76 research outputs found
UNC-13 Interaction with Syntaxin Is Required for Synaptic Transmission
SummaryNeurotransmitter secretion at synapses is controlled by several processesâmorphological docking of vesicles at release sites, priming of docked vesicles to make them fusion competent, and calcium-dependent fusion of vesicles with the plasma membrane [1, 2]. In worms, flies, and mice, mutants lacking UNC-13 have defects in vesicle priming [3â5]. Current models propose that UNC-13 primes vesicles by stabilizing Syntaxin's âopenâ conformation by directly interacting with its amino-terminal regulatory domain [6â8]. However, the functional significance of the UNC-13/Syntaxin interaction has not been tested directly. A truncated protein containing the Munc homology domains (MHD1 and MHD2) and the carboxy-terminal C2 domain partially rescued both the behavioral and secretion defects of unc-13 mutants in C. elegans. A double mutation in MHD2 (F1000A/K1002A) disrupts the UNC-13/Syntaxin interaction. The rate of endogenous synaptic events and the amplitude of nerve-evoked excitatory post-synaptic currents (EPSCs) were both significantly reduced in UNC-13S(F1000A/K1002A). However, the pool of primed (i.e., fusion-competent) vesicles was normal. These results suggest that the UNC-13/Syntaxin interaction is conserved in C. elegans and that, contrary to current models, the UNC-13/Syntaxin interaction is required for nerve-evoked vesicle fusion rather than synaptic-vesicle priming. Thus, UNC-13 may regulate multiple steps of the synaptic-vesicle cycle
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Permafrost Formation in a Meandering River Floodplain
Permafrost influences 25% of land in the Northern Hemisphere, where it stabilizes the ground beneath communities and infrastructure and sequesters carbon. However, the coevolution of permafrost, river dynamics, and vegetation in Arctic environments remains poorly understood. As rivers meander, they erode the floodplain at cutbanks and build new land through bar deposition, creating sequences of landforms with distinct formation ages. Here we mapped these sequences along the Koyukuk River floodplain, Alaska, analyzing permafrost occurrence, and landform and vegetation types. We used radiocarbon and optically stimulated luminescence (OSL) dating to develop a floodplain age map. Deposit ages ranged from modern to 10 ka, with more younger deposits near the modern channel. Permafrost rapidly reached 50% areal extent in all deposits older than 200 years then gradually increased up to âź85% extent for deposits greater than 4 Kyr old. Permafrost extent correlated with increases in black spruce and wetland abundance, as well as increases in permafrost extent within wetland, and shrub and scrub vegetation classes. We developed an inverse model to constrain permafrost formation rate as a function of air temperature. Permafrost extent initially increased by âź25% per century, in pace with vegetation succession, before decelerating to <10% per millennia as insulating overbank mud and moss slowly accumulated. Modern permafrost extent on the Koyukuk floodplain therefore reflects a dynamic balance between widespread, time-varying permafrost formation and rapid, localized degradation due to cutbank erosion that might trigger a rapid loss of permafrost with climatic warming
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Cis-Acting Regulation of Brain-Specific ANK3 Gene Expression by a Genetic Variant Associated with Bipolar Disorder
Several genome-wide association studies (GWAS) for bipolar disorder (BD) have found a strong association of the Ankyrin3 (ANK3) gene. This association spans numerous linked single nucleotide polymorphisms (SNPs) in a ~250 kb genomic region overlapping ANK3. The associated region encompasses predicted regulatory elements as well as two of six validated alternative first exons, which encode distinct protein domains at the N-terminus of the protein also known as ankyrin-G (AnkG). Using RNA Ligase-Mediated Rapid Amplification of cDNA Ends (RLM-RACE) to identify novel transcripts in conjunction with a highly sensitive, exon-specific multiplexed mRNA expression assay, we detected differential regulation of distinct ANK3 transcription start sites (TSSs) and coupling of specific 5â ends with 3â mRNA splicing events in post-mortem human brain and human stem cell-derived neural progenitors and neurons. Furthermore, allelic variation at the BDâassociated SNP rs1938526 correlated with a significant difference in cerebellar expression of a brain-specific ANK3 transcript. These findings suggest a brain-specific cis-regulatory transcriptional effect of ANK3 may be relevant to BD pathophysiology
Disrupted in Schizophrenia 1 Regulates Neuronal Progenitor Proliferation via Modulation of GSK3β/β-Catenin Signaling
The Disrupted in Schizophrenia 1 (DISC1) gene is disrupted by a balanced chromosomal translocation (1; 11) (q42; q14.3) in a Scottish family with a high incidence of major depression, schizophrenia, and bipolar disorder. Subsequent studies provided indications that DISC1 plays a role in brain development. Here, we demonstrate that suppression of DISC1 expression reduces neural progenitor proliferation, leading to premature cell cycle exit and differentiation. Several lines of evidence suggest that DISC1 mediates this function by regulating GSK3β. First, DISC1 inhibits GSK3β activity through direct physical interaction, which reduces β-catenin phosphorylation and stabilizes β-catenin. Importantly, expression of stabilized β-catenin overrides the impairment of progenitor proliferation caused by DISC1 loss of function. Furthermore, GSK3 inhibitors normalize progenitor proliferation and behavioral defects caused by DISC1 loss of function. Together, these results implicate DISC1 in GSK3β/β-catenin signaling pathways and provide a framework for understanding how alterations in this pathway may contribute to the etiology of psychiatric disorders.National Alliance for Research on Schizophrenia and Depression (U.S.) (Young Investigator Award)Natural Sciences and Engineering Research Council of Canada (Postdoctoral Award)Human Frontier Science Program (Strasbourg, France) (Fellowship)Singleton FellowshipNational Institutes of Health (U.S.) (Grant NS37007
AAK1 Identified as an Inhibitor of Neuregulin-1/ErbB4-Dependent Neurotrophic Factor Signaling Using Integrative Chemical Genomics and Proteomics
SummaryTarget identification remains challenging for the field of chemical biology. We describe an integrative chemical genomic and proteomic approach combining the use of differentially active analogs of small molecule probes with stable isotope labeling by amino acids in cell culture-mediated affinity enrichment, followed by subsequent testing of candidate targets using RNA interference-mediated gene silencing. We applied this approach to characterizing the natural product K252a and its ability to potentiate neuregulin-1 (Nrg1)/ErbB4 (v-erb-a erythroblastic leukemia viral oncogene homolog 4)-dependent neurotrophic factor signaling and neuritogenesis. We show that AAK1 (adaptor-associated kinase 1) is a relevant target of K252a, and that the loss of AAK1 alters ErbB4 trafficking and expression levels, providing evidence for a previously unrecognized role for AAK1 in Nrg1-mediated neurotrophic factor signaling. Similar strategies should lead to the discovery of novel targets for therapeutic development
Epigenetic Characterization of the FMR1 Gene and Aberrant Neurodevelopment in Human Induced Pluripotent Stem Cell Models of Fragile X Syndrome
Fragile X syndrome (FXS) is the most common inherited cause of intellectual disability. In addition to cognitive deficits, FXS patients exhibit hyperactivity, attention deficits, social difficulties, anxiety, and other autistic-like behaviors. FXS is caused by an expanded CGG trinucleotide repeat in the 5Ⲡuntranslated region of the Fragile X Mental Retardation (FMR1) gene leading to epigenetic silencing and loss of expression of the Fragile X Mental Retardation protein (FMRP). Despite the known relationship between FMR1 CGG repeat expansion and FMR1 silencing, the epigenetic modifications observed at the FMR1 locus, and the consequences of the loss of FMRP on human neurodevelopment and neuronal function remain poorly understood. To address these limitations, we report on the generation of induced pluripotent stem cell (iPSC) lines from multiple patients with FXS and the characterization of their differentiation into post-mitotic neurons and glia. We show that clones from reprogrammed FXS patient fibroblast lines exhibit variation with respect to the predominant CGG-repeat length in the FMR1 gene. In two cases, iPSC clones contained predominant CGG-repeat lengths shorter than measured in corresponding input population of fibroblasts. In another instance, reprogramming a mosaic patient having both normal and pre-mutation length CGG repeats resulted in genetically matched iPSC clonal lines differing in FMR1 promoter CpG methylation and FMRP expression. Using this panel of patient-specific, FXS iPSC models, we demonstrate aberrant neuronal differentiation from FXS iPSCs that is directly correlated with epigenetic modification of the FMR1 gene and a loss of FMRP expression. Overall, these findings provide evidence for a key role for FMRP early in human neurodevelopment prior to synaptogenesis and have implications for modeling of FXS using iPSC technology. By revealing disease-associated cellular phenotypes in human neurons, these iPSC models will aid in the discovery of novel therapeutics for FXS and other autism-spectrum disorders sharing common pathophysiology.FRAXA Research FoundationHarvard Stem Cell Institute (seed grant)Stanley Medical Research InstituteNational Institute of Mental Health (U.S.) (grant #R33MH087896
The 22q11.2 region regulates presynaptic gene-products linked to schizophrenia
How the 22q11.2 deletion predisposes to psychiatric disease is unclear. Here, the authors examine living human neuronal cells and show that 22q11.2 regulates the expression of genes linked to autism during early development, and genes linked to schizophrenia and synaptic biology in neurons. It is unclear how the 22q11.2 deletion predisposes to psychiatric disease. To study this, we generated induced pluripotent stem cells from deletion carriers and controls and utilized CRISPR/Cas9 to introduce the heterozygous deletion into a control cell line. Here, we show that upon differentiation into neural progenitor cells, the deletion acted in trans to alter the abundance of transcripts associated with risk for neurodevelopmental disorders including autism. In excitatory neurons, altered transcripts encoded presynaptic factors and were associated with genetic risk for schizophrenia, including common and rare variants. To understand how the deletion contributed to these changes, we defined the minimal protein-protein interaction network that best explains gene expression alterations. We found that many genes in 22q11.2 interact in presynaptic, proteasome, and JUN/FOS transcriptional pathways. Our findings suggest that the 22q11.2 deletion impacts genes that may converge with psychiatric risk loci to influence disease manifestation in each deletion carrier.Peer reviewe
Pheromone-sensing neurons regulate peripheral lipid metabolism in <i>Caenorhabditis elegans</i>
It is now established that the central nervous system plays an important role in regulating whole body metabolism and energy balance. However, the extent to which sensory systems relay environmental information to modulate metabolic events in peripheral tissues has remained poorly understood. In addition, it has been challenging to map the molecular mechanisms underlying discrete sensory modalities with respect to their role in lipid metabolism. In previous work our lab has identified instructive roles for serotonin signaling as a surrogate for food availability, as well as oxygen sensing, in the control of whole body metabolism. In this study, we now identify a role for a pair of pheromone-sensing neurons in regulating fat metabolism in C. elegans, which has emerged as a tractable and highly informative model to study the neurobiology of metabolism. A genetic screen revealed that GPA-3, a member of the GÎą family of G proteins, regulates body fat content in the intestine, the major metabolic organ for C. elegans. Genetic and reconstitution studies revealed that the potent body fat phenotype of gpa-3 null mutants is controlled from a pair of neurons called ADL(L/R). We show that cAMP functions as the second messenger in the ADL neurons, and regulates body fat stores via the neurotransmitter acetylcholine, from downstream neurons. We find that the pheromone ascr#3, which is detected by the ADL neurons, regulates body fat stores in a GPA-3-dependent manner. We define here a third sensory modality, pheromone sensing, as a major regulator of body fat metabolism. The pheromone ascr#3 is an indicator of population density, thus we hypothesize that pheromone sensing provides a salient 'denominator' to evaluate the amount of food available within a population and to accordingly adjust metabolic rate and body fat levels
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