An, H., Elvers, K.T., Gillespie, J.A. et al. (4 more authors) (2022) A toolkit for the identification of NEAT1_2/paraspeckle modulators. Nucleic Acids Research. ISSN 0305-1048
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.
Metadata
Authors/Creators: |
|
||||
---|---|---|---|---|---|
Copyright, Publisher and Additional Information: | © 2022 The Authors. This is an Open Access article distributed under the terms of the Creative Commons Attribution Licence (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. | ||||
Keywords: | Cell biology; Nucleic Acid Enzymology | ||||
Dates: |
|
||||
Institution: | The University of Sheffield | ||||
Academic Units: | The University of Sheffield > Faculty of Medicine, Dentistry and Health (Sheffield) > Department of Neuroscience (Sheffield) | ||||
Funding Information: |
|
||||
Depositing User: | Symplectic Sheffield | ||||
Date Deposited: | 24 Oct 2022 09:47 | ||||
Last Modified: | 24 Oct 2022 09:47 | ||||
Status: | Published online | ||||
Publisher: | Oxford University Press (OUP) | ||||
Refereed: | Yes | ||||
Identification Number: | https://doi.org/10.1093/nar/gkac771 | ||||
Related URLs: |