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A toolkit for the identification of NEAT1_2/paraspeckle modulators

An, Haiyan, Elvers, Karen T., Gillespie, Jason A., Jones, Kimberley, Atack, John R. ORCID: https://orcid.org/0000-0002-3410-791X, Grubisha, Olivera ORCID: https://orcid.org/0000-0003-1067-1230 and Shelkovnikova, Tatyana A. ORCID: https://orcid.org/0000-0003-1367-5309 2022. A toolkit for the identification of NEAT1_2/paraspeckle modulators. Nucleic Acids Research 50 (20) , e119. 10.1093/nar/gkac771

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Abstract

Paraspeckles are ribonucleoprotein granules assembled by NEAT1_2 lncRNA, an isoform of Nuclear Paraspeckle Assembly Transcript 1 (NEAT1). Dysregulation of NEAT1_2/paraspeckles has been linked to multiple human diseases making them an attractive drug target. However currently NEAT1_2/paraspeckle-focused translational research and drug discovery are hindered by a limited toolkit. To fill this gap, we developed and validated a set of tools for the identification of NEAT1_2 binders and modulators comprised of biochemical and cell-based assays. The NEAT1_2 triple helix stability element was utilized as the target in the biochemical assays, and the cellular assay (‘ParaQuant’) was based on high-content imaging of NEAT1_2 in fixed cells. As a proof of principle, these assays were used to screen a 1,200-compound FDA-approved drug library and a 170-compound kinase inhibitor library and to confirm the screening hits. The assays are simple to establish, use only commercially-available reagents and are scalable for higher throughput. In particular, ParaQuant is a cost-efficient assay suitable for any cells growing in adherent culture and amenable to multiplexing. Using ParaQuant, we identified dual PI3K/mTOR inhibitors as potent negative modulators of paraspeckles. The tools we describe herein should boost paraspeckle studies and help guide the search, validation and optimization of NEAT1_2/paraspeckle-targeted small molecules.

Item Type: Article
Date Type: Publication
Status: Published
Schools: Biosciences
Publisher: Oxford University Press
ISSN: 0305-1048
Date of First Compliant Deposit: 28 October 2022
Date of Acceptance: 29 August 2022
Last Modified: 12 Jan 2023 12:02
URI: https://orca.cardiff.ac.uk/id/eprint/153837

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