Exploring Excitotoxicity and Regulation of a Constitutively Active TRP Ca<sup>2+</sup> Channel in Drosophila

Shieh, Bih‐Hwa; Nuzum, Lucinda; Kristaponyte, Inga

doi:10.1080/19336934.2020.1851586

Cited by 3 publications

(3 citation statements)

References 61 publications

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“…Specifically, we employed the GMR driver ( 24 ) to direct the expression of double-strand RNA using the UAS/GAL4 binary system ( 25 ). Subsequently, we examined the retinas of live flies using Arrestin 2-GFP (Arr2-GFP) ( 26 ) or Actin-GFP ( 27 , 28 ). This use of two GFP reporters allows us to explore specifically whether Rh1 or the actin cytoskeleton of the rhabdomere may be affected ( Fig.…”

Section: Resultsmentioning

confidence: 99%

A conventional PKC critical for both the light-dependent and the light-independent regulation of the actin cytoskeleton in Drosophila photoreceptors

Shieh

Sun

Ferng

2023

Journal of Biological Chemistry

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Section: Resultsmentioning

confidence: 99%

A conventional PKC critical for both the light-dependent and the light-independent regulation of the actin cytoskeleton in Drosophila photoreceptors

Shieh

Sun

Ferng

2023

Journal of Biological Chemistry

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“…The following transgenic stocks were used: ppk-GAL4, UAS-CD4::tdGFP ( Bhogal et al . 2016 ); UAS-mCherry.scramble.sponge (Bloomington Stock 61501); ppk-GAL4 (Bloomington Stock 32079); UAS-mCherry.miR-276a.sponge (Bloomington Stock 61406); UAS-mCherry.miR-276b.sponge (Bloomington Stock 61407); UAS-mCherry.mir-9c.sponge (Bloomington Stock 61376); UAS-mCherry.mir-125.sponge (Bloomington Stock 61393); UAS-Fmr1.Z (Bloomington Stock 6931); UAS-fmr1-RNAi (Bloomington Stock 34944; TRiP HMS00248); UAS-Nf1-RNAi (Bloomington Stock 53322; TRiP HMC03551) (validated by Moscato et al 2020 ); UAS-nej-RNAi (Bloomington Stock 37489; TRiP HMS01507) (validated by Jia et al 2015 ); UAS-inaE-RNAi (Bloomington Stock 64885; TRiP HMC05758) (validated by Shieh et al 2021 ); UAS-Hers-RNAi (Bloomington Stock 61858; TRiP HMJ23347); UAS-Ric-RNAi (Bloomington Stock 82973; TRiP HMC06651); UAS-Mkp3-RNAi (Bloomington Stock 57030; TRiP HMS04475); and UAS-Axn-RNAi (Bloomington Stock 62434; TRiP HMJ23888) (validated by Nye et al 2020 ). ppk-GAL4 was used to drive expression of UAS transgenes specifically in C4da neurons.…”

Section: Methodsmentioning

confidence: 99%

Drosophila FMRP controls miR-276-mediated regulation of nejire mRNA for space-filling dendrite development

Gavis

2022

G3 Genes|Genomes|Genetics

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MicroRNAs (miRNAs) are enriched in neurons and play important roles in dendritic spine development and synaptic plasticity. miRNA activity is controlled by a wide range of RNA-binding proteins (RBPs). Fragile X mental retardation protein (FMRP), a highly conserved RBP, has been linked to miRNA-mediated gene regulation in axonal development and dendritic spine formation. FMRP also participates in dendritic arbor morphogenesis, but whether and how miRNAs contribute to its function in this process remains to be elucidated. Here, using Drosophila larval sensory neurons, we show that a FMRP-associated miRNA, miR-276, functions in FMRP-mediated space filling dendrite morphogenesis. Using EGFP miRNA sensors, we demonstrate that FMRP likely acts by regulating miR-276a RNA targeting rather than by modulating miRNA levels. Supporting this conclusion, miR-276a co-immunoprecipitated with FMRP and this association was dependent on the FMRP KH domains. By testing putative targets of the FMRP-miR-276a regulatory axis, we identified nejire (nej) as a FMRP-associated mRNA and, using EGFP reporters, showed that the nej 3′ UTR is a target of miR-276a in vivo. Genetic analysis places nej downstream of the FMRP-miR-276a pathway in regulating dendrite patterning. Together, our findings support a model in which FMRP facilitates miR-276a-mediated control of nej for proper dendrite space filling morphology, and shed light on miRNA-dependent dendrite developmental pathology of Fragile X syndrome.

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“…Conventional PKC has been implicated in regulating the actin cytoskeleton. Therefore, we examined how the actin-based rhabdomeres might be affected using Actin-GFP that is expressed in R1-8 photoreceptors as the reporter (Roper et al, 2005;Shieh et al, 2021). Indeed, Pkc53E-RNAi results in missing rhabdomeres, which is accompanied by the abnormal arrangement of ommatidia clusters (Figure 2B).…”

Section: Downregulation Of Either Pkc53e or Eye-pkc Leads To Retinal Degeneration Via Two Gfp Reportersmentioning

confidence: 99%

Transmembrane Signaling Regulates the Remodeling of the Actin Cytoskeleton: Roles of PKC and Adducin

Shieh

Sun

Ferng

2021

Preprint

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Members of the conventional protein kinase C (cPKC) family are activated by both DAG and Ca2+ and have been implicated in the regulation of the actin cytoskeleton. Drosophila contains two cPKCs, Pkc53E (Pkc1) and eye-PKC (Pkc2); mutants missing each PKC lead to retinal degeneration. While eye-PKC is critical for the visual signaling, the role of Pkc53E is not known. We identified a photoreceptor-specific isoform of Pkc53E and show Pkc53E-RNAi negatively impacts the actin cytoskeleton of rhabdomeres. Interestingly, Pkc53E-RNAi enhances the degeneration of norpAP24 photoreceptors, suggesting Pkc53E could be activated independently of NorpA/PLCβ4. We further demonstrate that in norpAP24 photoreceptors Plc21C can be activated by Gq, which is responsible for the activation of Pkc53E. We explored whether Pkc53E regulates adducin in Drosophila photoreceptors. Adducin cross-links the actin cytoskeleton to the spectrin network, which is blunted by PKC phosphorylation. Importantly, we observed that phosphorylation of adducin was greatly reduced in a null allele of pkc53E. Downregulation of hts that encodes Drosophila adducin, exerts a more severe effect than Pkc53E-RNAi to impact the actin cytoskeleton. In contrast, overexpression of a mCherry tagged Add2, one of the three Drosophila adducin isoforms, led to the apical expansion of rhabdomeres with overgrowth of the actin cytoskeleton. This phenotype is likely caused by the dominant-negative activity of the tagged Add2 as it also was observed in α-spectrin-RNAi or β-spectrin-RNAi. Interestingly, downregulation of Pkc53E does not suppress the expansion of rhabdomeres during development, but negatively affects the appearance of rhabdomeres in adult photoreceptors. We conclude that Drosophila adducin has two distinct functions: in pupal photoreceptors, it regulates rhabdomere morphogenesis, which is independent of Pkc53E. In adult photoreceptors, it promotes the maintenance of the actin cytoskeleton, which is regulated by Pkc53E in response to the light-induced activation of the PLCβ activity.

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Exploring Excitotoxicity and Regulation of a Constitutively Active TRP Ca²⁺ Channel in Drosophila

Cited by 3 publications

References 61 publications

A conventional PKC critical for both the light-dependent and the light-independent regulation of the actin cytoskeleton in Drosophila photoreceptors

A conventional PKC critical for both the light-dependent and the light-independent regulation of the actin cytoskeleton in Drosophila photoreceptors

Drosophila FMRP controls miR-276-mediated regulation of nejire mRNA for space-filling dendrite development

Transmembrane Signaling Regulates the Remodeling of the Actin Cytoskeleton: Roles of PKC and Adducin

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Exploring Excitotoxicity and Regulation of a Constitutively Active TRP Ca2+ Channel in Drosophila

Cited by 3 publications

References 61 publications

A conventional PKC critical for both the light-dependent and the light-independent regulation of the actin cytoskeleton in Drosophila photoreceptors

A conventional PKC critical for both the light-dependent and the light-independent regulation of the actin cytoskeleton in Drosophila photoreceptors

Drosophila FMRP controls miR-276-mediated regulation of nejire mRNA for space-filling dendrite development

Transmembrane Signaling Regulates the Remodeling of the Actin Cytoskeleton: Roles of PKC and Adducin

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Exploring Excitotoxicity and Regulation of a Constitutively Active TRP Ca²⁺ Channel in Drosophila