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Nonsense-mediated decay as a terminating mechanism for antisense oligonucleotides
Antisense oligonucleotides (ASOs) are synthetic oligonucleotides that alter expression of disease-associated transcripts via Watson–Crick hybridization. ASOs that function through RNase H or the RNA-induced silencing complex (RISC) result in enzymatic degradation of target RNA. ASOs designed to ster...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Oxford University Press
2014
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4027159/ https://www.ncbi.nlm.nih.gov/pubmed/24589581 http://dx.doi.org/10.1093/nar/gku184 |
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author | Ward, Amanda J. Norrbom, Michaela Chun, Seung Bennett, C. Frank Rigo, Frank |
author_facet | Ward, Amanda J. Norrbom, Michaela Chun, Seung Bennett, C. Frank Rigo, Frank |
author_sort | Ward, Amanda J. |
collection | PubMed |
description | Antisense oligonucleotides (ASOs) are synthetic oligonucleotides that alter expression of disease-associated transcripts via Watson–Crick hybridization. ASOs that function through RNase H or the RNA-induced silencing complex (RISC) result in enzymatic degradation of target RNA. ASOs designed to sterically block access of proteins to the RNA modulate mRNA metabolism but do not typically cause degradation. Here, we rationally design steric blocking ASOs to promote mRNA reduction and characterize the terminating mechanism. Transfection of ASOs complementary to constitutive exons in STAT3 and Sod1 results in greater than 70% reduction of mRNA and protein. The ASOs promote aberrant exon skipping and generation of premature termination codon (PTC)-containing mRNAs. We inhibit the nonsense-mediated mRNA decay (NMD) pathway and show that the PTC-containing mRNAs are recognized by the UPF1 ATPase, cleaved by the SMG6 endonuclease and degraded by the XRN1 cytoplasmic exonuclease. NMD surveillance, however, does not entirely explain the mechanism of decreased STAT3 expression. In addition to exon skipping, ASO treatment causes intron retention and reduction of chromatin-associated STAT3 mRNA. The application of steric blocking ASOs to promote RNA degradation allows one to explore more nucleotide modifications than tolerated by RNase H or RISC-dependent ASOs, with the goal of improving ASO drug properties. |
format | Online Article Text |
id | pubmed-4027159 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2014 |
publisher | Oxford University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-40271592014-05-28 Nonsense-mediated decay as a terminating mechanism for antisense oligonucleotides Ward, Amanda J. Norrbom, Michaela Chun, Seung Bennett, C. Frank Rigo, Frank Nucleic Acids Res RNA Antisense oligonucleotides (ASOs) are synthetic oligonucleotides that alter expression of disease-associated transcripts via Watson–Crick hybridization. ASOs that function through RNase H or the RNA-induced silencing complex (RISC) result in enzymatic degradation of target RNA. ASOs designed to sterically block access of proteins to the RNA modulate mRNA metabolism but do not typically cause degradation. Here, we rationally design steric blocking ASOs to promote mRNA reduction and characterize the terminating mechanism. Transfection of ASOs complementary to constitutive exons in STAT3 and Sod1 results in greater than 70% reduction of mRNA and protein. The ASOs promote aberrant exon skipping and generation of premature termination codon (PTC)-containing mRNAs. We inhibit the nonsense-mediated mRNA decay (NMD) pathway and show that the PTC-containing mRNAs are recognized by the UPF1 ATPase, cleaved by the SMG6 endonuclease and degraded by the XRN1 cytoplasmic exonuclease. NMD surveillance, however, does not entirely explain the mechanism of decreased STAT3 expression. In addition to exon skipping, ASO treatment causes intron retention and reduction of chromatin-associated STAT3 mRNA. The application of steric blocking ASOs to promote RNA degradation allows one to explore more nucleotide modifications than tolerated by RNase H or RISC-dependent ASOs, with the goal of improving ASO drug properties. Oxford University Press 2014-05-01 2014-04-03 /pmc/articles/PMC4027159/ /pubmed/24589581 http://dx.doi.org/10.1093/nar/gku184 Text en © The Author(s) 2014. Published by Oxford University Press [on behalf of Nucleic Acids Research]. http://creativecommons.org/licenses/by/3.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | RNA Ward, Amanda J. Norrbom, Michaela Chun, Seung Bennett, C. Frank Rigo, Frank Nonsense-mediated decay as a terminating mechanism for antisense oligonucleotides |
title | Nonsense-mediated decay as a terminating mechanism for antisense oligonucleotides |
title_full | Nonsense-mediated decay as a terminating mechanism for antisense oligonucleotides |
title_fullStr | Nonsense-mediated decay as a terminating mechanism for antisense oligonucleotides |
title_full_unstemmed | Nonsense-mediated decay as a terminating mechanism for antisense oligonucleotides |
title_short | Nonsense-mediated decay as a terminating mechanism for antisense oligonucleotides |
title_sort | nonsense-mediated decay as a terminating mechanism for antisense oligonucleotides |
topic | RNA |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4027159/ https://www.ncbi.nlm.nih.gov/pubmed/24589581 http://dx.doi.org/10.1093/nar/gku184 |
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