAs regulating actin cytoskeleton dynamics, CME and cell motility with a special focus on neurodegenerative diseases, and cancer. We investigate whether current literature provides an evidence that microRNA-mediated regulation of essential cellular processes, such as CME and cell motility, is conserved in neurons, and cancer cells. We argue that more research effort should be addressed to study the neuron-specific functions on microRNAs. Disease-associated microRNAs affecting essential cellular processes deserve special attention both from the view of fundamental science and as future neurorestorative or anti-cancer therapies.