Nuclear Localization of Huntingtin mRNA Is Specific to Cells of Neuronal Origin

Didiot, Marie-Cécile; Ferguson, Chantal; Ly, Socheata; Coles, Andrew H.; Smith, Abigail O.; Bicknell, Alicia A.; Hall, Lauren M; Sapp, Ellen; Echeverria, Dimas; Pai, Athma A.; DiFiglia, Marian; Moore, Melissa J.; Hayward, Lawrence J.; Aronin, Neil; Khvorova, Anastasia

doi:10.1016/j.celrep.2018.07.106

Cited by 36 publications

(33 citation statements)

References 24 publications

(31 reference statements)

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“…The extent of HTT protein silencing was always greater than Htt mRNA (> 99% vs ~70%) ( Fig. 2c,d) because a fraction of Htt mRNA localizes to the nucleus, making it more resistant to RNAi-based silencing 29 . More than 80% silencing of HTT protein was maintained at four months in all brain regions analyzed ( Fig.…”

Section: The Efficacy Of High-dose Di-sirna Exceeds Six Monthsmentioning

confidence: 96%

A divalent siRNA chemical scaffold for potent and sustained modulation of gene expression throughout the central nervous system

et al. 2019

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Section: The Efficacy Of High-dose Di-sirna Exceeds Six Monthsmentioning

confidence: 96%

A divalent siRNA chemical scaffold for potent and sustained modulation of gene expression throughout the central nervous system

et al. 2019

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“…Incomplete knockdown can be explained by a variety of reasons, including the presence of mRNA isoforms detectable by the qPCR probes that are not affected by the sdRNA, or by the nuclear localization of mRNA. 85 However, it is not always advantageous to have complete knockdown. We have consistently observed that small changes in USP10 expression relate to large phenotypic changes in cells.…”

Section: Discussionmentioning

confidence: 99%

USP10 Targeted Self-Deliverable siRNA to Prevent Scarring in the Cornea

Boumil

Castro

Phillips

et al. 2020

Molecular Therapy - Nucleic Acids

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Ocular scarring after surgery, trauma, or infection leads to vision loss. The transparent cornea is an excellent model system to test anti-scarring therapies. Cholesterol-conjugated fully modified asymmetric small interfering RNAs (siRNAs) (selfdeliverable siRNAs [sdRNAs]) are a novel modality for in vivo gene knockdown, transfecting cells and tissues without any additional formulations. Myofibroblasts are a main contributor to scarring and fibrosis. a v integrins play a central role in myofibroblast pathological adhesion, overcontraction, and transforming growth factor b (TGF-b) activation. Previously, we demonstrated that a v integrins are protected from intracellular degradation after wounding by upregulation of the deubiquitinase (DUB) ubiquitin-specific protease 10 (USP10), leading to integrin cell surface accumulation. In this study, we tested whether knockdown of USP10 with a USP10-targeting sdRNA (termed US09) will reduce scarring after wounding a rabbit cornea in vivo. The wounded corneal stroma was treated once with US09 or non-targeting control (NTC) sdRNA. At 6 weeks US09 treatment resulted in faster wound closure, limited scarring, and suppression of fibrotic markers and immune response. Specifically, fibronectin-extra domain A (EDA), collagen III, and a-smooth muscle actin (p < 0.05), CD45 + cell infiltration (p < 0.01), and apoptosis at 24 (p < 0.01) and 48 h (p < 0.05) were reduced post-wounding. Corneal thickness and cell proliferation were restored to unwounded parameters. Targeting the DUB, USP10 is a novel strategy to reduce scarring. This study indicates that ubiquitin-mediated pathways should be considered in the pathogenesis of fibrotic healing.

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“…We have previously shown that cellular origin has an impact on subcellular localization of huntingtin mRNA in both wild-type mice 23 and mutant models of Huntington's disease models. We found that in cells of neuronal origin, a larger portion of Htt mRNA was localized to the nucleus, corresponding to a lower degree of observable mRNA silencing.…”

Section: Cell Type Impacts Efficacy Of Sirnas Targeting Apoementioning

confidence: 99%

“…We found that in cells of neuronal origin, a larger portion of Htt mRNA was localized to the nucleus, corresponding to a lower degree of observable mRNA silencing. 23 Thus, as a next step, we evaluated ApoE intracellular localization in different cell types.…”

Section: Cell Type Impacts Efficacy Of Sirnas Targeting Apoementioning

confidence: 99%

Cell Type Impacts Accessibility of mRNA to Silencing by RNA Interference

Ferguson

Echeverria

Hassler

et al. 2020

Molecular Therapy - Nucleic Acids

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RNA interference (RNAi) is a potent mechanism that silences mRNA and protein expression in all cells and tissue types. RNAi is known to exert many of its functional effects in the cytoplasm, and thus, the cellular localization of target mRNA may impact observed potency. Here, we demonstrate that cell identity has a profound impact on accessibility of apolipoprotein E (ApoE) mRNA to RNAi. We show that, whereas both neuronal and glial cell lines express detectable ApoE mRNA, in neuronal cells, ApoE mRNA is not targetable by RNAi. Screening of a panel of thirty-five chemically modified small interfering RNAs (siRNAs) did not produce a single hit in a neuronal cell line, whereas up to fifteen compounds showed strong efficacy in glial cells. Further investigation of the cellular localization of ApoE mRNA demonstrates that ApoE mRNA is partially spliced and preferentially localized to the nucleus ($80%) in neuronal cells, whereas more than 90% of ApoE mRNA is cytoplasmic in glial cells. Such an inconsistency in intracellular localization and splicing might provide an explanation for functional differences in RNAi compounds. Thus, cellular origin might have an impact on accessibility of mRNA to RNAi and should be taken into account during the screening process.

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Nuclear Localization of Huntingtin mRNA Is Specific to Cells of Neuronal Origin

Cited by 36 publications

References 24 publications

A divalent siRNA chemical scaffold for potent and sustained modulation of gene expression throughout the central nervous system

A divalent siRNA chemical scaffold for potent and sustained modulation of gene expression throughout the central nervous system

USP10 Targeted Self-Deliverable siRNA to Prevent Scarring in the Cornea

Cell Type Impacts Accessibility of mRNA to Silencing by RNA Interference

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