Adenoviral expression of <scp>TDP</scp>‐43 and <scp>FUS</scp> genes and <scp>shRNAs</scp> for protein degradation pathways in rodent motoneurons in vitro and in vivo

Watabe, Kazuhiko; Akiyama, Keiko; Kawakami, Emiko; Ishii, Takemochi; Endo, Kentaro; Yanagisawa, Hiroko; Sango, Kazunori; Tsukamoto, Masami

doi:10.1111/neup.12058

Cited by 17 publications

(50 citation statements)

References 75 publications

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“…In the presence of retinoic acid, 1464R cells differentiate predominantly into TuJ1-positive neurons and, to a lesser extent, into GFAP-positive astrocytes or O4-positive oligodendrocytes. We have demonstrated cytoplasmic aggregate formation in these neurons and glial cells by co-transduction of adenoviruses expressing DsRed-tagged WT and CTF (208-414aa; 25kD) TDP-43 in the presence of proteasome inhibitor MG-132 [26]. In this study, we confirmed that these cytoplasmic TDP-43 aggregates were strongly immunoreactive for phosphorylated TDP-43 (Ser409/Ser410) and ubiquitin (Fig 1).…”

Section: Resultssupporting

confidence: 74%

“…In the present study we used a neural stem cell (NSC) line 1464R established from the adult rat brainstem [26] to examine aggregate formation of TDP-43. In the presence of retinoic acid, 1464R cells differentiate predominantly into TuJ1-positive neurons and, to a lesser extent, into GFAP-positive astrocytes or O4-positive oligodendrocytes.…”

Section: Resultsmentioning

confidence: 99%

“…Protein misfolding and aggregate deposition cause cellular dysfunction and ultimately lead to cell death [24,25]. We have previously demonstrated that proteasome inhibition enhanced the formation of adenovirus-transduced artificial TDP-43 cytoplasmic aggregates both in vitro and in vivo , suggesting that impairment of protein degradation pathways accelerates the formation of TDP-43-positive aggregates [26]. However, the molecular mechanisms underlying aggregate formation are largely unknown, and whether these aggregates play toxic or protective roles remains controversial [25,27].…”

Section: Introductionmentioning

confidence: 99%

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Formation and spreading of TDP-43 aggregates in cultured neuronal and glial cells demonstrated by time-lapse imaging

et al. 2017

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TAR DNA-binding protein 43 (TDP-43) is a main constituent of cytoplasmic aggregates in neuronal and glial cells in cases of amyotrophic lateral sclerosis and frontotemporal lobar degeneration. We have previously demonstrated that adenovirus-transduced artificial TDP-43 cytoplasmic aggregates formation is enhanced by proteasome inhibition in vitro and in vivo. However, the relationship between cytoplasmic aggregate formation and cell death remains unclear. In the present study, rat neural stem cell lines stably transfected with EGFP- or Sirius-expression vectors under the control of tubulin beta III, glial fibrillary acidic protein, or 2′,3′-cyclic nucleotide 3′-phosphodiesterase promoter were differentiated into neurons, astrocytes, and oligodendrocytes, respectively, in the presence of retinoic acid. The differentiated cells were then transduced with adenoviruses expressing DsRed-tagged human wild type and C-terminal fragment TDP-43 under the condition of proteasome inhibition. Time-lapse imaging analyses revealed growing cytoplasmic aggregates in the transduced neuronal and glial cells, followed by collapse of the cell. The aggregates remained insoluble in culture media, consisted of sarkosyl-insoluble granular materials, and contained phosphorylated TDP-43. Moreover, the released aggregates were incorporated into neighboring neuronal cells, suggesting cell-to-cell spreading. The present study provides a novel tool for analyzing the detailed molecular mechanisms of TDP-43 proteinopathy in vitro.

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Section: Resultssupporting

confidence: 74%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Formation and spreading of TDP-43 aggregates in cultured neuronal and glial cells demonstrated by time-lapse imaging

et al. 2017

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“…Mutant FUS may also inhibit the proteasome because raising cAMP levels with forskolin induces the clearance of insoluble and soluble fraction of FUS aggregates (Lokireddy et al, 2015). Furthermore, inhibition of the autophagy, proteasome or endosome/ESCRT systems with shRNAS targeting proteins involved in these pathways; ATG5, PMSC1 and VPS24; increased the localization of FUS within SG (Watabe et al, 2014). In addition, induction of autophagy using rapamycin reduced mutant FUS-positive SGs, neurite fragmentation and apoptosis in primary neurons overexpressing mutant FUS, under conditions of oxidative stress (Ryu et al, 2014).…”

Section: Fusmentioning

confidence: 99%

Protein Quality Control and the Amyotrophic Lateral Sclerosis/Frontotemporal Dementia Continuum

Shahheydari

Ragagnin

Walker

et al. 2017

Front. Mol. Neurosci.

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Protein homeostasis, or proteostasis, has an important regulatory role in cellular function. Protein quality control mechanisms, including protein folding and protein degradation processes, have a crucial function in post-mitotic neurons. Cellular protein quality control relies on multiple strategies, including molecular chaperones, autophagy, the ubiquitin proteasome system, endoplasmic reticulum (ER)-associated degradation (ERAD) and the formation of stress granules (SGs), to regulate proteostasis. Neurodegenerative diseases are characterized by the presence of misfolded protein aggregates, implying that protein quality control mechanisms are dysfunctional in these conditions. Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are neurodegenerative diseases that are now recognized to overlap clinically and pathologically, forming a continuous disease spectrum. In this review article, we detail the evidence for dysregulation of protein quality control mechanisms across the whole ALS-FTD continuum, by discussing the major proteins implicated in ALS and/or FTD. We also discuss possible ways in which protein quality mechanisms could be targeted therapeutically in these disorders and highlight promising protein quality control-based therapeutics for clinical trials.

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“…Motor neurons derived from mESC transfected with FUS adenoviral vectors displayed cytoplasmic aggregate formation. 48 Additionally, Wainger et al 26 found that motor neurons derived from iPSC of patients carrying mutations in FUS were hyperexcitable. 26 Sporadic disease iPSC-derived motor neurons from known genetic cases have revealed several insights in ALS pathology as we have described above.…”

Section: Fusmentioning

confidence: 99%

A perspective on stem cell modeling of amyotrophic lateral sclerosis

Boer

Eggan

2015

Cell Cycle

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Adenoviral expression of TDP‐43 and FUS genes and shRNAs for protein degradation pathways in rodent motoneurons in vitro and in vivo

Abstract: Formation of cytoplasmic aggregates in neuronal and glial cells is one of the pathological hallmarks of amyotrophic lateral sclerosis (ALS). Mutations in two genes

Cited by 17 publications

References 75 publications

Formation and spreading of TDP-43 aggregates in cultured neuronal and glial cells demonstrated by time-lapse imaging

Formation and spreading of TDP-43 aggregates in cultured neuronal and glial cells demonstrated by time-lapse imaging

Protein Quality Control and the Amyotrophic Lateral Sclerosis/Frontotemporal Dementia Continuum

A perspective on stem cell modeling of amyotrophic lateral sclerosis

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