4.7 Article

Artificial miRNAs targeting CAG repeat expansion in ORFs cause rapid deadenylation and translation inhibition of mutant transcripts

Journal

CELLULAR AND MOLECULAR LIFE SCIENCES
Volume 78, Issue 4, Pages 1577-1596

Publisher

SPRINGER BASEL AG
DOI: 10.1007/s00018-020-03596-7

Keywords

miRNA; CAG repeats; Polyglutamine diseases; Huntington's disease; Translational inhibition

Funding

  1. National Science Centre [2014/15/B/NZ1/01880, 2015/17/D/NZ5/03443, 2015/19/B/NZ2/02453, 2015/17/N/NZ2/01916]
  2. Polish Ministry of Science and Higher Education [DI 2011 0278 41, 01/KNOW2/2014]

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Polyglutamine diseases, such as Huntington's disease, are incurable neurological disorders caused by CAG repeat expansion in specific genes. The use of CAG repeat-targeting artificial miRNAs shows promise in selectively decreasing mutant protein levels. Research suggests that the efficiency and selectivity of art-miRNAs are influenced by the localization and sequence context of CAG repeats in the transcript. Additionally, the downregulation of mutant polyQ proteins involves rapid activation of translational repression and transcript deadenylation, with the slicer activity of AGO2 being dispensable in this process.
Polyglutamine (polyQ) diseases are incurable neurological disorders caused by CAG repeat expansion in the open reading frames (ORFs) of specific genes. This type of mutation in theHTTgene is responsible for Huntington's disease (HD). CAG repeat-targeting artificial miRNAs (art-miRNAs) were shown as attractive therapeutic approach for polyQ disorders as they caused allele-selective decrease in the level of mutant proteins. Here, using polyQ disease models, we aimed to demonstrate how miRNA-based gene expression regulation is dependent on target sequence features. We show that the silencing efficiency and selectivity of art-miRNAs is influenced by the localization of the CAG repeat tract within transcript and the specific sequence context. Furthermore, we aimed to reveal the events leading to downregulation of mutant polyQ proteins and found very rapid activation of translational repression andHTTtranscript deadenylation. Slicer-activity of AGO2 was dispensable in this process, as determined inAGO2knockout cells generated with CRISPR-Cas9 technology. We also showed highly allele-selective downregulation of huntingtin in human HD neural progenitors (NPs). Taken together, art-miRNA activity may serve as a model of the cooperative activity and targeting of ORF regions by endogenous miRNAs.

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