Reporters transiently transfected into mammalian cells are highly sensitive to translational repression induced by dsRNA expression.

In mammals, double-stranded RNA (dsRNA) can mediate sequence-specific RNA interference, activate sequence-independent interferon response, or undergo RNA editing by adenosine deaminases. We showed that long hairpin dsRNA expression had negligible effects on mammalian somatic cells--expressed dsRNA w...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Jana Nejepinska, Radek Malik, Susan Wagner, Petr Svoboda
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2014
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/f4804606d4fb43caa33a3f785c5e8e0e
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:f4804606d4fb43caa33a3f785c5e8e0e
record_format	dspace
spelling	oai:doaj.org-article:f4804606d4fb43caa33a3f785c5e8e0e2021-11-18T08:35:30ZReporters transiently transfected into mammalian cells are highly sensitive to translational repression induced by dsRNA expression.1932-620310.1371/journal.pone.0087517https://doaj.org/article/f4804606d4fb43caa33a3f785c5e8e0e2014-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/24475301/?tool=EBIhttps://doaj.org/toc/1932-6203In mammals, double-stranded RNA (dsRNA) can mediate sequence-specific RNA interference, activate sequence-independent interferon response, or undergo RNA editing by adenosine deaminases. We showed that long hairpin dsRNA expression had negligible effects on mammalian somatic cells--expressed dsRNA was slightly edited, poorly processed into siRNAs, and it did not activate the interferon response. At the same time, we noticed reduced reporter expression in transient co-transfections, which was presumably induced by expressed dsRNA. Since transient co-transfections are frequently used for studying gene function, we systematically explored the role of expressed dsRNA in this silencing phenomenon. We demonstrate that dsRNA expressed from transiently transfected plasmids strongly inhibits the expression of co-transfected reporter plasmids but not the expression of endogenous genes or reporters stably integrated in the genome. The inhibition is concentration-dependent, it is found in different cell types, and it is independent of transfection method and dsRNA sequence. The inhibition occurs at the level of translation and involves protein kinase R, which binds the expressed dsRNA. Thus, dsRNA expression represents a hidden danger in transient transfection experiments and must be taken into account during interpretation of experimental results.Jana NejepinskaRadek MalikSusan WagnerPetr SvobodaPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 9, Iss 1, p e87517 (2014)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Jana Nejepinska Radek Malik Susan Wagner Petr Svoboda Reporters transiently transfected into mammalian cells are highly sensitive to translational repression induced by dsRNA expression.
description	In mammals, double-stranded RNA (dsRNA) can mediate sequence-specific RNA interference, activate sequence-independent interferon response, or undergo RNA editing by adenosine deaminases. We showed that long hairpin dsRNA expression had negligible effects on mammalian somatic cells--expressed dsRNA was slightly edited, poorly processed into siRNAs, and it did not activate the interferon response. At the same time, we noticed reduced reporter expression in transient co-transfections, which was presumably induced by expressed dsRNA. Since transient co-transfections are frequently used for studying gene function, we systematically explored the role of expressed dsRNA in this silencing phenomenon. We demonstrate that dsRNA expressed from transiently transfected plasmids strongly inhibits the expression of co-transfected reporter plasmids but not the expression of endogenous genes or reporters stably integrated in the genome. The inhibition is concentration-dependent, it is found in different cell types, and it is independent of transfection method and dsRNA sequence. The inhibition occurs at the level of translation and involves protein kinase R, which binds the expressed dsRNA. Thus, dsRNA expression represents a hidden danger in transient transfection experiments and must be taken into account during interpretation of experimental results.
format	article
author	Jana Nejepinska Radek Malik Susan Wagner Petr Svoboda
author_facet	Jana Nejepinska Radek Malik Susan Wagner Petr Svoboda
author_sort	Jana Nejepinska
title	Reporters transiently transfected into mammalian cells are highly sensitive to translational repression induced by dsRNA expression.
title_short	Reporters transiently transfected into mammalian cells are highly sensitive to translational repression induced by dsRNA expression.
title_full	Reporters transiently transfected into mammalian cells are highly sensitive to translational repression induced by dsRNA expression.
title_fullStr	Reporters transiently transfected into mammalian cells are highly sensitive to translational repression induced by dsRNA expression.
title_full_unstemmed	Reporters transiently transfected into mammalian cells are highly sensitive to translational repression induced by dsRNA expression.
title_sort	reporters transiently transfected into mammalian cells are highly sensitive to translational repression induced by dsrna expression.
publisher	Public Library of Science (PLoS)
publishDate	2014
url	https://doaj.org/article/f4804606d4fb43caa33a3f785c5e8e0e
work_keys_str_mv	AT jananejepinska reporterstransientlytransfectedintomammaliancellsarehighlysensitivetotranslationalrepressioninducedbydsrnaexpression AT radekmalik reporterstransientlytransfectedintomammaliancellsarehighlysensitivetotranslationalrepressioninducedbydsrnaexpression AT susanwagner reporterstransientlytransfectedintomammaliancellsarehighlysensitivetotranslationalrepressioninducedbydsrnaexpression AT petrsvoboda reporterstransientlytransfectedintomammaliancellsarehighlysensitivetotranslationalrepressioninducedbydsrnaexpression
_version_	1718421597946118144

Reporters transiently transfected into mammalian cells are highly sensitive to translational repression induced by dsRNA expression.

Ejemplares similares