289,744 research outputs found
Neto-β is a novel Neto isoform at the <i>Drosophila</i> NMJ.
No full text<p>(A) Diagram of the <i>Drosophila neto</i> locus. Two isoforms are generated by alternative splicing, Neto-α and Neto-β. They have different cytoplasmic domains, but share highly conserved domains, CUB (complement <u>C</u>1r/C1s, <u>U</u>egf, <u>B</u>MP1), LDLa (LDL receptor class a) and transmembrane (TM), with Neto proteins from vertebrates and <i>C</i>. <i>elegans</i>. (B) Confocal images of third instar larvae NMJ4 from <i>neto</i><sup><i>null</i></sup> animals rescued with <i>neto</i>-<i>α</i> (<i>neto</i><sup><i>36</i></sup>;<i>G14>neto</i><sup><i>α</i></sup>), <i>neto</i>-<i>β</i> (<i>neto</i><sup><i>36</i></sup>;<i>G14>neto</i><sup>β</sup>), and a <i>neto</i> transgene lacking any intracellular part, <i>neto</i>Δ<sup>intra</sup><i>(neto</i><sup><i>36</i></sup>;<i>G14>neto</i>Δ<sup>intra</sup>). These Neto variants can rescue the viability and NMJ development in <i>neto</i><sup><i>null</i></sup> animals. The Neto-ex signals mark all NMJs, but the Neto-β antibodies specifically label control and <i>neto-β</i> rescued NMJs. (C-D) Confocal images of late embryos ventral muscle fields (indicated in white) labeled for (C) Neto-ex (red), Neto-β (green), HRP (blue) and (D) GluRIIA (red), Neto-β (green), HRP (blue) indicating the presence of Neto-β in the early stages of the larvae development. Genotypes: control (<i>w</i><sup><i>1118</i></sup>); <i>neto</i><sup><i>36</i></sup>;<i>G14>neto</i><sup><i>α</i></sup> (<i>neto</i><sup><i>36</i></sup><i>/Y; G14-Gal4/UAS-neto-A9</i>); <i>neto</i><sup><i>36</i></sup>;<i>G14>neto</i><sup>β</sup> (<i>neto</i><sup><i>36</i></sup>, <i>UAS-neto-B6/Y; G14-Gal4/+</i>); <i>neto</i><sup><i>36</i></sup>;<i>G14>neto</i>Δ<sup>intra</sup>(<i>neto</i><sup><i>36</i></sup><i>/Y; G14-Gal4/ UAS-neto</i>Δ<sup>intra</sup><i>-H4</i>). Scale bars: (B) 20 μm (C-D) 10 μm; 5 μm in details.</p
Neto-β, but not Neto-α, restores PAK recruitment and mEJP amplitude at <i>neto</i><sup><i>null</i></sup> NMJs.
No full text<p>(A) Confocal images of larval NMJ4 labeled for Neto-ex (red), PAK (green) and HRP (blue). Neto-β, but not Neto-α, restores PAK synaptic accumulation over a large range of concentrations tested. (B) Western blot comparison of Neto levels in muscle extracts from control (first lane), and <i>neto</i><sup><i>null</i></sup> larvae rescued with <i>neto-α</i> transgenes (low, medium, and high expression) (magenta gradient), or <i>neto-β</i> transgenes (low, medium, high, and very high expression)(blue gradient). Arrows indicate unprocessed and processed Neto-α (magenta) and Neto-β (blue). (B’)—low exposure. Tubulin was used as a loading control. (C-H) Electrophysiological recordings of <i>neto</i><sup><i>null</i></sup> NMJs rescued by various levels of Neto-α or Neto-β. Data are reported relative to controls matched by rearing at 18°C (empty bars) or 25°C (hatched bars). Representative traces of mEJPs and EJPs are shown in (C) and (F), respectively. The mEJPs amplitude is reduced at NMJs rescued by low and medium levels of Neto-α (D). The mEJPs frequency appears less dependent on Neto levels/isoforms, but is significantly increased in larvae reared at 18°C (E). The EJPs amplitudes are largely normal (G), likely due to subtle, but significant increases in quantal content at Neto-α-rescued NMJs (H). (I) Confocal images of NMJ4 boutons (segment A3) in control (<i>w</i><sup><i>1118</i></sup>) and <i>GluRIIA</i><sup><i>SP16/Df</i></sup> (<i>GluRIIA</i><sup><i>SP16</i></sup>/ <i>Df(2L)cl</i><sup><i>h4</i></sup>) third instar larvae labeled for Neto-ex (red), PAK (green), and GluRIIA (blue). PAK is normally present at <i>GluRIIA</i> mutant synapses, indicating that the synaptic recruitment PAK does not depend on GluRIIA. Genotypes: control (<i>G14-Gal4/+</i>); <i>neto</i><sup><i>α low</i></sup> (<i>neto</i><sup><i>36</i></sup><i>/Y</i>; <i>G14-Gal4/UAS-neto-A9</i>), reared at 25°C); <i>neto</i><sup><i>α med</i></sup> (<i>neto</i><sup><i>36</i></sup><i>/Y</i>; <i>G14-Gal4/UAS-neto-A3</i>, 18°C); <i>neto</i><sup><i>α high</i></sup> (<i>neto</i><sup><i>36</i></sup><i>/Y</i>; <i>G14-Gal4/UAS-neto-A3</i>, 25°C); <i>neto</i><sup><i>β low</i></sup> (<i>neto</i><sup><i>36</i></sup>, <i>UAS-neto-B6/Y</i>; <i>G14-Gal4/+</i>, 18°C); <i>neto</i><sup><i>β med</i></sup> (<i>neto</i><sup><i>36</i></sup>, <i>UAS-neto-B6</i>/Y; <i>G14-Gal4/+</i>, 25°C); <i>neto</i><sup><i>β high</i></sup> (<i>neto</i><sup><i>36</i></sup><i>/Y</i>; <i>G14-Gal4/UAS-neto-B3</i>, 18°C); <i>neto</i><sup><i>β very high</i></sup> (<i>neto</i><sup><i>36</i></sup><i>/Y</i>; <i>G14-Gal4/UAS-neto-B3</i>, 25°C). Error bars indicate SEM. ***; p<0.001, **; p<0.005, *; p<0.05, ns; p>0.05. Scale bars: 20 μm, 2 μm in details and (I).</p
Leituras críticas da obra de João Simões Lopes Neto: Província de São Pedro e Caderno de Sábado
Get PDFDissertação (mestrado) - Universidade Federal de Santa Catarina, Centro de Comunicação e Expressão. Programa de Pós-graduação em LiteraturaEste trabalho tem como objetivo reunir e recuperar leituras críticas esparsas da obra de João Simões Lopes Neto publicadas na revista Província de São Pedro e no suplemento literário Caderno de Sábado, facilitando a pesquisa de outros estudiosos interessados em sua obra. O trabalho de transcrição foi realizado seguindo as normas padronizadas pela Filologia contemporânea, a fim de determinar os critérios adotados para a transcrição dos textos. Num primeiro momento, apresenta-se a apreciação da obra simoniana por parte dos críticos e, num segundo momento, são apresentados os vinte e oito textos selecionados para a transcrição
Cearagrylloides Martins-Neto
No full text<i>Cearagrylloides</i> Martins-Neto n. gen. <p> <b>Etymology.</b> A quite <i>Cearagryllus</i>.</p> <p> <b>Type species.</b> <i>Cearagryllus perforatorius</i> Martins-Neto, 1991, designated here. <b>Diagnosis</b> (female). Robust cearagryllids with relatively small head. forewing length equal to the body length; ovipositor long and setiform, longer than the forewing/body length.</p> <p> <b>Discussion.</b> The Araripe female cearagryllines, although reasonably similar to the male ones, cannot be confidently placed in any of the genera <i>Cearagryllus</i>, <i>Notocearagryllus, Allocearagryllus</i> or <i>Cryptocearagryllus</i>. The generic separation is necessary, although artificial, to prevent mistakes of interpretation. The new genus is useful for paleoecological purposes, as discussed below.</p>Published as part of <i>Martins-Neto, Rafael Gioia & Tassi, Lara Vaz, 2009, The Orthoptera (Ensifera) from the Santana formation (Early Cretaceous, Northeast Brazil): A statistical and paleoecological approach, with description of new taxa, pp. 21-37 in Zootaxa 2080</i> on page 27, DOI: <a href="http://zenodo.org/record/187344">10.5281/zenodo.187344</a>
Generation and characterization of <i>neto-β</i> isoform specific alleles.
No full text<p>(A) Schematics of the <i>Minos</i> transposomal elements and the small lesions corresponding to various <i>neto</i> alleles. <i>MB07125</i> was mobilized to generate precise excision control and isoform specific <i>neto-β</i> alleles, <i>neto</i><sup><i>βshort</i></sup> (short cytoplasmic tail) and <i>neto</i><sup><i>βnull</i></sup>. The breakpoint coordinates are indicated. (B) Diagram of the predicted Neto-β proteins. The bars mark the antigens for Neto-ex, Neto-β1 and Neto-β2 antibodies. (C) Western blot analysis of lysates from S2 cells (left) and larval muscle (right). S2 cells were transfected with empty vector (lane 1), Neto-β (2), Neto-β<sup>short</sup> (3), and Neto-α (4) expression constructs and the lysates were compared with muscle extracts from control (5), <i>neto</i><sup><i>βshort</i></sup> (6), and <i>neto</i><sup><i>βnull</i></sup> (7) third instar larvae. Full length (black arrow) and truncated (white arrow) Neto-β are indicated. No specific signal was detected in <i>neto</i><sup><i>βnull</i></sup> animals. (D-E) Confocal images of boutons at NMJ4 of third instar larvae stained with Neto-ex (red) and with either Neto-β2 (D) or Neto-β1 (E) (green). As expected, Neto-ex signals were detected in control (precise excision) and <i>neto</i> alleles. <i>neto</i><sup><i>βshort</i></sup> NMJs show Neto-β2 signals (D) but both <i>neto-β</i> alleles lack Neto-β1 synaptic signals (E). In contrast, Neto-β1 puncta are present at <i>neto</i><sup><i>hypo</i></sup> NMJs. Scale bars: 2 μm.</p
The prodomain restricts Neto activities.
No full text<p>(A) Diagram and Western blot analysis of constitutively active (CA) and processing mutant (PM) Neto-GFP variants. (B) Fluorescence images of salivary glands from third instar larvae expressing Neto-GFP variants as indicated (<i>G14>neto-GFP</i>). Prodomain processing did not affect the apical localization of Neto. (C) Co-immunoprecipitation (IP: α-GFP, WB: α-GluRIIC) from muscle extracts from control (<i>y<sup>1</sup>w<sup>1118</sup></i>) and <i>neto</i> rescued larvae (<i>neto<sup>36</sup>;G14>CA-neto-GFP</i> and <i>neto<sup>36</sup>;G14>PM-neto-GFP</i>) showed that both CA- and PM-Neto bind iGluRs. (D-E) PM-Neto is less efficient in rescuing the embryonic lethality and adult viability of <i>neto</i> null mutants compared with control or CA-Neto. Genotypes: control (<i>neto<sup>36</sup>;G14>neto-GFP</i> at 25°C); CA-Neto normal (<i>neto<sup>36</sup>;G14>CA-neto-GFP-N4</i> at 18°C); CA-Neto high (<i>neto<sup>36</sup>;G14>CA-neto-GFP-N4</i> at 25°C); PM-Neto normal (<i>neto<sup>36</sup>;G14>PM-neto-GFP-D2</i> at 25°C); PM-Neto high (<i>neto<sup>36</sup>;G14>PM-neto-GFP-D1</i> at 25°C). The numbers of animals analyzed are indicated in each bar. Bars: 10 μm.</p
Modulation of Neto processing.
No full text<p>(A) Western blot comparison of the extent of Neto processing in fed and starved larvae. Third instar larvae (<i>neto<sup>36</sup>;24B>neto-V5</i>) were separated into two groups: one group was fed with yeast paste and the other was starved. Muscle extracts were analyzed at indicated time points and the relative ratio of processed Neto was calculated against total Neto (B). Values were averaged from three individual experiments. **; <i>p</i><0.01</p
iGluRs synaptic accumulation is perturbed at <i>neto-β</i> mutant NMJs.
No full text<p>(A) Confocal images of NMJ4 boutons in larvae of indicated genotypes labeled for Brp (red), GluRIIC (green), and HRP (blue) (quantified in B-C). <i>neto-β</i> mutant NMJs have increased number of synaptic contacts. The intensity of the presynaptic active zone marker Brp appears to be normal, but the GluRIIC synaptic signals are reduced at <i>neto-β</i> mutant NMJs compared with control (precise excision). (D) Western blot comparison of GluRIIC protein levels in muscle lysates from <i>neto-β</i> third instar larvae. Tubulin was used as a loading control. (E) Confocal images of NMJ4 boutons in larvae of indicated genotypes labeled for GluRIIA (red), GluRIIB (green), and HRP (blue). The synaptic accumulation of GluRIIA is severely reduced at <i>neto-β</i> mutant NMJs (quantified in C). In contrast, the GluRIIB synaptic signals are slightly increased at <i>neto</i><sup><i>βshort</i></sup> NMJs and significantly reduced (by 31%) at <i>neto</i><sup><i>βnull</i></sup> NMJs. (F) Confocal images of NMJ4 boutons in control and <i>neto-α</i><sup><i>RNAi</i></sup> larvae labeled for GluRIIA (red), GluRIIB (green), and HRP (blue). The GluRIIB signals, but not GluRIIA are reduced at Neto-α-depleted NMJs (quantified in G). (H) Western blot analysis of larval muscle extracts from <i>neto</i><sup><i>null</i></sup> mutants rescued by V5-tagged Neto- and carried through RNAi-mediated knockdown as indicated. The <i>neto-</i><sup>RNAi</sup> appears to be more effective than the <i>CUB1</i><sup><i>RNAi</i></sup> in knocking down V5-tagged Neto- relative to the Tubulin control. Error bars indicate SEM. ***; p<0.001, **; p<0.005, *; p<0.05, ns; p>0.05. Scale bars: 2 μm.</p
Ultrastructure defects at <i>neto-β</i> mutant boutons.
No full text<p>(A–C) Electron micrographs of type Ib boutons in third instar larvae of indicated genotypes. The upper panels show entire boutons; the active zones (AZ, arrows), mitochondria (m), and subsynaptic reticulum (SSR, brackets) are indicated. The <i>neto-β</i> mutant boutons have numerous synaptic contacts, but their active zones often have abnormal T-bar structures (B’-B’”, C’-C’”), including closely spaced, distorted, fused, and floating T-bars. The PSDs length, quantified in serial section (D), is significantly reduced at <i>neto-β</i> mutant boutons. The boutons appear enlarged and the SSR area and thickness diminished in both <i>neto-β</i> mutants (quantified in E-H). Error bars indicate SEM. ***; p<0.001, **; p<0.005. Scale bars: 1 μm, 200 nm in details.</p
Prodomain processing affects Neto-mediated iGluR clustering.
No full text<p>(A-B) Confocal images of NMJ4 (A) and bouton details (B) in larvae of indicated genotypes labeled for Brp (green), GluRIIC (red), and Neto (blue). Similar to <i>neto</i> transgenes, <i>CA-neto</i> induced dose-dependent gain-of-function NMJ phenotypes. In contrast, <i>PM-neto</i> transgenes severely disrupted the NMJ morphology and the synaptic contacts. (C) Confocal images of NMJ4 labeled for GluRIIA (green), GluRIIB (red), and HRP (blue) uncovered a drastic reduction of GluRIIA synaptic signals at PM-Neto rescued NMJs. (D) Western blot comparison of Neto expression levels in muscle extracts from third instar larvae rescued with: (i) untagged Neto (lane 2, <i>neto<sup>36</sup>;G14>neto-A3</i>); (ii) CA-Neto-GFP, normal (lane 3) and high (lane 4); and (iii) PM-Neto-GFP, normal (lane 5) and high (lane 6). (*) uncleaved proteins. (E-F) Quantification of various synaptic signals at <i>neto</i> null NMJs rescued with normal levels of CA- and PM-Neto. (G) Bouton numbers were severely reduced at PM-Neto rescued NMJs. The numbers of NMJs examined are indicated in each bar. Genotypes: CA-Neto normal (<i>neto<sup>36</sup>;G14>CA-neto-GFP-N4</i> at 18°C); CA-Neto high (<i>neto<sup>36</sup>;G14>CA-neto-GFP-N4</i> at 25°C); PM-Neto normal (<i>neto<sup>36</sup>;G14>PM-neto-GFP-D2</i> at 25°C); PM-Neto high (<i>neto<sup>36</sup>;G14>PM-neto-GFP-D1</i> at 25°C). Error bars indicate SEM. *; <i>p</i><0.001, **; <i>p</i><0.01. Bars: 10 μm, 1 μm in details.</p
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