The mammalian forebrain is a tissue of stunning complexity comprised of numerous regions each containing many distinct cell types that differ in their intrinsic and synaptic physiology, morphology and connectivity. These differences are likely conferred by differential gene expression, but the extent and nature of cell type specific gene expression is largely unknown. Here, we carried out microarray analysis of twelve major classes of fluorescently labelled neurons within the forebrain and provide the first comprehensive view of gene expression differences. The results demonstrate a profound molecular heterogeneity among neuronal subtypes, represented disproportionately by gene paralogs, and begin to reveal the genetic programs underlying the fundamental divisions between neuronal classes including that between glutamatergic and GABAergic neurons.
Molecular taxonomy of major neuronal classes in the adult mouse forebrain.
Sex, Specimen part
View SamplesMutations in methyl-CpG-binding protein 2 (MeCP2) cause Rett syndrome and related autism spectrum disorders. MeCP2 is believed to be required for proper regulation of brain gene expression, but prior microarray studies in Mecp2 knockout mice using brain tissue homogenates have revealed only subtle changes in gene expression. Here, by profiling discrete subtypes of neurons we uncovered more dramatic effects of MeCP2 on gene expression, overcoming the "dilution problem" associated with assaying homogenates of complex tissues. The results reveal misregulation of genes involved in neuronal connectivity and communication. Importantly, genes up-regulated following loss of MeCP2 are biased toward longer genes but this is not true for down-regulated genes, suggesting MeCP2 may selectively repress long genes. Since genes involved in neuronal connectivity and communication, such as cell adhesion and cell-cell signaling genes, are enriched among longer genes, their misregulation following loss of MeCP2 suggests a possible etiology for altered circuit function in Rett syndrome.
Cell-type-specific repression by methyl-CpG-binding protein 2 is biased toward long genes.
Sex, Age, Specimen part
View SamplesThis SuperSeries is composed of the SubSeries listed below.
A role for microRNAs in the Drosophila circadian clock.
Specimen part, Time
View SamplesLittle is known about the contribution of translational control to circadian rhythms. To address this issue and in particular translational control by microRNAs (miRNAs), we knocked down the miRNA biogenesis pathway in Drosophila circadian tissues. In combination with an increase in circadian-mediated transcription, this severely affected Drosophila behavioral rhythms, indicating that miRNAs function in circadian timekeeping. To identify miRNAmRNA pairs important for this regulation, immunoprecipitation of AGO1 followed by microarray analysis identified mRNAs under miRNA-mediated control. They included three core clock mRNAsclock (clk), vrille (vri), and clockworkorange (cwo). To identify miRNAs involved in circadian timekeeping, we exploited circadian cell-specific inhibition of the miRNA biogenesis pathway followed by tiling array analysis. This approach identified miRNAs expressed in fly head circadian tissue. Behavioral and molecular experiments show that one of these miRNAs, the developmental regulator bantam, has a role in the core circadian pacemaker. S2 cell biochemical experiments indicate that bantam regulates the translation of clk through an association with three target sites located within the clk 39 untranslated region (UTR). Moreover, clk transgenes harboring mutated bantam sites in their 39 UTRs rescue rhythms of clk mutant flies much less well than wild-type CLK transgenes.
No associated publication
Specimen part, Time
View SamplesGene expression profiling of distinct members of a neuronal circuit has the potential to identify candidate molecules and mechanisms that underlie the formation, organization and function of the circuit. To this end, we report here the application of a novel method to characterize RNAs from small numbers of specific Drosophila brain neurons, which belong to the circadian circuit. We identified three different sets of mRNAs enriched in different subclasses of clock neurons: one is enriched in all clock neurons, a second is enriched in PDF-positive clock neurons and a third is enriched in PDF-negative clock neurons. Moreover, we characterized 2 novel genes, Fer2 and dnocturnin, one from each subgroup, which highlight subgroup-specific features and play important roles in circadian rhythms. The methodology is a powerful tool not only to dissect the cellular and molecular basis of circadian rhythms but also to molecularly characterize other Drosophila neuronal circuits.
No associated publication
Specimen part
View SamplesFast-spiking (FS) interneurons are important elements of neocortical circuitry that constitute the primary source of synaptic inhibition in adult cortex and impart temporal organization on ongoing cortical activity. The highly specialized intrinsic membrane and firing properties that allow cortical FS interneurons to perform these functions are attributable to equally specialized gene expression, which is ultimately coordinated by cell-type-specific transcriptional regulation. Although embryonic transcriptional events govern the initial steps of cell-type specification in most cortical interneurons, including FS cells, the electrophysiological properties that distinguish adult cortical cell types emerge relatively late in postnatal development, and the transcriptional events that drive this maturational process are not known. To address this, we used mouse whole-genome microarrays and whole-cell patch clamp to characterize the transcriptional and electrophysiological maturation of cortical FS interneurons between postnatal day 7 (P7) and P40. We found that the intrinsic and synaptic physiology of FS cells undergoes profound regulation over the first 4 postnatal weeks and that these changes are correlated with primarily monotonic but bidirectional transcriptional regulation of thousands of genes belonging to multiple functional classes. Using our microarray screen as a guide, we discovered that upregulation of two-pore K leak channels between P10 and P25 contributes to one of the major differences between the intrinsic membrane properties of immature and adult FS cells and found a number of other candidate genes that likely confer cell-type specificity on mature FS cells.
Transcriptional and electrophysiological maturation of neocortical fast-spiking GABAergic interneurons.
Specimen part
View SamplesThe transcriptional events accompanying synaptogenesis are largely unknown, or have been studied in systems in which synapse formation occurs gradually over time. With a system in which synaptogenesis is synchronized and controllable, molecular or biochemical techniques can be used to examine cellular events across cultures on a wide scale, as synapses develop.
Synaptogenesis in purified cortical subplate neurons.
No sample metadata fields
View SamplesIn order to identify genes regulated by VE-cadherin expression, we compared a mouse VE-cadherin null cell line (VEC null) with the same line reconstituted with VE-cadherin wild type cDNA (VEC positive). The morphological and functional properties of these cell lines were described previously [Lampugnani,M.G. et al. Contact inhibition of VEGF-induced proliferation requires vascular endothelial cadherin, beta-catenin, and the phosphatase DEP-1/CD148. J. Cell Biol. 161, 793-804 (2003)]. By Affymetrix gene expression analysis we found several genes up-regulated by VE-cadherin, among which claudin-5 reached remarkably high levels. The up-regulation of these genes required not only VE-cadherin expression but also cell confluence suggesting that VE-cadherin clustering at junctions was needed.
Endothelial adherens junctions control tight junctions by VE-cadherin-mediated upregulation of claudin-5.
No sample metadata fields
View SamplesEndothelial cells (ECs) express two members of the cadherin family, VE- and N-cadherin. While VE-cadherin induces EC homotypic adhesion, N-cadherin function in ECs remains largely unknown. EC-specific inactivation of either VE- or N-cadherin leads to early foetal lethality suggesting that these cadherins play a non-redundant role in vascular development.
Overlapping and divergent signaling pathways of N-cadherin and VE-cadherin in endothelial cells.
Specimen part, Cell line
View SamplesLipo-chitooligosaccharides (LCOs) produced by N2-fixing rhizobacteria initiate host nodule formation. Foliar application of LCOs has been shown to induce stress-related genes under optimal growth conditions. To study the effects of LCO foliar spray under stressed conditions, soybean seedlings grown at optimal temperature were exposed to sub-optimal temperature. After a 5-day acclimation period, the first trifoliolate leaves were sprayed with 10-7 M LCO (NodBj-V (C18:1, MeFuc)) produced by Bradyrhizobium japonicum, and harvested at 0 and 48 h following treatment. Microarray analysis was performed using Affymetrix GeneChip Soybean Genome Arrays. A total of 147 genes were differentially expressed 48 h after LCO treatment, including a number of stress-related genes and transcription factors. In addition, during the 48 h following treatment, hundreds of genes were differentially expressed in LCO-treated plants, indicating that the dynamic soybean foliar transcriptome was highly responsive to LCO treatment. The microarray data was supported by quantitative real-time PCR data.
No associated publication
Specimen part, Treatment, Time
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