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Neuro-2a

RRID:CVCL_0470

Organism

Mus musculus

Comments

Characteristics: Capable of differentiating into various types of neurons. Doubling time: ~70 hours (DSMZ). Omics: Deep proteome analysis. Omics: SNP array analysis. Breed/subspecies: A/J. DT Created: 04-04-12; Last updated: 06-09-19; Version: 22

Proper Citation

RCB Cat# RCB2639, RRID:CVCL_0470

Category

Cancer cell line DT Created: 04-04-12; Last updated: 06-09-19; Version: 22

Sex

DT Created: 04-04-12; Last updated: 06-09-19; Version: 22

Synonyms

NEURO-2A, Neuro 2a, Neuro2a, Neuro2A, N-2a, N2a, N2A, Nb2a, NB2a DT Created: 04-04-12, Last updated: 06-09-19, Version: 22

Vendor

RCB

Cat Num

RCB2639

Cross References

BTO; BTO:0001976 CLO; CLO_0008154 CLO; CLO_0050180 MCCL; MCC:0000368 CLDB; cl3682 CLDB; cl3683 CLDB; cl3684 CLDB; cl3685 CLDB; cl4956 CLDB; cl5236 ATCC; CCL-131 BCRC; 60026 BCRJ; 0189 CCLV; CCLV-RIE 0132 CCRID; 3111C0001CCC000291 CCRID; 3131C0001000800029 CCRID; 3142C0001000001719 CCTCC; GDC0162 ChEMBL-Cells; CHEMBL3307521 ChEMBL-Targets; CHEMBL614062 CLS; 400394/p451_Neuro-2A DSMZ; ACC-148 ECACC; 89121404 ICLC; ATL99007 IZSLER; BS TCL 128 JCRB; IFO50081 KCB; KCB 2014025YJ Lonza; 451 PRIDE; PXD000065 PRIDE; PXD002290 RCB; RCB2639 TKG; TKG 0509 TOKU-E; 2746 TOKU-E; 3552 TOKU-E; 3563 Wikidata; Q18391478 DT Created: 04-04-12; Last updated: 06-09-19; Version: 22

Mitoregulin: A lncRNA-Encoded Microprotein that Supports Mitochondrial Supercomplexes and Respiratory Efficiency.

  • Stein CS
  • Cell Rep
  • 2018 Jun 26

Literature context:


Abstract:

Mitochondria are composed of many small proteins that control protein synthesis, complex assembly, metabolism, and ion and reactive oxygen species (ROS) handling. We show that a skeletal muscle- and heart-enriched long non-coding RNA, LINC00116, encodes a highly conserved 56-amino-acid microprotein that we named mitoregulin (Mtln). Mtln localizes to the inner mitochondrial membrane, where it binds cardiolipin and influences protein complex assembly. In cultured cells, Mtln overexpression increases mitochondrial membrane potential, respiration rates, and Ca2+ retention capacity while decreasing mitochondrial ROS and matrix-free Ca2+. Mtln-knockout mice display perturbations in mitochondrial respiratory (super)complex formation and activity, fatty acid oxidation, tricarboxylic acid (TCA) cycle enzymes, and Ca2+ retention capacity. Blue-native gel electrophoresis revealed that Mtln co-migrates alongside several complexes, including the complex I assembly module, complex V, and supercomplexes. Under denaturing conditions, Mtln remains in high-molecular-weight complexes, supporting its role as a sticky molecular tether that enhances respiratory efficiency by bolstering protein complex assembly and/or stability.

Funding information:
  • NIGMS NIH HHS - GM034869(United States)

Phospholipase PLA2G6, a Parkinsonism-Associated Gene, Affects Vps26 and Vps35, Retromer Function, and Ceramide Levels, Similar to α-Synuclein Gain.

  • Lin G
  • Cell Metab.
  • 2018 Jun 7

Literature context:


Abstract:

Mutations in PLA2G6 (PARK14) cause neurodegenerative disorders in humans, including autosomal recessive neuroaxonal dystrophy and early-onset parkinsonism. We show that loss of iPLA2-VIA, the fly homolog of PLA2G6, reduces lifespan, impairs synaptic transmission, and causes neurodegeneration. Phospholipases typically hydrolyze glycerol phospholipids, but loss of iPLA2-VIA does not affect the phospholipid composition of brain tissue but rather causes an elevation in ceramides. Reducing ceramides with drugs, including myriocin or desipramine, alleviates lysosomal stress and suppresses neurodegeneration. iPLA2-VIA binds the retromer subunits Vps35 and Vps26 and enhances retromer function to promote protein and lipid recycling. Loss of iPLA2-VIA impairs retromer function, leading to a progressive increase in ceramide. This induces a positive feedback loop that affects membrane fluidity and impairs retromer function and neuronal function. Similar defects are observed upon loss of vps26 or vps35 or overexpression of α-synuclein, indicating that these defects may be common in Parkinson disease.

Funding information:
  • NCI NIH HHS - R01 CA130893(United States)

Defects in the Alternative Splicing-Dependent Regulation of REST Cause Deafness.

  • Nakano Y
  • Cell
  • 2018 Jun 25

Literature context:


Abstract:

The DNA-binding protein REST forms complexes with histone deacetylases (HDACs) to repress neuronal genes in non-neuronal cells. In differentiating neurons, REST is downregulated predominantly by transcriptional silencing. Here we report that post-transcriptional inactivation of REST by alternative splicing is required for hearing in humans and mice. We show that, in the mechanosensory hair cells of the mouse ear, regulated alternative splicing of a frameshift-causing exon into the Rest mRNA is essential for the derepression of many neuronal genes. Heterozygous deletion of this alternative exon of mouse Rest causes hair cell degeneration and deafness, and the HDAC inhibitor SAHA (Vorinostat) rescues the hearing of these mice. In humans, inhibition of the frameshifting splicing event by a novel REST variant is associated with dominantly inherited deafness. Our data reveal the necessity for alternative splicing-dependent regulation of REST in hair cells, and they identify a potential treatment for a group of hereditary deafness cases.

Funding information:
  • NIMH NIH HHS - 5 F32 MH064339-03(United States)

Retrograde transport of γ-secretase from endosomes to the trans-Golgi network regulates Aβ42 production.

  • Kanatsu K
  • J. Neurochem.
  • 2018 May 31

Literature context:


Abstract:

The aberrant metabolism of amyloid-β protein (Aβ) in the human brain has been implicated in the etiology of Alzheimer disease (AD). γ-Secretase is the enzyme that generates various forms of Aβ, such as Aβ40 and Aβ42, the latter being an aggregation-prone toxic peptide that is involved in the pathogenesis of AD. Recently, we found that clathrin-mediated endocytosis of γ-secretase affects the production and deposition of Aβ42 in vivo, suggesting that the membrane trafficking of γ-secretase affects its enzymatic activity. However, the detailed intracellular trafficking pathway of γ-secretase and its contribution to Aβ42 generation remain unclear. Here we show that Retro-2, which inhibits the retrograde transport, elevated the Aβ42-generating activity both in cultured cells and mice brain. However, the result of in vitro γ-secretase assay using a recombinant substrate suggested that Retro-2 did not elevate the intrinsic Aβ42-production activity of γ-secretase. Immunocytochemistry and cell-surface biotinylation experiments revealed that γ-secretase is recycled via the endosome-to-trans-Golgi network (TGN) transport. In addition, γ-secretase is retrogradely transported by syntaxin 5/6, known as targets of Retro-2, independent pathway. Conversely, TPT-260, which enhances the trafficking function of retromers, lowered Aβ42 levels and the Aβ42/(Aβ40 + Aβ42) ratio in secreted Aβ from cultured cells. Our results strongly suggest that the endosome-to-TGN trafficking of γ-secretase regulates its Aβ42 production activity. Modulation of this trafficking pathway might be a potential target for the development of Aβ42-lowering AD therapeutics. This article is protected by copyright. All rights reserved.

Funding information:
  • NCI NIH HHS - P30 CA08748(United States)

Palladin Is a Neuron-Specific Translational Target of mTOR Signaling That Regulates Axon Morphogenesis.

  • Umegaki Y
  • J. Neurosci.
  • 2018 May 23

Literature context:


Abstract:

The mTOR signaling pathway regulates protein synthesis and diverse aspects of neuronal morphology that are important for brain development and function. To identify proteins controlled translationally by mTOR signaling, we performed ribosome profiling analyses in mouse cortical neurons and embryonic stem cells upon acute mTOR inhibition. Among proteins whose translation was significantly affected by mTOR inhibition selectively in neurons, we identified the cytoskeletal regulator protein palladin, which is localized within the cell body and axons in hippocampal neurons. Knockdown of palladin eliminated supernumerary axons induced by suppression of the tuberous sclerosis complex protein TSC1 in neurons, demonstrating that palladin regulates neuronal morphogenesis downstream of mTOR signaling. Our findings provide novel insights into an mTOR-dependent mechanism that controls neuronal morphogenesis through translational regulation.SIGNIFICANCE STATEMENT This study reports the discovery of neuron-specific protein translational responses to alterations of mTOR activity. By using ribosome profiling analysis, which can reveal the location and quantity of translating ribosomes on mRNAs, multiple aspects of protein translation were quantitatively analyzed in mouse embryonic stem cells and cortical neurons upon acute mTOR inhibition. Neurons displayed distinct patterns of ribosome occupancy for each codon and ribosome stalling during translation at specific positions of mRNAs. Importantly, the cytoskeletal regulator palladin was identified as a translational target protein of mTOR signaling in neurons. Palladin operates downstream of mTOR to modulate axon morphogenesis. This study identifies a novel mechanism of neuronal morphogenesis regulated by mTOR signaling through control of translation of the key protein palladin.

Funding information:
  • Medical Research Council - (United Kingdom)
  • NINDS NIH HHS - R01 NS051255()

Coordinated Control of mRNA and rRNA Processing Controls Embryonic Stem Cell Pluripotency and Differentiation.

  • Corsini NS
  • Cell Stem Cell
  • 2018 Apr 5

Literature context:


Abstract:

Stem cell-specific transcriptional networks are well known to control pluripotency, but constitutive cellular processes such as mRNA splicing and protein synthesis can add complex layers of regulation with poorly understood effects on cell-fate decisions. Here, we show that the RNA binding protein HTATSF1 controls embryonic stem cell differentiation by regulating multiple aspects of RNA processing during ribosome biogenesis. HTATSF1, in a complex with splicing factor SF3B1, controls intron removal from ribosomal protein transcripts and regulates ribosomal RNA transcription and processing, thereby controlling 60S ribosomal abundance and protein synthesis. HTATSF1-dependent protein synthesis is essential for naive pre-implantation epiblast to transition into post-implantation epiblast, a stage with transiently low protein synthesis, and further differentiation toward neuroectoderm. Together, these results identify coordinated regulation of ribosomal RNA and protein synthesis by HTATSF1 and show that this essential mechanism controls protein synthesis during early mammalian embryogenesis.

Funding information:
  • NIGMS NIH HHS - GM077403(United States)

An Image-Based miRNA Screen Identifies miRNA-135s As Regulators of CNS Axon Growth and Regeneration by Targeting Krüppel-like Factor 4.

  • van Battum EY
  • J. Neurosci.
  • 2018 Jan 17

Literature context:


Abstract:

During embryonic development, axons extend over long distances to establish functional connections. In contrast, axon regeneration in the adult mammalian CNS is limited in part by a reduced intrinsic capacity for axon growth. Therefore, insight into the intrinsic control of axon growth may provide new avenues for enhancing CNS regeneration. Here, we performed one of the first miRNome-wide functional miRNA screens to identify miRNAs with robust effects on axon growth. High-content screening identified miR-135a and miR-135b as potent stimulators of axon growth and cortical neuron migration in vitro and in vivo in male and female mice. Intriguingly, both of these developmental effects of miR-135s relied in part on silencing of Krüppel-like factor 4 (KLF4), a well known intrinsic inhibitor of axon growth and regeneration. These results prompted us to test the effect of miR-135s on axon regeneration after injury. Our results show that intravitreal application of miR-135s facilitates retinal ganglion cell (RGC) axon regeneration after optic nerve injury in adult mice in part by repressing KLF4. In contrast, depletion of miR-135s further reduced RGC axon regeneration. Together, these data identify a novel neuronal role for miR-135s and the miR-135-KLF4 pathway and highlight the potential of miRNAs as tools for enhancing CNS axon regeneration.SIGNIFICANCE STATEMENT Axon regeneration in the adult mammalian CNS is limited in part by a reduced intrinsic capacity for axon growth. Therefore, insight into the intrinsic control of axon growth may provide new avenues for enhancing regeneration. By performing an miRNome-wide functional screen, our studies identify miR-135s as stimulators of axon growth and neuron migration and show that intravitreal application of these miRNAs facilitates CNS axon regeneration after nerve injury in adult mice. Intriguingly, these developmental and regeneration-promoting effects rely in part on silencing of Krüppel-like factor 4 (KLF4), a well known intrinsic inhibitor of axon regeneration. Our data identify a novel neuronal role for the miR-135-KLF4 pathway and support the idea that miRNAs can be used for enhancing CNS axon regeneration.

Funding information:
  • NCI NIH HHS - R01 CA112054(United States)

Neuronal activity-dependent local activation of dendritic unfolded protein response promotes expression of brain-derived neurotrophic factor in cell soma.

  • Saito A
  • J. Neurochem.
  • 2018 Jan 2

Literature context:


Abstract:

Unfolded protein response (UPR) has roles not only in resolving the accumulation of unfolded proteins owing to endoplasmic reticulum (ER) stress, but also in regulation of cellular physiological functions. ER stress transducers providing the branches of UPR signaling are known to localize in distal dendritic ER of neurons. These reports suggest that local activation of UPR branches may produce integrated outputs for distant communication, and allow regulation of local events in highly polarized neurons. Here, we demonstrated that synaptic activity- and brain-derived neurotrophic factor (BDNF)-dependent local activation of UPR signaling could be associated with dendritic functions through retrograde signal propagation by using murine neuroblastoma cell line, Neuro-2A and primary cultured hippocampal neurons derived from postnatal day 0 litter C57BL/6 mice. ER stress transducer, inositol-requiring kinase 1 (IRE1), was activated at postsynapses in response to excitatory synaptic activation. Activated dendritic IRE1 accelerated accumulation of the downstream transcription factor, x-box-binding protein 1 (XBP1), in the nucleus. Interestingly, excitatory synaptic activation-dependent up-regulation of XBP1 directly facilitated transcriptional activation of BDNF. BDNF in turn drove its own expression via IRE1-XBP1 pathway in a protein kinase A-dependent manner. Exogenous treatment with BDNF promoted extension and branching of dendrites through the protein kinase A-IRE1-XBP1 cascade. Taken together, our findings indicate novel mechanisms for communication between soma and distal sites of polarized neurons that are coordinated by local activation of IRE1-XBP1 signaling. Synaptic activity- and BDNF-dependent distinct activation of dendritic IRE1-XBP1 cascade drives BDNF expression in cell soma and may be involved in dendritic extension. Cover Image for this issue: doi. 10.1111/jnc.14159.

Role of the GM1 ganglioside oligosaccharide portion in the TrkA-dependent neurite sprouting in neuroblastoma cells.

  • Chiricozzi E
  • J. Neurochem.
  • 2017 Dec 20

Literature context:


Abstract:

GM1 ganglioside (II3 NeuAc-Gg4 Cer) is known to promote neurite formation in neuroblastoma cells by activating TrkA-MAPK pathway. The molecular mechanism by which GM1 is involved in the neurodifferentiation process is still unknown, however, in vitro and in vivo evidences have suggested that the oligosaccharide portion of this ganglioside could be involved. Here, we report that, similarly to the entire GM1 molecule, its oligosaccharide II3 NeuAc-Gg4, rather than its ceramide (Cer) portion is responsible for the neurodifferentiation process by augmenting neurite elongation and increasing the neurofilament protein expression in murine neuroblastoma cells, Neuro2a. Conversely, asialo-GM1, GM2 and GM3 oligosaccharides are not effective in neurite elongation on Neuro2a cells, whereas the effect exerted by the Fuc-GM1 oligosaccharide (IV2 αFucII3 Neu5Ac-Gg4 ) is similar to that exerted by GM1 oligosaccharide. The neurotrophic properties of GM1 oligosaccharide are exerted by activating the TrkA receptor and the following phosphorylation cascade. By photolabeling experiments performed with a nitrophenylazide containing GM1 oligosaccharide, labeled with tritium, we showed a direct interaction between the GM1 oligosaccharide and the extracellular domain of TrkA receptor. Moreover, molecular docking analyses confirmed that GM1 oligosaccharide binds the TrkA-nerve growth factor complex leading to a binding free energy of approx. -11.5 kcal/mol, acting as a bridge able to increase and stabilize the TrkA-nerve growth factor molecular interactions.

Intracerebroventricular administration of Cystatin C ameliorates disease in SOD1-linked amyotrophic lateral sclerosis mice.

  • Watanabe S
  • J. Neurochem.
  • 2017 Dec 29

Literature context:


Abstract:

Cystatin C (CysC) is a major protein component of Bunina bodies, which are a pathological hallmark observed in the remaining motor neurons of patients with amyotrophic lateral sclerosis (ALS). Dominant mutations in the SOD1 gene, encoding Cu/Zn superoxide dismutase (SOD1), are causative for a subset of inherited ALS cases. Our previous study showed that CysC exerts a neuroprotective effect against mutant SOD1-mediated toxicity in vitro; however, in vivo evidence of the beneficial effects mediated by CysC remains obscure. Here we examined the therapeutic potential of recombinant human CysC in vivo using a mouse model of ALS in which the ALS-linked mutated SOD1 gene is expressed (SOD1G93A mice). Intracerebroventricular administration of CysC during the early symptomatic SOD1G93A mice extended their survival times. Administered CysC was predominantly distributed in ventral horn neurons including motor neurons, and induced autophagy through AMP-activated kinase activation to reduce the amount of insoluble mutant SOD1 species. Moreover, PGC-1α, a disease modifier of ALS, was restored by CysC through AMP-activated kinase activation. Finally, the administration of CysC also promoted aggregation of CysC in motor neurons, which is similar to Bunina bodies. Taken together, our findings suggest that CysC represents a promising therapeutic candidate for ALS.

Funding information:
  • NIAMS NIH HHS - R01-AR050478(United States)

Inducible and reversible phenotypes in a novel mouse model of Friedreich's Ataxia.

  • Chandran V
  • Elife
  • 2017 Dec 19

Literature context:


Abstract:

Friedreich's ataxia (FRDA), the most common inherited ataxia, is caused by recessive mutations that reduce the levels of frataxin (FXN), a mitochondrial iron binding protein. We developed an inducible mouse model of Fxn deficiency that enabled us to control the onset and progression of disease phenotypes by the modulation of Fxn levels. Systemic knockdown of Fxn in adult mice led to multiple phenotypes paralleling those observed in human patients across multiple organ systems. By reversing knockdown after clinical features appear, we were able to determine to what extent observed phenotypes represent reversible cellular dysfunction. Remarkably, upon restoration of near wild-type FXN levels, we observed significant recovery of function, associated pathology and transcriptomic dysregulation even after substantial motor dysfunction and pathology were observed. This model will be of broad utility in therapeutic development and in refining our understanding of the relative contribution of reversible cellular dysfunction at different stages in disease.

Funding information:
  • NCRR NIH HHS - S10 RR019391(United States)

Drp1 Mitochondrial Fission in D1 Neurons Mediates Behavioral and Cellular Plasticity during Early Cocaine Abstinence.

  • Chandra R
  • Neuron
  • 2017 Dec 20

Literature context:


Abstract:

Altered brain energy homeostasis is a key adaptation occurring in the cocaine-addicted brain, but the effect of cocaine on the fundamental source of energy, mitochondria, is unknown. We demonstrate an increase of dynamin-related protein-1 (Drp1), the mitochondrial fission mediator, in nucleus accumbens (NAc) after repeated cocaine exposure and in cocaine-dependent individuals. Mdivi-1, a demonstrated fission inhibitor, blunts cocaine seeking and locomotor sensitization, while blocking c-Fos induction and excitatory input onto dopamine receptor-1 (D1) containing NAc medium spiny neurons (MSNs). Drp1 and fission promoting Drp1 are increased in D1-MSNs, consistent with increased smaller mitochondria in D1-MSN dendrites after repeated cocaine. Knockdown of Drp1 in D1-MSNs blocks drug seeking after cocaine self-administration, while enhancing the fission promoting Drp1 enhances seeking after long-term abstinence from cocaine. We demonstrate a role for altered mitochondrial fission in the NAc, during early cocaine abstinence, suggesting potential therapeutic treatment of disrupting mitochondrial fission in cocaine addiction.

Funding information:
  • NCI NIH HHS - R01 CA140198(United States)
  • NIAAA NIH HHS - R01 AA024845()
  • NIDA NIH HHS - R01 DA037257()
  • NIDA NIH HHS - R01 DA038613()
  • NIGMS NIH HHS - R25 GM055036()
  • NIGMS NIH HHS - SC2 GM109811()

Cannabinoid Receptor Type 1 Agonist ACEA Protects Neurons from Death and Attenuates Endoplasmic Reticulum Stress-Related Apoptotic Pathway Signaling.

  • Vrechi TA
  • Neurotox Res
  • 2017 Nov 15

Literature context:


Abstract:

Neurodegeneration is the result of progressive destruction of neurons in the central nervous system, with unknown causes and pathological mechanisms not yet fully elucidated. Several factors contribute to neurodegenerative processes, including neuroinflammation, accumulation of neurotoxic factors, and misfolded proteins in the lumen of the endoplasmic reticulum (ER). Endocannabinoid signaling has been pointed out as an important modulatory system in several neurodegeneration-related processes, inhibiting the inflammatory response and increasing neuronal survival. Thus, we investigated the presumptive protective effect of the selective cannabinoid type 1 (CB1) receptor agonist arachidonyl-2'-chloroethylamide (ACEA) against inflammatory (lipopolysaccharide, LPS) and ER stress (tunicamycin) stimuli in an in vitro neuronal model (Neuro-2a neuroblastoma cells). Cell viability analysis revealed that ACEA was able to protect against cell death induced by LPS and tunicamycin. This neuroprotective effect occurs via the CB1 receptor in the inflammation process and via the transient receptor potential of vanilloid type-1 (TRPV1) channel in ER stress. Furthermore, the immunoblotting analyses indicated that the neuroprotective effect of ACEA seems to involve the modulation of eukaryotic initiation factor 2 (eIF2α), transcription factor C/EBP homologous protein (CHOP), and caspase 12, as well as the survival/death p44/42 MAPK, ERK1/2-related signaling pathways. Together, these data suggest that the endocannabinoid system is a potential therapeutic target in neurodegenerative processes, especially in ER-related neurodegenerative diseases.

Funding information:
  • Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - 1233360()
  • Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - 1279985()
  • Fundação de Amparo à Pesquisa do Estado de São Paulo - 2014/06372-0()

Mammalian FMRP S499 Is Phosphorylated by CK2 and Promotes Secondary Phosphorylation of FMRP.

  • Bartley CM
  • eNeuro
  • 2017 Oct 25

Literature context:


Abstract:

The fragile X mental retardation protein (FMRP) is an mRNA-binding regulator of protein translation that associates with 4-6% of brain transcripts and is central to neurodevelopment. Autism risk genes' transcripts are overrepresented among FMRP-binding mRNAs, and FMRP loss-of-function mutations are responsible for fragile X syndrome, the most common cause of monogenetic autism. It is thought that FMRP-dependent translational repression is governed by the phosphorylation of serine residue 499 (S499). However, recent evidence suggests that S499 phosphorylation is not modulated by metabotropic glutamate receptor class I (mGluR-I) or protein phosphatase 2A (PP2A), two molecules shown to regulate FMRP translational repression. Moreover, the mammalian FMRP S499 kinase remains unknown. We found that casein kinase II (CK2) phosphorylates murine FMRP S499. Further, we show that phosphorylation of FMRP S499 permits phosphorylation of additional, nearby residues. Evidence suggests that these nearby residues are modulated by mGluR-I and PP2A pathways. These data support an alternative phosphodynamic model of FMRP that is harmonious with prior studies and serves as a framework for further investigation.

cTag-PAPERCLIP Reveals Alternative Polyadenylation Promotes Cell-Type Specific Protein Diversity and Shifts Araf Isoforms with Microglia Activation.

  • Hwang HW
  • Neuron
  • 2017 Sep 13

Literature context:


Abstract:

Alternative polyadenylation (APA) is increasingly recognized to regulate gene expression across different cell types, but obtaining APA maps from individual cell types typically requires prior purification, a stressful procedure that can itself alter cellular states. Here, we describe a new platform, cTag-PAPERCLIP, that generates APA profiles from single cell populations in intact tissues; cTag-PAPERCLIP requires no tissue dissociation and preserves transcripts in native states. Applying cTag-PAPERCLIP to profile four major cell types in the mouse brain revealed common APA preferences between excitatory and inhibitory neurons distinct from astrocytes and microglia, regulated in part by neuron-specific RNA-binding proteins NOVA2 and PTBP2. We further identified a role of APA in switching Araf protein isoforms during microglia activation, impacting production of downstream inflammatory cytokines. Our results demonstrate the broad applicability of cTag-PAPERCLIP and a previously undiscovered role of APA in contributing to protein diversity between different cell types and cellular states within the brain.

Funding information:
  • NHGRI NIH HHS - UM1 HG008901()
  • NINDS NIH HHS - R01 NS034389()
  • NINDS NIH HHS - R01 NS081706()
  • NINDS NIH HHS - R35 NS097404()
  • NINDS NIH HHS - R56 NS034389()

Prosapip1-Dependent Synaptic Adaptations in the Nucleus Accumbens Drive Alcohol Intake, Seeking, and Reward.

  • Laguesse S
  • Neuron
  • 2017 Sep 27

Literature context:


Abstract:

The mammalian target of rapamycin complex 1 (mTORC1), a transducer of local dendritic translation, participates in learning and memory processes as well as in mechanisms underlying alcohol-drinking behaviors. Using an unbiased RNA-seq approach, we identified Prosapip1 as a novel downstream target of mTORC1 whose translation and consequent synaptic protein expression are increased in the nucleus accumbens (NAc) of mice excessively consuming alcohol. We demonstrate that alcohol-dependent increases in Prosapip1 levels promote the formation of actin filaments, leading to changes in dendritic spine morphology of NAc medium spiny neurons (MSNs). We further demonstrate that Prosapip1 is required for alcohol-dependent synaptic localization of GluA2 lacking AMPA receptors in NAc shell MSNs. Finally, we present data implicating Prosapip1 in mechanisms underlying alcohol self-administration and reward. Together, these data suggest that Prosapip1 in the NAc is a molecular transducer of structural and synaptic alterations that drive and/or maintain excessive alcohol use.

CpG and UpA dinucleotides in both coding and non-coding regions of echovirus 7 inhibit replication initiation post-entry.

  • Fros JJ
  • Elife
  • 2017 Sep 29

Literature context:


Abstract:

Most vertebrate and plant RNA and small DNA viruses suppress genomic CpG and UpA dinucleotide frequencies, apparently mimicking host mRNA composition. Artificially increasing CpG/UpA dinucleotides attenuates viruses through an entirely unknown mechanism. Using the echovirus 7 (E7) model in several cell types, we show that the restriction in E7 replication in mutants with increased CpG/UpA dinucleotides occurred immediately after viral entry, with incoming virions failing to form replication complexes. Sequences of CpG/UpA-high virus stocks showed no evidence of increased mutational errors that would render them replication defective, these viral RNAs were not differentially sequestered in cytoplasmic stress granules nor did they induce a systemic antiviral state. Importantly, restriction was not mediated through effects on translation efficiency since replicons with high CpG/UpA sequences inserted into a non-coding region were similarly replication defective. Host-cells thus possess intrinsic defence pathways that prevent replication of viruses with increased CpG/UpA frequencies independently of codon usage.

Body Temperature Cycles Control Rhythmic Alternative Splicing in Mammals.

  • Preußner M
  • Mol. Cell
  • 2017 Aug 3

Literature context:


Abstract:

The core body temperature of all mammals oscillates with the time of the day. However, direct molecular consequences of small, physiological changes in body temperature remain largely elusive. Here we show that body temperature cycles drive rhythmic SR protein phosphorylation to control an alternative splicing (AS) program. A temperature change of 1°C is sufficient to induce a concerted splicing switch in a large group of functionally related genes, rendering this splicing-based thermometer much more sensitive than previously described temperature-sensing mechanisms. AS of two exons in the 5' UTR of the TATA-box binding protein (Tbp) highlights the general impact of this mechanism, as it results in rhythmic TBP protein levels with implications for global gene expression in vivo. Together our data establish body temperature-driven AS as a core clock-independent oscillator in mammalian peripheral clocks.

Golgi-Resident Gαo Promotes Protrusive Membrane Dynamics.

  • Solis GP
  • Cell
  • 2017 Aug 24

Literature context:


Abstract:

To form protrusions like neurites, cells must coordinate their induction and growth. The first requires cytoskeletal rearrangements at the plasma membrane (PM), the second requires directed material delivery from cell's insides. We find that the Gαo-subunit of heterotrimeric G proteins localizes dually to PM and Golgi across phyla and cell types. The PM pool of Gαo induces, and the Golgi pool feeds, the growing protrusions by stimulated trafficking. Golgi-residing KDELR binds and activates monomeric Gαo, atypically for G protein-coupled receptors that normally act on heterotrimeric G proteins. Through multidimensional screenings identifying > 250 Gαo interactors, we pinpoint several basic cellular activities, including vesicular trafficking, as being regulated by Gαo. We further find small Golgi-residing GTPases Rab1 and Rab3 as direct effectors of Gαo. This KDELR → Gαo → Rab1/3 signaling axis is conserved from insects to mammals and controls material delivery from Golgi to PM in various cells and tissues.

A Requirement for Mena, an Actin Regulator, in Local mRNA Translation in Developing Neurons.

  • Vidaki M
  • Neuron
  • 2017 Aug 2

Literature context:


Abstract:

During neuronal development, local mRNA translation is required for axon guidance and synaptogenesis, and dysregulation of this process contributes to multiple neurodevelopmental and cognitive disorders. However, regulation of local protein synthesis in developing axons remains poorly understood. Here, we uncover a novel role for the actin-regulatory protein Mena in the formation of a ribonucleoprotein complex that involves the RNA-binding proteins HnrnpK and PCBP1 and regulates local translation of specific mRNAs in developing axons. We find that translation of dyrk1a, a Down syndrome- and autism spectrum disorders-related gene, is dependent on Mena, both in steady-state conditions and upon BDNF stimulation. We identify hundreds of additional mRNAs that associate with the Mena complex, suggesting that it plays broader role(s) in post-transcriptional gene regulation. Our work establishes a dual role for Mena in neurons, providing a potential link between regulation of actin dynamics and local translation.

Reduced Expression of Foxp1 as a Contributing Factor in Huntington's Disease.

  • Louis Sam Titus ASC
  • J. Neurosci.
  • 2017 Jul 5

Literature context:


Abstract:

Huntington's disease (HD) is an inherited neurodegenerative disease caused by a polyglutamine expansion in the huntington protein (htt). The neuropathological hallmark of HD is the loss of neurons in the striatum and, to a lesser extent, in the cortex. Foxp1 is a member of the Forkhead family of transcription factors expressed selectively in the striatum and the cortex. In the brain, three major Foxp1 isoforms are expressed: isoform-A (∼90 kDa), isoform-D (∼70 kDa), and isoform-C (∼50 kDa). We find that expression of Foxp1 isoform-A and -D is selectively reduced in the striatum and cortex of R6/2 HD mice as well as in the striatum of HD patients. Furthermore, expression of mutant htt in neurons results in the downregulation of Foxp1 Elevating expression of isoform-A or -D protects cortical neurons from death caused by the expression of mutant htt On the other hand, knockdown of Foxp1 promotes death in otherwise healthy neurons. Neuroprotection by Foxp1 is likely to be mediated by the transcriptional stimulation of the cell-cycle inhibitory protein p21Waf1/Cip1 Consistently, Foxp1 activates transcription of the p21Waf1/Cip1 gene promoter, and overexpression of Foxp1 in neurons results in the elevation of p21 expression. Moreover, knocking down of p21Waf1/Cip1 blocks the ability of Foxp1 to protect neurons from mut-Htt-induced neurotoxicity. We propose that the selective vulnerability of neurons of the striatum and cortex in HD is related to the loss of expression of Foxp1, a protein that is highly expressed in these neurons and required for their survival.SIGNIFICANCE STATEMENT Although the mutant huntingtin gene is expressed widely, neurons of the striatum and cortex are selectively affected in Huntington's disease (HD). Our results suggest that this selectivity is attributable to the reduced expression of Foxp1, a protein expressed selectively in striatal and cortical neurons that plays a neuroprotective role in these cells. We show that protection by Foxp1 involves stimulation of the p21Waf1/Cip1 (Cdkn1a) gene. Although three major Foxp1 isoforms (A, C, and D) are expressed in the brain, only isoform-A has been studied in the nervous system. We show that isoform-D is also expressed selectively, neuroprotective and downregulated in HD mice and patients. Our results suggest that Foxp1 might be an attractive therapeutic target for HD.

Funding information:
  • NINDS NIH HHS - R01 NS040408()

Reduced Slc6a15 in Nucleus Accumbens D2-Neurons Underlies Stress Susceptibility.

  • Chandra R
  • J. Neurosci.
  • 2017 Jul 5

Literature context:


Abstract:

Previous research demonstrates that Slc6a15, a neutral amino acid transporter, is associated with depression susceptibility. However, no study examined Slc6a15 in the ventral striatum [nucleus accumbens (NAc)] in depression. Given our previous characterization of Slc6a15 as a striatal dopamine receptor 2 (D2)-neuron-enriched gene, we examined the role of Slc6a15 in NAc D2-neurons in mediating susceptibility to stress in male mice. First, we showed that Slc6a15 mRNA was reduced in NAc of mice susceptible to chronic social defeat stress (CSDS), a paradigm that produces behavioral and molecular adaptations that resemble clinical depression. Consistent with our preclinical data, we observed Slc6a15 mRNA reduction in NAc of individuals with major depressive disorder (MDD). The Slc6a15 reduction in NAc occurred selectively in D2-neurons. Next, we used Cre-inducible viruses combined with D2-Cre mice to reduce or overexpress Slc6a15 in NAc D2-neurons. Slc6a15 reduction in D2-neurons caused enhanced susceptibility to a subthreshold social defeat stress (SSDS) as observed by reduced social interaction, while a reduction in social interaction following CSDS was not observed when Slc6a15 expression in D2-neurons was restored. Finally, since both D2-medium spiny neurons (MSNs) and D2-expressing choline acetyltransferase (ChAT) interneurons express Slc6a15, we examined Slc6a15 protein in these interneurons after CSDS. Slc6a15 protein was unaltered in ChAT interneurons. Consistent with this, reducing Slc5a15 selectively in NAc D2-MSNs, using A2A-Cre mice that express Cre selectively in D2-MSNs, caused enhanced susceptibility to SSDS. Collectively, our data demonstrate that reduced Slc6a15 in NAc occurs in MDD individuals and that Slc6a15 reduction in NAc D2-neurons underlies stress susceptibility.SIGNIFICANCE STATEMENT Our study demonstrates a role for reduced Slc6a15, a neutral amino acid transporter, in nucleus accumbens (NAc) in depression and stress susceptibility. The reduction of Slc6a15 occurs selectively in the NAc D2-neurons. Genetic reduction of Slc6a15 induces susceptibility to a subthreshold stress, while genetic overexpression in D2-neurons prevents social avoidance after chronic social defeat stress.

Funding information:
  • NIMH NIH HHS - R01 MH106500()

Physiological and pathophysiological control of synaptic GluN2B-NMDA receptors by the C-terminal domain of amyloid precursor protein.

  • Pousinha PA
  • Elife
  • 2017 Jul 6

Literature context:


Abstract:

The amyloid precursor protein (APP) harbors physiological roles at synapses and is central to Alzheimer's disease (AD) pathogenesis. Evidence suggests that APP intracellular domain (AICD) could regulate synapse function, but the underlying molecular mechanisms remain unknown. We addressed AICD actions at synapses, per se, combining in vivo AICD expression, ex vivo AICD delivery or APP knock-down by in utero electroporation of shRNAs with whole-cell electrophysiology. We report a critical physiological role of AICD in controlling GluN2B-containing NMDA receptors (NMDARs) at immature excitatory synapses, via a transcription-dependent mechanism. We further show that AICD increase in mature neurons, as reported in AD, alters synaptic NMDAR composition to an immature-like GluN2B-rich profile. This disrupts synaptic signal integration, via over-activation of SK channels, and synapse plasticity, phenotypes rescued by GluN2B antagonism. We provide a new physiological role for AICD, which becomes pathological upon AICD increase in mature neurons. Thus, AICD could contribute to AD synaptic failure.

Mov10 suppresses retroelements and regulates neuronal development and function in the developing brain.

  • Skariah G
  • BMC Biol.
  • 2017 Jun 29

Literature context:


Abstract:

BACKGROUND: Moloney leukemia virus 10 (Mov10) is an RNA helicase that mediates access of the RNA-induced silencing complex to messenger RNAs (mRNAs). Until now, its role as an RNA helicase and as a regulator of retrotransposons has been characterized exclusively in cell lines. We investigated the role of Mov10 in the mouse brain by examining its expression over development and attempting to create a Mov10 knockout mouse. Loss of both Mov10 copies led to early embryonic lethality. RESULTS: Mov10 was significantly elevated in postnatal murine brain, where it bound retroelement RNAs and mRNAs. Mov10 suppressed retroelements in the nucleus by directly inhibiting complementary DNA synthesis, while cytosolic Mov10 regulated cytoskeletal mRNAs to influence neurite outgrowth. We verified this important function by observing reduced dendritic arborization in hippocampal neurons from the Mov10 heterozygote mouse and shortened neurites in the Mov10 knockout Neuro2A cells. Knockdown of Fmrp also resulted in shortened neurites. Mov10, Fmrp, and Ago2 bound a common set of mRNAs in the brain. Reduced Mov10 in murine brain resulted in anxiety and increased activity in a novel environment, supporting its important role in the development of normal brain circuitry. CONCLUSIONS: Mov10 is essential for normal neuronal development and brain function. Mov10 preferentially binds RNAs involved in actin binding, neuronal projection, and cytoskeleton. This is a completely new and critically important function for Mov10 in neuronal development and establishes a precedent for Mov10 being an important candidate in neurological disorders that have underlying cytoarchitectural causes like autism and Alzheimer's disease.

Funding information:
  • NHLBI NIH HHS - R01 HL126845()

The N-terminus of the prion protein is a toxic effector regulated by the C-terminus.

  • Wu B
  • Elife
  • 2017 May 20

Literature context:


Abstract:

PrPC, the cellular isoform of the prion protein, serves to transduce the neurotoxic effects of PrPSc, the infectious isoform, but how this occurs is mysterious. Here, using a combination of electrophysiological, cellular, and biophysical techniques, we show that the flexible, N-terminal domain of PrPC functions as a powerful toxicity-transducing effector whose activity is tightly regulated in cis by the globular C-terminal domain. Ligands binding to the N-terminal domain abolish the spontaneous ionic currents associated with neurotoxic mutants of PrP, and the isolated N-terminal domain induces currents when expressed in the absence of the C-terminal domain. Anti-PrP antibodies targeting epitopes in the C-terminal domain induce currents, and cause degeneration of dendrites on murine hippocampal neurons, effects that entirely dependent on the effector function of the N-terminus. NMR experiments demonstrate intramolecular docking between N- and C-terminal domains of PrPC, revealing a novel auto-inhibitory mechanism that regulates the functional activity of PrPC.

Funding information:
  • NIGMS NIH HHS - P20 GM104316()
  • NIGMS NIH HHS - R01 GM065790()
  • NINDS NIH HHS - R01 NS065244()

Circ-ZNF609 Is a Circular RNA that Can Be Translated and Functions in Myogenesis.

  • Legnini I
  • Mol. Cell
  • 2017 Apr 6

Literature context:


Abstract:

Circular RNAs (circRNAs) constitute a family of transcripts with unique structures and still largely unknown functions. Their biogenesis, which proceeds via a back-splicing reaction, is fairly well characterized, whereas their role in the modulation of physiologically relevant processes is still unclear. Here we performed expression profiling of circRNAs during in vitro differentiation of murine and human myoblasts, and we identified conserved species regulated in myogenesis and altered in Duchenne muscular dystrophy. A high-content functional genomic screen allowed the study of their functional role in muscle differentiation. One of them, circ-ZNF609, resulted in specifically controlling myoblast proliferation. Circ-ZNF609 contains an open reading frame spanning from the start codon, in common with the linear transcript, and terminating at an in-frame STOP codon, created upon circularization. Circ-ZNF609 is associated with heavy polysomes, and it is translated into a protein in a splicing-dependent and cap-independent manner, providing an example of a protein-coding circRNA in eukaryotes.

Funding information:
  • Telethon - GGP16213()

Multilayered Control of Alternative Splicing Regulatory Networks by Transcription Factors.

  • Han H
  • Mol. Cell
  • 2017 Feb 2

Literature context:


Abstract:

Networks of coordinated alternative splicing (AS) events play critical roles in development and disease. However, a comprehensive knowledge of the factors that regulate these networks is lacking. We describe a high-throughput system for systematically linking trans-acting factors to endogenous RNA regulatory events. Using this system, we identify hundreds of factors associated with diverse regulatory layers that positively or negatively control AS events linked to cell fate. Remarkably, more than one-third of the regulators are transcription factors. Further analyses of the zinc finger protein Zfp871 and BTB/POZ domain transcription factor Nacc1, which regulate neural and stem cell AS programs, respectively, reveal roles in controlling the expression of specific splicing regulators. Surprisingly, these proteins also appear to regulate target AS programs via binding RNA. Our results thus uncover a large "missing cache" of splicing regulators among annotated transcription factors, some of which dually regulate AS through direct and indirect mechanisms.

Long Noncoding RNA Malat1 Regulates Cerebrovascular Pathologies in Ischemic Stroke.

  • Zhang X
  • J. Neurosci.
  • 2017 Feb 15

Literature context:


Abstract:

The study was designed to determine the role of long noncoding RNA (lncRNA), metastasis-associated lung adenocarcinoma transcript 1 (Malat1), in ischemic stroke outcome. Primary mouse brain microvascular endothelial cells (BMECs) were cultured and treated with Malat1 GapmeR before 16 h oxygen and glucose depravation (OGD). Cell death was assayed by LDH and MTT methods. Malat1 knock-out and wild-type mice were subjected to 1 h of middle cerebral artery occlusion (MCAO) and 24-72 h of reperfusion. To explore the underlying mechanism, apoptotic and inflammatory factors were measured by qPCR, ELISA, and Western blotting. The physical interaction between Malat1 and apoptotic or inflammatory factors was measured by RNA immunoprecipitation. Increased Malat1 levels were found in cultured mouse BMECs after OGD as well as in isolated cerebral microvessels in mice after MCAO. Silencing of Malat1 by Malat1 GapmeR significantly increased OGD-induced cell death and Caspase 3 activity in BMECs. Silencing of Malat1 also significantly aggravated OGD-induced expression of the proapoptotic factor Bim and proinflammatory cytokines MCP-1, IL-6, and E-selectin. Moreover, Malat1 KO mice presented larger brain infarct size, worsened neurological scores, and reduced sensorimotor functions. Consistent with in vitro findings, significantly increased expression of proapoptotic and proinflammatory factors was also found in the cerebral cortex of Malat1 KO mice after ischemic stroke compared with WT controls. Finally, we demonstrated that Malat1 binds to Bim and E-selectin both in vitro and in vivo Our study suggests that Malat1 plays critical protective roles in ischemic stroke.SIGNIFICANCE STATEMENT Accumulative studies have demonstrated the important regulatory roles of microRNAs in vascular and neural damage after ischemic stroke. However, the functional significance and mechanisms of other classes of noncoding RNAs in cerebrovascular pathophysiology after stroke are less studied. Here we demonstrate a novel role of Malat1, a long noncoding RNA that has been originally identified as a prognostic marker for non-small cell lung cancer, in cerebrovascular pathogenesis of ischemic stroke. Our experiments have provided the first evidence that Malat1 plays anti-apoptotic and anti-inflammatory roles in brain microvasculature to reduce ischemic cerebral vascular and parenchymal damages. Our studies also suggest that lncRNAs can be therapeutically targeted to minimize poststroke brain damage.

Funding information:
  • NINDS NIH HHS - R01 NS086820()
  • NINDS NIH HHS - R01 NS091175()
  • NINDS NIH HHS - R21 NS094930()

Suppression of C9orf72 RNA repeat-induced neurotoxicity by the ALS-associated RNA-binding protein Zfp106.

  • Celona B
  • Elife
  • 2017 Jan 10

Literature context:


Abstract:

Expanded GGGGCC repeats in the first intron of the C9orf72 gene represent the most common cause of familial amyotrophic lateral sclerosis (ALS), but the mechanisms underlying repeat-induced disease remain incompletely resolved. One proposed gain-of-function mechanism is that repeat-containing RNA forms aggregates that sequester RNA binding proteins, leading to altered RNA metabolism in motor neurons. Here, we identify the zinc finger protein Zfp106 as a specific GGGGCC RNA repeat-binding protein, and using affinity purification-mass spectrometry, we show that Zfp106 interacts with multiple other RNA binding proteins, including the ALS-associated factors TDP-43 and FUS. We also show that Zfp106 knockout mice develop severe motor neuron degeneration, which can be suppressed by transgenic restoration of Zfp106 specifically in motor neurons. Finally, we show that Zfp106 potently suppresses neurotoxicity in a Drosophila model of C9orf72 ALS. Thus, these studies identify Zfp106 as an RNA binding protein with important implications for ALS.

Funding information:
  • BLRD VA - I01 BX001108()
  • NHLBI NIH HHS - P01 HL089707()
  • NHLBI NIH HHS - R01 HL064658()
  • NIA NIH HHS - P01 AG019724()
  • NIA NIH HHS - P50 AG023501()
  • NINDS NIH HHS - R01 NS098516()
  • RRD VA - I01 RX002133()

Deficiency of CPEB2-Confined Choline Acetyltransferase Expression in the Dorsal Motor Nucleus of Vagus Causes Hyperactivated Parasympathetic Signaling-Associated Bronchoconstriction.

  • Lai YT
  • J. Neurosci.
  • 2016 Dec 14

Literature context:


Abstract:

Cytoplasmic polyadenylation element binding protein 2 (CPEB2) is an RNA-binding protein and translational regulator. To understand the physiological function of CPEB2, we generated CPEB2 knock-out (KO) mice and found that most died within 3 d after birth. CPEB2 is highly expressed in the brainstem, which controls vital functions, such as breathing. Whole-body plethysmography revealed that KO neonates had aberrant respiration with frequent apnea. Nevertheless, the morphology and function of the respiratory rhythm generator and diaphragm neuromuscular junctions appeared normal. We found that upregulated translation of choline acetyltransferase in the CPEB2 KO dorsal motor nucleus of vagus resulted in hyperactivation of parasympathetic signaling-induced bronchoconstriction, as evidenced by increased pulmonary acetylcholine and phosphorylated myosin light chain 2 in bronchial smooth muscles. Specific deletion of CPEB2 in cholinergic neurons sufficiently caused increased apnea in neonatal pups and airway hyper-reactivity in adult mice. Moreover, inhalation of an anticholinergic bronchodilator reduced apnea episodes in global and cholinergic CPEB2-KO mice. Together, the elevated airway constriction induced by cholinergic transmission in KO neonates may account for the respiratory defect and mortality. SIGNIFICANCE STATEMENT: This study first generated and characterized cpeb2 gene-deficient mice. CPEB2-knock-out (KO) mice are born alive but most die within 3 d after birth showing no overt defects in anatomy. We found that the KO neonates showed severe apnea and altered respiratory pattern. Such respiratory defects could be recapitulated in mice with pan-neuron-specific or cholinergic neuron-specific ablation of the cpeb2 gene. Further investigation revealed that cholinergic transmission in the KO dorsal motor nucleus of vagus was overactivated because KO mice lack CPEB2-suppressed translation of the rate-limiting enzyme in the production of acetylcholine (i.e., choline acetyltransferase). Consequently, increased parasympathetic signaling leads to hyperactivated bronchoconstriction and abnormal respiration in the KO neonates.

The Ubiquitin Receptor ADRM1 Modulates HAP40-Induced Proteasome Activity.

  • Huang ZN
  • Mol. Neurobiol.
  • 2016 Nov 7

Literature context:


Abstract:

Huntington's disease (HD) is a progressive neurodegenerative disorder caused by an N-terminal expansion of polyglutamine stretch (polyQ) of huntingtin (Htt) protein. HAP40 is a huntingtin-associated protein with unknown cellular functions. Increased HAP40 expression has been reported in the brain of HD patients and HD mouse model. However, the relationship between the elevation of HAP40 and HD etiology remains elusive. In this study, we demonstrated that overexpression of HAP40 enhanced accumulation of mutant Htt aggregates and caused defects in proteasome function. Specifically, excess HAP40 interfered with adhesion-regulating molecule 1 (ADRM1), a proteasome ubiquitin receptor, to regulate the proteasome-dependent pathway. Increasing ADRM1 in the presence of excess HAP40 alleviated mutant Htt aggregates and at the same time, restored the cell viability. Reducing ADRM1 in the absence of excess HAP40; on the other hand, increased mutant Htt aggregates and decreased the cell viability. Our data provide compelling evidence to support that ADRM1 plays an important role in mediating removal of mutant Htt aggregates when excess HAP40 is present. ADRM1-dependent ubiquitin proteasome system (UPS) may be a general mechanism to guard cells from mutant Htt toxicity.

Funding information:
  • NINDS NIH HHS - R37NS008174(United States)

Loss of Frataxin activates the iron/sphingolipid/PDK1/Mef2 pathway in mammals.

  • Chen K
  • Elife
  • 2016 Nov 30

Literature context:


Abstract:

Friedreich's ataxia (FRDA) is an autosomal recessive neurodegenerative disease caused by mutations in Frataxin (FXN). Loss of FXN causes impaired mitochondrial function and iron homeostasis. An elevated production of reactive oxygen species (ROS) was previously proposed to contribute to the pathogenesis of FRDA. We recently showed that loss of frataxin homolog (fh), a Drosophila homolog of FXN, causes a ROS independent neurodegeneration in flies (Chen et al., 2016). In fh mutants, iron accumulation in the nervous system enhances the synthesis of sphingolipids, which in turn activates 3-phosphoinositide dependent protein kinase-1 (Pdk1) and myocyte enhancer factor-2 (Mef2) to trigger neurodegeneration of adult photoreceptors. Here, we show that loss of Fxn in the nervous system in mice also activates an iron/sphingolipid/PDK1/Mef2 pathway, indicating that the mechanism is evolutionarily conserved. Furthermore, sphingolipid levels and PDK1 activity are also increased in hearts of FRDA patients, suggesting that a similar pathway is affected in FRDA.

Funding information:
  • NIDDK NIH HHS - U24 DK059637(United States)

C9orf72 Dipeptide Repeats Impair the Assembly, Dynamics, and Function of Membrane-Less Organelles.

  • Lee KH
  • Cell
  • 2016 Oct 20

Literature context:


Abstract:

Expansion of a hexanucleotide repeat GGGGCC (G4C2) in C9ORF72 is the most common cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Transcripts carrying (G4C2) expansions undergo unconventional, non-ATG-dependent translation, generating toxic dipeptide repeat (DPR) proteins thought to contribute to disease. Here, we identify the interactome of all DPRs and find that arginine-containing DPRs, polyGly-Arg (GR) and polyPro-Arg (PR), interact with RNA-binding proteins and proteins with low complexity sequence domains (LCDs) that often mediate the assembly of membrane-less organelles. Indeed, most GR/PR interactors are components of membrane-less organelles such as nucleoli, the nuclear pore complex and stress granules. Genetic analysis in Drosophila demonstrated the functional relevance of these interactions to DPR toxicity. Furthermore, we show that GR and PR altered phase separation of LCD-containing proteins, insinuating into their liquid assemblies and changing their material properties, resulting in perturbed dynamics and/or functions of multiple membrane-less organelles.

Enhancer Variants Synergistically Drive Dysfunction of a Gene Regulatory Network In Hirschsprung Disease.

  • Chatterjee S
  • Cell
  • 2016 Oct 6

Literature context:


Abstract:

Common sequence variants in cis-regulatory elements (CREs) are suspected etiological causes of complex disorders. We previously identified an intronic enhancer variant in the RET gene disrupting SOX10 binding and increasing Hirschsprung disease (HSCR) risk 4-fold. We now show that two other functionally independent CRE variants, one binding Gata2 and the other binding Rarb, also reduce Ret expression and increase risk 2- and 1.7-fold. By studying human and mouse fetal gut tissues and cell lines, we demonstrate that reduced RET expression propagates throughout its gene regulatory network, exerting effects on both its positive and negative feedback components. We also provide evidence that the presence of a combination of CRE variants synergistically reduces RET expression and its effects throughout the GRN. These studies show how the effects of functionally independent non-coding variants in a coordinated gene regulatory network amplify their individually small effects, providing a model for complex disorders.

Funding information:
  • NINDS NIH HHS - R56 NS046367(United States)

Cell-Type-Specific Alternative Splicing Governs Cell Fate in the Developing Cerebral Cortex.

  • Zhang X
  • Cell
  • 2016 Aug 25

Literature context:


Abstract:

Alternative splicing is prevalent in the mammalian brain. To interrogate the functional role of alternative splicing in neural development, we analyzed purified neural progenitor cells (NPCs) and neurons from developing cerebral cortices, revealing hundreds of differentially spliced exons that preferentially alter key protein domains-especially in cytoskeletal proteins-and can harbor disease-causing mutations. We show that Ptbp1 and Rbfox proteins antagonistically govern the NPC-to-neuron transition by regulating neuron-specific exons. Whereas Ptbp1 maintains apical progenitors partly through suppressing a poison exon of Flna in NPCs, Rbfox proteins promote neuronal differentiation by switching Ninein from a centrosomal splice form in NPCs to a non-centrosomal isoform in neurons. We further uncover an intronic human mutation within a PTBP1-binding site that disrupts normal skipping of the FLNA poison exon in NPCs and causes a brain-specific malformation. Our study indicates that dynamic control of alternative splicing governs cell fate in cerebral cortical development.

Autism-related deficits via dysregulated eIF4E-dependent translational control.

  • Gkogkas CG
  • Nature
  • 2013 Jan 17

Literature context:


Abstract:

Hyperconnectivity of neuronal circuits due to increased synaptic protein synthesis is thought to cause autism spectrum disorders (ASDs). The mammalian target of rapamycin (mTOR) is strongly implicated in ASDs by means of upstream signalling; however, downstream regulatory mechanisms are ill-defined. Here we show that knockout of the eukaryotic translation initiation factor 4E-binding protein 2 (4E-BP2)-an eIF4E repressor downstream of mTOR-or eIF4E overexpression leads to increased translation of neuroligins, which are postsynaptic proteins that are causally linked to ASDs. Mice that have the gene encoding 4E-BP2 (Eif4ebp2) knocked out exhibit an increased ratio of excitatory to inhibitory synaptic inputs and autistic-like behaviours (that is, social interaction deficits, altered communication and repetitive/stereotyped behaviours). Pharmacological inhibition of eIF4E activity or normalization of neuroligin 1, but not neuroligin 2, protein levels restores the normal excitation/inhibition ratio and rectifies the social behaviour deficits. Thus, translational control by eIF4E regulates the synthesis of neuroligins, maintaining the excitation-to-inhibition balance, and its dysregulation engenders ASD-like phenotypes.

Funding information:
  • Canadian Institutes of Health Research - MOP-10848()
  • Canadian Institutes of Health Research - MOP-114994()
  • Canadian Institutes of Health Research - MOP-44050()
  • Canadian Institutes of Health Research - MOP-93679()
  • NCI NIH HHS - R01 CA140456()
  • NCI NIH HHS - R01 CA154916()
  • NIGMS NIH HHS - R01 GM088813()
  • PHS HHS - U19 AUI082715(United States)

Direct conversion of fibroblasts to neurons by reprogramming PTB-regulated microRNA circuits.

  • Xue Y
  • Cell
  • 2013 Jan 17

Literature context:


Abstract:

The induction of pluripotency or trans-differentiation of one cell type to another can be accomplished with cell-lineage-specific transcription factors. Here, we report that repression of a single RNA binding polypyrimidine-tract-binding (PTB) protein, which occurs during normal brain development via the action of miR-124, is sufficient to induce trans-differentiation of fibroblasts into functional neurons. Besides its traditional role in regulated splicing, we show that PTB has a previously undocumented function in the regulation of microRNA functions, suppressing or enhancing microRNA targeting by competitive binding on target mRNA or altering local RNA secondary structure. A key event during neuronal induction is the relief of PTB-mediated blockage of microRNA action on multiple components of the REST complex, thereby derepressing a large array of neuronal genes, including miR-124 and multiple neuronal-specific transcription factors, in nonneuronal cells. This converts a negative feedback loop to a positive one to elicit cellular reprogramming to the neuronal lineage.

Funding information:
  • NCRR NIH HHS - R24 RR021346(United States)