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1.
Elife ; 112022 03 10.
Artigo em Inglês | MEDLINE | ID: mdl-35266450

RESUMO

Mutations altering the scaffolding protein Shank are linked to several psychiatric disorders, and to synaptic and behavioral defects in mice. Among its many binding partners, Shank directly binds CaV1 voltage activated calcium channels. Here, we show that the Caenorhabditis elegans SHN-1/Shank promotes CaV1 coupling to calcium activated potassium channels. Mutations inactivating SHN-1, and those preventing SHN-1 binding to EGL-19/CaV1 all increase action potential durations in body muscles. Action potential repolarization is mediated by two classes of potassium channels: SHK-1/KCNA and SLO-1 and SLO-2 BK channels. BK channels are calcium-dependent, and their activation requires tight coupling to EGL-19/CaV1 channels. SHN-1's effects on AP duration are mediated by changes in BK channels. In shn-1 mutants, SLO-2 currents and channel clustering are significantly decreased in both body muscles and neurons. Finally, increased and decreased shn-1 gene copy number produce similar changes in AP width and SLO-2 current. Collectively, these results suggest that an important function of Shank is to promote microdomain coupling of BK with CaV1.


Assuntos
Proteínas de Caenorhabditis elegans , Canais de Potássio Ativados por Cálcio de Condutância Alta , Potenciais de Ação , Animais , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Cálcio/metabolismo , Cálcio da Dieta , Proteínas de Transporte/metabolismo , Humanos , Canais de Potássio Ativados por Cálcio de Condutância Alta/genética , Canais de Potássio Ativados por Cálcio de Condutância Alta/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Camundongos
2.
Elife ; 102021 03 31.
Artigo em Inglês | MEDLINE | ID: mdl-33787493

RESUMO

The development of functional synapses in the nervous system is important for animal physiology and behaviors, and its disturbance has been linked with many neurodevelopmental disorders. The synaptic transmission efficacy can be modulated by the environment to accommodate external changes, which is crucial for animal reproduction and survival. However, the underlying plasticity of synaptic transmission remains poorly understood. Here we show that in Caenorhabditis elegans, the male environment increases the hermaphrodite cholinergic transmission at the neuromuscular junction (NMJ), which alters hermaphrodites' locomotion velocity and mating efficiency. We identify that the male-specific pheromones mediate this synaptic transmission modulation effect in a developmental stage-dependent manner. Dissection of the sensory circuits reveals that the AWB chemosensory neurons sense those male pheromones and further transduce the information to NMJ using cGMP signaling. Exposure of hermaphrodites to the male pheromones specifically increases the accumulation of presynaptic CaV2 calcium channels and clustering of postsynaptic acetylcholine receptors at cholinergic synapses of NMJ, which potentiates cholinergic synaptic transmission. Thus, our study demonstrates a circuit mechanism for synaptic modulation and behavioral flexibility by sexual dimorphic pheromones.


Assuntos
Caenorhabditis elegans/fisiologia , Feromônios/metabolismo , Transmissão Sináptica , Animais , Feminino , Masculino , Junção Neuromuscular/fisiologia , Fatores Sexuais
3.
Cell Rep ; 28(11): 2979-2995.e4, 2019 09 10.
Artigo em Inglês | MEDLINE | ID: mdl-31509756

RESUMO

Munc13 proteins play several roles in regulating short-term synaptic plasticity. However, the underlying molecular mechanisms remain largely unclear. Here we report that C. elegans UNC-13L, a Munc13-1 ortholog, has three domains that inhibit synaptic vesicle (SV) exocytosis. These include the X (sequence between C2A and C1), C1, and C2B domains. Deleting all three inhibitory domains produces a hyperactive UNC-13 (sUNC-13) that exhibits dramatically increased neurotransmitter release, Ca2+ sensitivity of release, and release probability. The vesicular pool in unc-13 mutants rescued by sUNC-13 exhibits a faster synaptic recovery and replenishment rate, demonstrating an important role of sUNC-13 in regulating synaptic plasticity. Analysis of double mutants suggests that sUNC-13 enhances tonic release by increasing the open probability of UNC-64/syntaxin-1A, whereas its effects on evoked release appear to be mediated by additional functions, presumably by further regulating the activity of the assembled soluble N-ethylmaleimide-sensitive factor activating protein receptor (SNARE) complex.


Assuntos
Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Cálcio/metabolismo , Exocitose/fisiologia , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Transmissão Sináptica/genética , Vesículas Sinápticas/metabolismo , Animais , Animais Geneticamente Modificados , Caenorhabditis elegans/genética , Exocitose/genética , Mutação , Plasticidade Neuronal/genética , Plasticidade Neuronal/fisiologia , Probabilidade , Domínios Proteicos , Proteínas SNARE/metabolismo , Transmissão Sináptica/fisiologia , Vesículas Sinápticas/genética , Sintaxina 1/metabolismo
4.
Elife ; 82019 02 25.
Artigo em Inglês | MEDLINE | ID: mdl-30802206

RESUMO

UNC-13 proteins play an essential role in synaptic transmission by recruiting synaptic vesicles (SVs) to become available for release, which is termed SV priming. Here we show that the C2A domain of UNC-13L, like the corresponding domain in mammalian Munc13-1, displays two conserved binding modes: forming C2A/C2A homodimers, or forming a heterodimer with the zinc finger domain of UNC-10/RIM (C2A/RIM). Functional analysis revealed that UNC-13L's C2A promotes synaptic transmission by regulating a post-priming process. Stimulus-evoked release but not SV priming, was impaired in unc-10 mutants deficient for C2A/RIM heterodimerization, leading to decreased release probability. Disrupting C2A/C2A homodimerization in UNC-13L-rescued animals had no effect on synaptic transmission, but fully restored the evoked release and the release probability of unc-10/RIM mutants deficient for C2A/RIM heterodimerization. Thus, our results support the model that RIM binding C2A releases UNC-13L from an autoinhibitory homodimeric complex to become fusion-competent by functioning as a switch only.


Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Proteínas de Transporte/metabolismo , Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Multimerização Proteica , Transmissão Sináptica , Vesículas Sinápticas/metabolismo , Animais , Ligação Proteica , Domínios Proteicos
5.
Elife ; 72018 07 17.
Artigo em Inglês | MEDLINE | ID: mdl-30014846

RESUMO

Nitric oxide (NO) is released into the air by NO-producing organisms; however, it is unclear if animals utilize NO as a sensory cue. We show that C. elegans avoids Pseudomonas aeruginosa (PA14) in part by detecting PA14-produced NO. PA14 mutants deficient for NO production fail to elicit avoidance and NO donors repel worms. PA14 and NO avoidance are mediated by a chemosensory neuron (ASJ) and these responses require receptor guanylate cyclases and cyclic nucleotide gated ion channels. ASJ exhibits calcium increases at both the onset and removal of NO. These NO-evoked ON and OFF calcium transients are affected by a redox sensing protein, TRX-1/thioredoxin. TRX-1's trans-nitrosylation activity inhibits the ON transient whereas TRX-1's de-nitrosylation activity promotes the OFF transient. Thus, C. elegans exploits bacterially produced NO as a cue to mediate avoidance and TRX-1 endows ASJ with a bi-phasic response to NO exposure.


Assuntos
Caenorhabditis elegans/microbiologia , Caenorhabditis elegans/fisiologia , Células Quimiorreceptoras/fisiologia , Neurotransmissores/metabolismo , Óxido Nítrico/metabolismo , Pseudomonas aeruginosa/metabolismo , Tiorredoxinas/metabolismo , Animais , Comportamento Animal/efeitos dos fármacos , Caenorhabditis elegans/efeitos dos fármacos , Sinalização do Cálcio/efeitos dos fármacos , Células Quimiorreceptoras/efeitos dos fármacos , Processamento de Proteína Pós-Traducional
6.
Elife ; 62017 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-28477407

RESUMO

Shank is a post-synaptic scaffolding protein that has many binding partners. Shank mutations and copy number variations (CNVs) are linked to several psychiatric disorders, and to synaptic and behavioral defects in mice. It is not known which Shank binding partners are responsible for these defects. Here we show that the C. elegans SHN-1/Shank binds L-type calcium channels and that increased and decreased shn-1 gene dosage alter L-channel current and activity-induced expression of a CRH-1/CREB transcriptional target (gem-4 Copine), which parallels the effects of human Shank copy number variations (CNVs) on Autism spectrum disorders and schizophrenia. These results suggest that an important function of Shank proteins is to regulate L-channel current and activity induced gene expression.


Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Canais de Cálcio Tipo L/metabolismo , Cálcio/metabolismo , Cátions Bivalentes/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Fatores de Transcrição/metabolismo , Animais , Caenorhabditis elegans , Músculos/fisiologia
7.
Genetics ; 204(3): 1151-1159, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27585848

RESUMO

During larval molts, Caenorhabditis elegans exhibits a sleep-like state (termed lethargus) that is characterized by the absence of feeding and profound locomotion quiescence. The rhythmic pattern of locomotion quiescence and arousal linked to the molting cycle is mediated by reciprocal changes in sensory responsiveness, whereby arousal is associated with increased responsiveness. Sensory neurons arouse locomotion via release of a neuropeptide (PDF-1) and glutamate. Here we identify a second arousing neuropeptide (FLP-2). We show that FLP-2 acts via an orexin-like receptor (FRPR-18), and that FLP-2 and PDF-1 secretion are regulated by reciprocal positive feedback. These results suggest that the aroused behavioral state is stabilized by positive feedback between two neuropeptides.


Assuntos
Nível de Alerta , Proteínas de Caenorhabditis elegans/genética , Caenorhabditis elegans/genética , Locomoção , Neuropeptídeos/genética , Animais , Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/fisiologia , Proteínas de Caenorhabditis elegans/metabolismo , Retroalimentação Fisiológica , Ácido Glutâmico/metabolismo , Neuropeptídeos/metabolismo , Receptores de Orexina/genética , Receptores de Orexina/metabolismo , Células Receptoras Sensoriais/metabolismo , Células Receptoras Sensoriais/fisiologia
8.
Nature ; 496(7444): 181-6, 2013 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-23542588

RESUMO

Meristems encompass stem/progenitor cells that sustain postembryonic growth of all plant organs. How meristems are activated and sustained by nutrient signalling remains enigmatic in photosynthetic plants. Combining chemical manipulations and chemical genetics at the photoautotrophic transition checkpoint, we reveal that shoot photosynthesis-derived glucose drives target-of-rapamycin (TOR) signalling relays through glycolysis and mitochondrial bioenergetics to control root meristem activation, which is decoupled from direct glucose sensing, growth-hormone signalling and stem-cell maintenance. Surprisingly, glucose-TOR signalling dictates transcriptional reprogramming of remarkable gene sets involved in central and secondary metabolism, cell cycle, transcription, signalling, transport and protein folding. Systems, cellular and genetic analyses uncover TOR phosphorylation of E2Fa transcription factor for an unconventional activation of S-phase genes, and glucose-signalling defects in e2fa root meristems. Our findings establish pivotal roles of glucose-TOR signalling in unprecedented transcriptional networks wiring central metabolism and biosynthesis for energy and biomass production, and integrating localized stem/progenitor-cell proliferation through inter-organ nutrient coordination to control developmental transition and growth.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , Glucose/metabolismo , Meristema/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Transdução de Sinais , Transcriptoma , Arabidopsis/crescimento & desenvolvimento , Citocininas/metabolismo , Fatores de Transcrição E2F/metabolismo , Ativação Enzimática , Redes Reguladoras de Genes/genética , Ácidos Indolacéticos/metabolismo , Meristema/genética , Meristema/crescimento & desenvolvimento , Fosforilação , Fotossíntese , Fase S/genética , Transcrição Gênica/genética , Ativação Transcricional , Transcriptoma/genética
9.
Proc Natl Acad Sci U S A ; 105(6): 2226-31, 2008 Feb 12.
Artigo em Inglês | MEDLINE | ID: mdl-18256186

RESUMO

Cytokinesis partitions the cell by a cleavage furrow in animals but by a new cross wall in plants. How this new wall assembles at the molecular level and connects with the mother cell wall remains unclear. A lethal Arabidopsis embryogenesis mutant designated root-, shoot-, hypocotyl-defective (rsh) provides some clues: RSH encodes extensin AtEXT3, a structural glycoprotein located in the nascent cross wall or "cell plate" and also in mature cell walls. Here we report that electron micrographs of rsh mutant cells lacking RSH extensin correspond to a wall phenotype typified by incomplete cross wall assembly. Biochemical characterization of the purified RSH glycoprotein isolated from wild-type Arabidopsis cell cultures confirmed its identity as AtEXT3: a (hydroxy)proline-rich glyco protein comprising 11 identical amphiphilic peptide repeats with a 28-residue periodicity: SOOOOKKHYVYKSOOOOVKHYSOOOVYH (O = Hyp), each repeat containing a hydrophobic isodityrosine cross-link motif (YVY, underlined). Atomic force microscopy of RSH glycoprotein imaged its propensity for self-assembly into a dendritic scaffold. Extensin peroxidase catalyzed in vitro formation of insoluble RSH gels with concomitant tyrosine cross-linking, hence this likelihood in muro. We conclude that self-assembling amphiphiles of lysine-rich RSH extensin form positively charged scaffolds in the cell plate. These react with negatively charged pectin to create an extensin pectate coacervate that may template further orderly deposition of the new cross wall at cytokinesis.


Assuntos
Arabidopsis/citologia , Parede Celular/ultraestrutura , Glicoproteínas/fisiologia , Proteínas de Plantas/fisiologia , Sequência de Aminoácidos , Glicoproteínas/química , Microscopia de Força Atômica , Microscopia Eletrônica de Transmissão , Dados de Sequência Molecular , Proteínas de Plantas/química
10.
Science ; 300(5617): 332-6, 2003 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-12690200

RESUMO

Glucose modulates many vital processes in photosynthetic plants. Analyses of Arabidopsis glucose insensitive2 (gin2) mutants define the physiological functions of a specific hexokinase (HXK1) in the plant glucose-signaling network. HXK1 coordinates intrinsic signals with extrinsic light intensity. HXK1 mutants lacking catalytic activity still support various signaling functions in gene expression, cell proliferation, root and inflorescence growth, and leaf expansion and senescence, thus demonstrating the uncoupling of glucose signaling from glucose metabolism. The gin2 mutants are also insensitive to auxin and hypersensitive to cytokinin. Plants use HXK as a glucose sensor to interrelate nutrient, light, and hormone signaling networks for controlling growth and development in response to the changing environment.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimologia , Glucose/metabolismo , Hexoquinase/metabolismo , Luz , Transdução de Sinais , Aminoácidos Cíclicos/farmacologia , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Catálise , Divisão Celular , Citocininas/metabolismo , Citocininas/farmacologia , Etilenos/metabolismo , Flores/crescimento & desenvolvimento , Expressão Gênica , Genes de Plantas , Hexoquinase/genética , Ácidos Indolacéticos/metabolismo , Ácidos Indolacéticos/farmacologia , Mutagênese , Fosforilação , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/metabolismo , Raízes de Plantas/crescimento & desenvolvimento , Plantas Geneticamente Modificadas
11.
Plant Cell ; 14(5): 1161-72, 2002 May.
Artigo em Inglês | MEDLINE | ID: mdl-12034904

RESUMO

Although a large number of embryo mutants have been studied, mostly at the morphological level, the critical molecular and cellular events responsible for embryogenesis are unknown. Here, we report that using an enhancer-trap embryo mutant of Arabidopsis, we identified a gene, ROOT-SHOOT-HYPOCOTYL-DEFECTIVE (RSH), that is essential for the correct positioning of the cell plate during cytokinesis in cells of the developing embryo. We traced the earliest point of influence of RSH to the first asymmetrical division of the zygote. Homozygous rsh embryos were defective morphologically, had irregular cell shape and size, and germinated to form agravitropic-defective seedlings incapable of further development. The RSH gene encodes a Hyp-rich glycoprotein-type cell wall protein. RSH localized to the cell wall throughout the embryo and to a few well-defined postembryonic sites. Although several lines of evidence from previous work suggest that the cell wall is involved in development, the protein(s) involved remained elusive.


Assuntos
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Glicoproteínas/genética , Proteínas de Plantas , Sementes/crescimento & desenvolvimento , Sequência de Aminoácidos , Arabidopsis/embriologia , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Divisão Celular/genética , Tamanho Celular , Parede Celular/metabolismo , Parede Celular/ultraestrutura , Mapeamento Cromossômico , Proteínas do Citoesqueleto , Perfilação da Expressão Gênica , Glicoproteínas/metabolismo , Microscopia Eletrônica , Dados de Sequência Molecular , Mutação , Homologia de Sequência de Aminoácidos
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