Neuronal Defects Research Articles

Methylation of lysine 36 on histone H3 is required to control transposon activities in somatic cells.

Transposable elements constitute a substantial portion of most eukaryotic genomes and their activity can lead to developmental and neuronal defects. In the germline, transposon activity is antagonized by the PIWI-interacting RNA pathway tasked with repression of transposon transcription and degrading transcripts that have already been produced. However, most of the genes required for transposon control are not expressed outside the germline, prompting the question: what causes deleterious transposons activity in the soma and how is it managed? Here, we show that disruptions of the Histone 3 lysine 36 methylation machinery led to increased transposon transcription in Drosophila melanogaster brains and that there is division of labour for the repression of transposable elements between the different methyltransferases Set2, NSD, and Ash1. Furthermore, we show that disruption of methylation leads to somatic activation of key genes in the PIWI-interacting RNA pathway and the preferential production of RNA from dual-strand piRNA clusters.

O-GlcNAcylation is crucial for sympathetic neuron development, maintenance, functionality and contributes to peripheral neuropathy.

O-GlcNAcylation is a post-translational modification (PTM) that regulates a wide range of cellular functions and has been associated with multiple metabolic diseases in various organs. The sympathetic nervous system (SNS) is the efferent portion of the autonomic nervous system that regulates metabolism of almost all organs in the body. How much the development and functionality of the SNS are influenced by O-GlcNAcylation, as well as how such regulation could contribute to sympathetic neuron (symN)-related neuropathy in diseased states, remains unknown. Here, we assessed the level of protein O-GlcNAcylation at various stages of symN development, using a human pluripotent stem cell (hPSC)-based symN differentiation paradigm. We found that pharmacological disruption of O-GlcNAcylation impaired both the growth and survival of hPSC-derived symNs. In the high glucose condition that mimics hyperglycemia, hPSC-derived symNs were hyperactive, and their regenerative capacity was impaired, which resembled typical neuronal defects in patients and animal models of diabetes mellitus. Using this model of sympathetic neuropathy, we discovered that O-GlcNAcylation increased in symNs under high glucose, which lead to hyperactivity. Pharmacological inhibition of O-GlcNAcylation rescued high glucose-induced symN hyperactivity and cell stress. This framework provides the first insight into the roles of O-GlcNAcylation in both healthy and diseased human symNs and may be used as a platform for therapeutic studies.

Molinate induces organ defects by promoting apoptosis, inflammation, and endoplasmic reticulum stress during the developmental stage of zebrafish

Molinate is classified as a thiocarbamate herbicide and is mainly used in paddy fields to culture rice. However, the toxic effects of molinate and the associated mechanisms in the process of development have not been completely elucidated. Therefore, in the present study, we demonstrated that molinate reduced the viability of zebrafish larvae and the probability of successful hatching using zebrafish (Danio rerio), one of the remarkable in vivo models for testing the toxicity of chemicals. In addition, molinate treatment triggered the occurrence of apoptosis, inflammation, and endoplasmic reticulum (ER) stress response in zebrafish larvae. Furthermore, we identified that an abnormal cardiovascular phenotype through wild type zebrafish, neuronal defects through transgenic olig2:dsRed zebrafish, and developmental toxicity in the liver through transgenic lfabp:dsRed zebrafish. Collectively, these results provide evidence of the hazardous effects of molinate on the developmental stage of non-target organisms by elucidating the toxic mechanisms of molinate in developing zebrafish.

C.elegans vab-6 encodes a KIF3A kinesin and functions cell non-autonomously to regulate epidermal morphogenesis.

Cells undergo strict regulation to develop their shape in a process called morphogenesis. Caenorhabditis elegans with mutations in the variable abnormal (vab) class of genes have been shown to display epidermal and neuronal morphological defects. While several vab genes have been well-characterized, the function of the vab-6 gene remains unknown. Here, we show that vab-6 is synonymous with a subunit of the kinesin-II heterotrimeric motor complex called klp-20/Kif3a, a motor well-understood to be involved in developing sensory cilia in the nervous system. We show that certain klp-20 alleles cause animals to develop a bumpy body phenotype that is variable but most severe in mutants containing single amino-acid substitutions in the catalytic head-domain sites of the protein. Surprisingly, animals carrying a klp-20 null allele do not show the bumpy epidermal phenotype suggesting genetic redundancy and only when mutant versions of the KLP-20 protein are present, the epidermal phenotype is observed. The bumpy epidermal phenotype was not observed in other kinesin-2 mutants, suggesting that KLP-20 is functioning independently from its role in intraflagellar transport (IFT) during ciliogenesis. Interestingly, despite having such a prominent epidermal phenotype, KLP-20 is not expressed in the epidermis, strongly suggesting a cell non-autonomous role in which it regulates epidermal morphogenesis.

Silver Nanoparticles Cause Neural and Vascular Disruption by Affecting Key Neuroactive Ligand-Receptor Interaction and VEGF Signaling Pathways.

Silver nanoparticles (AgNP) are widely used as coating materials. However, the potential risks of AgNP to human health, especially for neural and vascular systems, are still poorly understood. The vascular and neurotoxicity of various concentrations of AgNP in zebrafish were examined using fluorescence microscopy. In addition, Illumina high-throughput global transcriptome analysis was performed to explore the transcriptome profiles of zebrafish embryos after exposure to AgNP. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were conducted to elucidate the top 3000 differentially expressed genes (DEGs) between AgNP-exposed and control groups. We systematically investigated the neural and vascular developmental toxicities of AgNP exposure in zebrafish. The results demonstrated that AgNP exposure could cause neurodevelopmental anomalies, including a small-eye phenotype, neuronal morphology defects, and inhibition of athletic abilities. In addition, we found that AgNP exposure induces angiogenesis malformation in zebrafish embryos. Further RNA-seq revealed that DEGs were mainly enriched in the neuroactive ligand-receptor interaction and vascular endothelial growth factor (Vegf) signaling pathways in AgNP-treated zebrafish embryos. Specifically, the mRNA levels of the neuroactive ligand-receptor interaction pathway and Vegf signaling pathway-related genes, including si:ch73-55i23.1, nfatc2a, prkcg, si:ch211-132p1.2, lepa, mchr1b, pla2g4aa, rac1b, p2ry6, adrb2, chrnb1, and chrm1b, were significantly regulated in AgNP-treated zebrafish embryos. Our findings indicate that AgNP exposure transcriptionally induces developmental toxicity in neural and vascular development by disturbing neuroactive ligand-receptor interactions and the Vegf signaling pathway in zebrafish embryos.

Neuronal Membrane Repair Defect as a Contributor to Amyloid Beta Pathogenesis in Alzheimer's Disease

Alzheimer’s Disease (AD) is a common neurodegenerative disease that displays two hallmark protein accumulations in the brain – extracellular amyloid beta (Aβ) and intracellular hyperphosphorylated tau. Aβ has been shown to directly penetrate the neuronal cell membrane and damage the membrane. This compromises membrane integrity, increases membrane permeability that alters membrane conductance and elevates intracellular calcium concentrations which contribute to reactive oxygen species production and mitochondrial dysfunction. This damage to the plasma membrane requires a quick and efficient repair mechanism to restore the barrier function of the membrane and avoid neuronal cell death. However, a defect in membrane repair has not been examined as a contributor to AD. Here, we aimed to assess membrane repair capacity in AD in vitro models as a potential contributor to neuronal death. To assess the effect of Aβ on membrane repair kinetics, we exposed primary mouse hippocampal neurons to 1μM recombinant human Aβ42 (rhAβ42) for 16 hours prior to experimentation. Additionally, we treated wildtype neuronal cells with healthy and AD patient with elevated levels of Aβ (PSEN1 M146L mutation) cortical neuron iPSC conditioned media. We also exposed primary mouse hippocampal neurons to cerebrospinal fluid (CSF) from non-AD and AD patients. Treated cells were challenged with an established laser damage assay where a two-photon laser is used to damage the cell membrane in the presence of FM4-64 dye. Dye fluorescence at the injury site is used as a proxy for membrane repair kinetics. Lastly, we investigated autoantibody production targeting proteins involved in membrane repair in non-AD and AD sera and CSF via custom ELISAs. Overall, we observed an induced membrane repair defect when wildtype neuronal cells are treated with rhAβ42, AD iPSC conditioned media, and AD CSF. We also observed a significant increase in autoantibody production in AD biofluids as compared to the non-AD controls. Autoantibody levels were inversely correlated with efficient membrane repair capacity and positively correlated with Aβ concentrations. These results illustrate Aβ’s role in impairing neuronal membrane repair capacity, and the potential contribution of autoantibodies targeting membrane repair proteins in AD. This work is supported by The Ohio State University Dean's Discovery grant and Dept. of Physiology & Cell Biology Nishikawara Student Research grant. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Comprehensive Transformation of Escherichia coli for Nicotinamide Mononucleotide Production

Nicotinamide mononucleotide (NMN) is a key precursor of nicotinamide adenine dinucleotide and an important source of cellular energy. It can prevent neuronal mitochondrial defects and alleviate heart fibrosis. Strategies to improve NMN production have important implications for human health. Through plasmid expression technology and CRISPR/Cas9 technology, we engineered Escherichia coli for efficient NMN production. First, we upregulated the expression of genes encoding key enzymes in the NMN synthesis pathway, enabling E. coli to directly produce NMN, and established the important role of the nicotinamide mononucleotide transporter in the transport of NMN from cells. The content of NMN was 0.24 g·L−1 at 24 h. Second, we strengthened the adenosine triphosphate (ATP) cycle, and the concentration of NMN was 0.49 g·L−1 at 24 h. Third, we increased the synthesis of the NMN precursor 5-phosphate ribose-1-phosphate and obtained an NMN content of 0.49 g·L−1 at 12 h and 1.11 g·L−1 at 24 h. Fourth, we introduced nicotinamide riboside kinase (NRK) and found that it was effective only for a period of time. The content of NMN was 0.54 g·L−1 at 12 h but only 1.05 g·L−1 at 24 h. Finally, we combined these strategies to regulate the whole metabolic flow, revealing that integrating multiple pathways promoted NMN production. During fermentation, we added 1 g·L−1 nicotinamide and 10 g·L−1 glucose, yielding an extracellular NMN concentration of 1.11 g·L−1.

Human Cytomegalovirus IE1 Impairs Neuronal Migration by Downregulating Connexin 43.

Human cytomegalovirus (HCMV) is a leading cause of congenital birth defects. Though the underlying mechanisms remain poorly characterized, mouse models of congenital CMV infection have demonstrated that the neuronal migration process is damaged. In this study, we evaluated the effects of HCMV infection on connexin 43 (Cx43), a crucial adhesion molecule mediating neuronal migration. We show in multiple cellular models that HCMV infection downregulated Cx43 posttranslationally. Further analysis identified the immediate early protein IE1 as the viral protein responsible for the reduction of Cx43. IE1 was found to bind the Cx43 C terminus and promote Cx43 degradation through the ubiquitin-proteasome pathway. Deletion of the Cx43-binding site in IE1 rendered it incapable of inducing Cx43 degradation. We validated the IE1-induced loss of Cx43 in vivo by introducing IE1 into the fetal mouse brain. Noteworthily, ectopic IE1 expression induced cortical atrophy and neuronal migration defects. Several lines of evidence suggest that these damages result from decreased Cx43, and restoration of Cx43 levels partially rescued IE1-induced interruption of neuronal migration. Taken together, the results of our investigation reveal a novel mechanism of HCMV-induced neural maldevelopment and identify a potential intervention target. IMPORTANCE Congenital CMV (cCMV) infection causes neurological sequelae in newborns. Recent studies of cCMV pathogenesis in animal models reveal ventriculomegaly and cortical atrophy associated with impaired neural progenitor cell (NPC) proliferation and migration. In this study, we investigated the mechanisms underlying these NPC abnormalities. We show that Cx43, a critical adhesion molecule mediating NPC migration, is downregulated by HCMV infection in vitro and HCMV-IE1 in vivo. We provide evidence that IE1 interacts with the C terminus of Cx43 to promote its ubiquitination and consequent degradation through the proteasome. Moreover, we demonstrate that introducing IE1 into mouse fetal brains led to neuronal migration defects, which was associated with Cx43 reduction. Deletion of the Cx43-binding region in IE1 or ectopic expression of Cx43 rescued the IE1-induced migration defects in vivo. Our study provides insight into how cCMV infection impairs neuronal migration and reveals a target for therapeutic interventions.

Nifedipine Ameliorates Cellular Differentiation Defects of Smn-Deficient Motor Neurons and Enhances Neuromuscular Transmission in SMA Mice.

In spinal muscular atrophy (SMA), mutations in or loss of the Survival Motor Neuron 1 (SMN1) gene reduce full-length SMN protein levels, which leads to the degeneration of a percentage of motor neurons. In mouse models of SMA, the development and maintenance of spinal motor neurons and the neuromuscular junction (NMJ) function are altered. Since nifedipine is known to be neuroprotective and increases neurotransmission in nerve terminals, we investigated its effects on cultured spinal cord motor neurons and motor nerve terminals of control and SMA mice. We found that application of nifedipine increased the frequency of spontaneous Ca2+ transients, growth cone size, cluster-like formations of Cav2.2 channels, and it normalized axon extension in SMA neurons in culture. At the NMJ, nifedipine significantly increased evoked and spontaneous release at low-frequency stimulation in both genotypes. High-strength stimulation revealed that nifedipine increased the size of the readily releasable pool (RRP) of vesicles in control but not SMA mice. These findings provide experimental evidence about the ability of nifedipine to prevent the appearance of developmental defects in SMA embryonic motor neurons in culture and reveal to which extent nifedipine could still increase neurotransmission at the NMJ in SMA mice under different functional demands.

Synaptojanin1 Modifies Endolysosomal Parameters in Cultured Ventral Midbrain Neurons.

The accumulation of α-synuclein (α-syn)-enriched protein aggregates is thought to arise from dysfunction in degradation systems within the brain. Recently, missense mutations of SYNJ1 encoding the SAC1 and 5'-phosphatase domains have been found in families with hereditary early-onset Parkinsonism. Previous studies showed that Synj1 haploinsufficiency (Synj1+/-) leads to accumulation of the autophagy substrate p62 and pathologic α-syn proteins in the midbrain (MB) and striatum of aged mice. In this study, we aim to investigate the neuronal degradation pathway using the Synj1+/- MB culture from mouse pups of mixed sex as a model. Our data show that GFP-LC3 puncta formation and cumulative mKeima puncta formation are unaltered at baseline in Synj1+/- MB neurons. However, GFP-LAMP1 puncta is reduced with a similar decrease in endogenous proteins, including lysosomal-associated membrane protein (LAMP)1, LAMP2, and LAMP2A. The LAMP1 vesicles are hyperacidified with enhanced enzymatic activity in Synj1+/- MB neurons. Using a combination of light and electron microscopy (EM), we show that endolysosomal changes are primarily associated with a lack of SAC1 activity. Consistently, expressing the SYNJ1 R258Q mutant in N2a cells reduces the lysosome number. Interestingly, the endolysosomal defects in Synj1+/- neurons does not impact the clearance of exogenously expressed wild-type (WT) α-syn; however, the clearance of α-syn A53T was impaired in the axons of Synj1+/- MB neurons. Taken together, our results suggest axonal vulnerability to endolysosomal defects in Synj1-deficient MB neurons.

ROP16-mediated activation of STAT6 enhances cyst development of type III Toxoplasma gondii in neurons.

Toxoplasma gondii establishes a long-lived latent infection in the central nervous system (CNS) of its hosts. Reactivation in immunocompromised individuals can lead to life threatening disease. Latent infection is driven by the ability of the parasite to convert from the acute-stage tachyzoite to the latent-stage bradyzoite which resides in long-lived intracellular cysts. While much work has focused on the parasitic factors that drive cyst development, the host factors that influence encystment are not well defined. Here we show that a polymorphic secreted parasite kinase (ROP16), that phosphorylates host cell proteins, mediates efficient encystment of T. gondii in a stress-induced model of encystment and primary neuronal cell cultures (PNCs) in a strain-specific manner. Using short-hairpin RNA (shRNA) knockdowns in human foreskin fibroblasts (HFFs) and PNCs from transgenic mice, we determined that ROP16's cyst enhancing abilities are mediated, in part, by phosphorylation-and therefore activation-of the host cell transcription factor STAT6. To test the role of STAT6 in vivo, we infected wild-type (WT) and STAT6KO mice, finding that, compared to WT mice, STAT6KO mice have a decrease in CNS cyst burden but not overall parasite burden or dissemination to the CNS. Finally, we found a similar ROP16-dependent encystment defect in human pluripotent stem cell-derived neurons. Together, these findings identify a host cell factor (STAT6) that T. gondii manipulates in a strain-specific manner to generate a favorable encystment environment.

Cis mutagenesis in vivo reveals extensive noncanonical functions of Dscam1 isoforms in neuronal wiring.

Drosophila Down syndrome cell adhesion molecule 1 (Dscam1) encodes tens of thousands of cell recognition molecules via alternative splicing, which are required for neural function. A canonical self-avoidance model seems to provide a central mechanistic basis for Dscam1 functions in neuronal wiring. Here, we reveal extensive noncanonical functions of Dscam1 isoforms in neuronal wiring. We generated a series of allelic cis mutations in Dscam1, encoding a normal number of isoforms, but with an altered isoform composition. Despite normal dendritic self-avoidance and self-/nonself-discrimination in dendritic arborization (da) neurons, which is consistent with the canonical self-avoidance model, these mutants exhibited strikingly distinct spectra of phenotypic defects in the three types of neurons: up to ∼60% defects in mushroom bodies, a significant increase in branching and growth in da neurons, and mild axonal branching defects in mechanosensory neurons. Remarkably, the altered isoform composition resulted in increased dendrite growth yet inhibited axon growth. Moreover, reducing Dscam1 dosage exacerbated axonal defects in mushroom bodies and mechanosensory neurons but reverted dendritic branching and growth defects in da neurons. This splicing-tuned regulation strategy suggests that axon and dendrite growth in diverse neurons cell-autonomously require Dscam1 isoform composition. These findings provide important insights into the functions of Dscam1 isoforms in neuronal wiring.

Diminished Neuronal ESCRT-0 Function Exacerbates AMPA Receptor Derangement and Accelerates Prion-Induced Neurodegeneration.

Endolysosomal defects in neurons are central to the pathogenesis of prion and other neurodegenerative disorders. In prion disease, prion oligomers traffic through the multivesicular body (MVB) and are routed for degradation in lysosomes or for release in exosomes, yet how prions impact proteostatic pathways is unclear. We found that prion-affected human and mouse brain showed a marked reduction in Hrs and STAM1 (ESCRT-0), which route ubiquitinated membrane proteins from early endosomes into MVBs. To determine how the reduction in ESCRT-0 impacts prion conversion and cellular toxicity in vivo, we prion-challenged conditional knockout mice (male and female) having Hrs deleted from neurons, astrocytes, or microglia. The neuronal, but not astrocytic or microglial, Hrs-depleted mice showed a shortened survival and an acceleration in synaptic derangements, including an accumulation of ubiquitinated proteins, deregulation of phosphorylated AMPA and metabotropic glutamate receptors, and profoundly altered synaptic structure, all of which occurred later in the prion-infected control mice. Finally, we found that neuronal Hrs (nHrs) depletion increased surface levels of the cellular prion protein, PrPC, which may contribute to the rapidly advancing disease through neurotoxic signaling. Taken together, the reduced Hrs in the prion-affected brain hampers ubiquitinated protein clearance at the synapse, exacerbates postsynaptic glutamate receptor deregulation, and accelerates neurodegeneration.SIGNIFICANCE STATEMENT Prion diseases are rapidly progressive neurodegenerative disorders characterized by prion aggregate spread through the central nervous system. Early disease features include ubiquitinated protein accumulation and synapse loss. Here, we investigate how prion aggregates alter ubiquitinated protein clearance pathways (ESCRT) in mouse and human prion-infected brain, discovering a marked reduction in Hrs. Using a prion-infection mouse model with neuronal Hrs (nHrs) depleted, we show that low neuronal Hrs is detrimental and markedly shortens survival time while accelerating synaptic derangements, including ubiquitinated protein accumulation, indicating that Hrs loss exacerbates prion disease progression. Additionally, Hrs depletion increases the surface distribution of prion protein (PrPC), linked to aggregate-induced neurotoxic signaling, suggesting that Hrs loss in prion disease accelerates disease through enhancing PrPC-mediated neurotoxic signaling.

Deregulation of ER-mitochondria contact formation and mitochondrial calcium homeostasis mediated by VDAC in fragile X syndrome.

Loss of fragile X messenger ribonucleoprotein (FMRP) causes fragile X syndrome (FXS), the most prevalent form of inherited intellectual disability. Here, we show that FMRP interacts with the voltage-dependent anion channel (VDAC) to regulate the formation and function of endoplasmic reticulum (ER)-mitochondria contact sites (ERMCSs), structures that are critical for mitochondrial calcium (mito-Ca2+) homeostasis. FMRP-deficient cells feature excessive ERMCS formation and ER-to-mitochondria Ca2+ transfer. Genetic and pharmacological inhibition of VDAC or other ERMCS components restored synaptic structure, function, and plasticity and rescued locomotion and cognitive deficits of the Drosophila dFmr1 mutant. Expressing FMRP C-terminal domain (FMRP-C), which confers FMRP-VDAC interaction, rescued the ERMCS formation and mito-Ca2+ homeostasis defects in FXS patient iPSC-derived neurons and locomotion and cognitive deficits in Fmr1 knockout mice. These results identify altered ERMCS formation and mito-Ca2+ homeostasis as contributors to FXS and offer potential therapeutic targets.

Neuronal biomolecular condensates and their implications in neurodegenerative diseases.

Biomolecular condensates are subcellular organizations where functionally related proteins and nucleic acids are assembled through liquid-liquid phase separation, allowing them to develop on a larger scale without a membrane. However, biomolecular condensates are highly vulnerable to disruptions from genetic risks and various factors inside and outside the cell and are strongly implicated in the pathogenesis of many neurodegenerative diseases. In addition to the classical view of the nucleation-polymerization process that triggers the protein aggregation from the misfolded seed, the pathologic transition of biomolecular condensates can also promote the aggregation of proteins found in the deposits of neurodegenerative diseases. Furthermore, it has been suggested that several protein or protein-RNA complexes located in the synapse and along the neuronal process are neuron-specific condensates displaying liquid-like properties. As their compositional and functional modifications play a crucial role in the context of neurodegeneration, further research is needed to fully understand the role of neuronal biomolecular condensates. In this article, we will discuss recent findings that explore the pivotal role of biomolecular condensates in the development of neuronal defects and neurodegeneration.

The synthetic TRPML1 agonist ML-SA1 rescues Alzheimer-related alterations of the endosomal-autophagic-lysosomal system.

Abnormalities in the endosomal-autophagic-lysosomal (EAL) system are an early event in Alzheimer's disease (AD) pathogenesis. However, the mechanisms underlying these abnormalities are unclear. The transient receptor potential channel mucolipin 1(TRPML1, also known as MCOLN1), a vital endosomal-lysosomal Ca2+ channel whose loss of function leads to neurodegeneration, has not been investigated with respect to EAL pathogenesis in late-onset AD (LOAD). Here, we identify pathological hallmarks of TRPML1 dysregulation in LOAD neurons, including increased perinuclear clustering and vacuolation of endolysosomes. We reveal that induced pluripotent stem cell (iPSC)-derived human cortical neurons expressing APOE ε4, the strongest genetic risk factor for LOAD, have significantly diminished TRPML1-induced endolysosomal Ca2+ release. Furthermore, we found that blocking TRPML1 function in primary neurons by depleting the TRPML1 agonist PI(3,5)P2 via PIKfyve inhibition, recreated multiple features of EAL neuropathology evident in LOAD. This included increased endolysosomal Ca2+ content, enlargement and perinuclear clustering of endolysosomes, autophagic vesicle accumulation and early endosomal enlargement. Strikingly, these AD-like neuronal EAL defects were rescued by TRPML1 reactivation using its synthetic agonist ML-SA1. These findings implicate defects in TRPML1 in LOAD EAL pathogenesis and present TRPML1 as a potential therapeutic target.

Dysregulation of ribosome-associated quality control elicits cognitive disorders via overaccumulation of TTC3

Ribosome-associated quality control (RQC) pathway is responsible for degradation of nascent polypeptides in aberrantly stalled ribosomes, and its defects may lead to neurological diseases. However, the underlying molecular mechanism of how RQC dysfunction elicits neurological disorders remains poorly understood. Here we revealed that neurons with knockout (KO) of ubiquitin ligase LTN1, a key gene in the RQC pathway, show developmental defects in neurons via upregulation of TTC3 and UFMylation signaling proteins. The abnormally enhanced TTC3 protein in Ltn1 KO neurons reduced further accumulation of translationally arrested products by preventing translation initiation of selective genes. However, the overaccumulated TTC3 protein in turn caused dendritic abnormalities and reduced surface-localized GABAA receptors during neuronal development. Ltn1 KO mice showed behavioral deficits associated with cognitive disorders, a subset of which were restored by TTC3 knockdown in medial prefrontal cortex. Together, the overactivated cellular compensatory mechanism against defective RQC through TTC3 overaccumulation induced synaptic and cognitive deficits. More broadly, these findings represent a novel cellular mechanism underlying neuronal dysfunctions triggered by exaggerated cellular stress response to accumulated abnormal translation products in neurons.

Cross-Sectional Study of Assessment of Predominance of Prakrutiamong Patients with Diabetic Neuropathy in Wardha City

India is a developing country, so much of the Indian subcontinent are converting into urban areas, which lead to changes in lifestyle involving major tourist in battery plateau physical activities and increasing mental stress, which leads to insulin inertia. Approximately 246 million people worldwide have diabetes mellitus. It is estimated that 20 to 30 million people globally suffer from symptomatic diabetic neuropathy. The prevalence of diabetic neuropathy also rises over time. If type 1 diabetes is not well controlled, severe diabetic polyneuropathy and poor glycemic control can appear in young adults just a few months after the disease manifests. Ayurveda can be correlated with MajjaDhatuDushti (Structural and functional defects of neurons). Identification of the predominance of a specific type of Prakrutiin patients with diabetic neuropathy in Wardha City. The prime objectives of the present study are to find out the predominance of specific Prakriti in patients with diabetic neuropathy in Wardhacity&create awareness in them regarding complications of Diabetes mellitus, especially diabetic neuropathy and its relation with Prakruti. The current study was a cross-sectional study based on a personal interview of patients and Prakrutiquestionnaires. It occurred at Salod(H), Wardha, Maharashtra's Mahatma Gandhi Ayurved College, Hospital & Research Center. It was found that the prevalence of diabetic neuropathy is high in PittajPradhanVataPrakruti. In this present study, most of the affected patients were male. The present study concludes that diabetic neuropathy was found more prevalently in patients with Pitta PradhanVataPrakruti. Specific lifestyle modifications can be recommended for patients with Prakriti, as mentioned earlier, to prevent diabetic neuropathy in the future. Keywords: , , , ,

MED13L and its disease-associated variants influence the dendritic development of cerebral cortical neurons in the mammalian brain.

The mediator complex comprises multiple subcellular subunits that collectively function as a molecular interface between RNA polymerase II and gene-specific transcription factors. Recently, genetic variants to one subunit of the complex, known as MED13L (mediator complex subunit 13 like), have been implicated in syndromic intellectual disability and distinct facial features, frequently accompanied by congenital heart defects. We investigated the impact of five disease-associated MED13L variants on the subcellular localization and biochemical stability of MED13L protein in vitro and in vivo. In overexpression assays using cortical neurons from embryonic mouse cerebral cortices transduced by in utero electroporation-mediated gene transfer, we found that mouse orthologues of human MED13L-p.P866L and -p.T2162M missense variants accumulated in the nucleus, while the p.S2163L and p.S2177Y variants were diffusely distributed in the cytoplasm. In contrast, we found that the p.Q1922* truncation variant was barely detectable in transduced cells, a phenotype reminiscent of this variant that results in MED13L haploinsufficiency in humans. Next, we analyzed these variants for their effects on neuronal migration, dendritic growth, spine morphology, and axon elongation of cortical neurons in vivo. There, we found that overexpression of the p.P866L variant resulted in reduced number and length of dendrites of cortical layer II/III pyramidal neurons. Furthermore, we show that mMED13L-knockdown abrogated dendritic growth in vivo, and this effect was significantly rescued by co-electroporation of an RNAi-resistant mMED13L, but weakly by the p.T2162M variant, and not at all by the p.S2163L variant. However, overexpression of the p.S2163L variant inhibited mature dendritic spine formation in vivo. Expression of each of the 5 variants did not affect neuronal cell migration and callosal axon elongation in vivo. Taken together, our results demonstrate that MED13L expression is relevant to corticogenesis and influences the dendritic branching characteristics of cortical excitatory neurons. Our study also suggests that disease-associated MED13L variants may directly cause morphological and functional defects in cortical neurons in different ways.

Triose-phosphate isomerase deficiency is associated with a dysregulation of synaptic vesicle recycling in Drosophila melanogaster.

Numerous neurodegenerative diseases are associated with neuronal dysfunction caused by increased redox stress, often linked to aberrant production of redox-active molecules such as nitric oxide (NO) or oxygen free radicals. One such protein affected by redox-mediated changes is the glycolytic enzyme triose-phosphate isomerase (TPI), which has been shown to undergo 3-nitrotyrosination (a NO-mediated post-translational modification) rendering it inactive. The resulting neuronal changes caused by this modification are not well understood. However, associated glycation-induced cytotoxicity has been reported, thus potentially causing neuronal and synaptic dysfunction via compromising synaptic vesicle recycling. This work uses Drosophila melanogaster to identify the impacts of altered TPI activity on neuronal physiology, linking aberrant TPI function and redox stress to neuronal defects. We used Drosophila mutants expressing a missense allele of the TPI protein, M81T, identified in a previous screen and resulting in an inactive mutant of the TPI protein (TPIM81T , wstd1). We assessed synaptic physiology at the glutamatergic Drosophila neuromuscular junction (NMJ), synapse morphology and behavioural phenotypes, as well as impacts on longevity. Electrophysiological recordings of evoked and spontaneous excitatory junctional currents, alongside high frequency train stimulations and recovery protocols, were applied to investigate synaptic depletion and subsequent recovery. Single synaptic currents were unaltered in the presence of the wstd1 mutation, but frequencies of spontaneous events were reduced. Wstd1 larvae also showed enhanced vesicle depletion rates at higher frequency stimulation, and subsequent recovery times for evoked synaptic responses were prolonged. A computational model showed that TPI mutant larvae exhibited a significant decline in activity-dependent vesicle recycling, which manifests itself as increased recovery times for the readily-releasable vesicle pool. Confocal images of NMJs showed no morphological or developmental differences between wild-type and wstd1 but TPI mutants exhibited learning impairments as assessed by olfactory associative learning assays. Our data suggests that the wstd1 phenotype is partially due to altered vesicle dynamics, involving a reduced vesicle pool replenishment, and altered endo/exocytosis processes. This may result in learning and memory impairments and neuronal dysfunction potentially also presenting a contributing factor to other reported neuronal phenotypes.