Unspliced Precursors of NMD-Sensitive β-Globin Transcripts Exhibit Decreased Steady-State Levels in Erythroid Cells
Autor(a) principal: | |
---|---|
Data de Publicação: | 2012 |
Outros Autores: | , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
Texto Completo: | http://hdl.handle.net/10400.18/891 |
Resumo: | Nonsense-mediated mRNA decay (NMD) is a quality control mechanism that detects and rapidly degrades mRNAs carrying premature translation-termination codons (PTCs). Mammalian NMD depends on both splicing and translation, and requires recognition of the premature stop codon by the cytoplasmic ribosomes. Surprisingly, some published data have suggested that nonsense codons may also affect the nuclear metabolism of the nonsense-mutated transcripts. To determine if nonsense codons could influence nuclear events, we have directly assessed the steady-state levels of the unspliced transcripts of wild-type and PTC-containing human β-globin genes stably transfected in mouse erythroleukemia (MEL) cells, after erythroid differentiation induction, or in HeLa cells. Our analyses by ribonuclease protection assays and reverse transcription-coupled quantitative PCR show that β-globin pre-mRNAs carrying NMD-competent PTCs, but not those containing a NMD-resistant PTC, exhibit a significant decrease in their steady-state levels relatively to the wild-type or to a missense-mutated β-globin pre-mRNA. On the contrary, in HeLa cells, human β-globin pre-mRNAs carrying NMD-competent PTCs accumulate at normal levels. Functional analyses of these pre-mRNAs in MEL cells demonstrate that their low steady-state levels do not reflect significantly lower pre-mRNA stabilities when compared to the normal control. Furthermore, our results also provide evidence that the relative splicing efficiencies of intron 1 and 2 are unaffected. This set of data highlights potential nuclear pathways that might be promoter- and/or cell line-specific, which recognize the NMD-sensitive transcripts as abnormal. These specialized nuclear pathway(s) may be superimposed on the general NMD mechanism. |
id |
RCAP_fe1fbbc44fc1f89d51c04e019e2bfa4d |
---|---|
oai_identifier_str |
oai:repositorio.insa.pt:10400.18/891 |
network_acronym_str |
RCAP |
network_name_str |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
repository_id_str |
7160 |
spelling |
Unspliced Precursors of NMD-Sensitive β-Globin Transcripts Exhibit Decreased Steady-State Levels in Erythroid CellsDoenças GenéticasGenómica Funcional e EstruturalNonsense-mediated mRNA decay (NMD) is a quality control mechanism that detects and rapidly degrades mRNAs carrying premature translation-termination codons (PTCs). Mammalian NMD depends on both splicing and translation, and requires recognition of the premature stop codon by the cytoplasmic ribosomes. Surprisingly, some published data have suggested that nonsense codons may also affect the nuclear metabolism of the nonsense-mutated transcripts. To determine if nonsense codons could influence nuclear events, we have directly assessed the steady-state levels of the unspliced transcripts of wild-type and PTC-containing human β-globin genes stably transfected in mouse erythroleukemia (MEL) cells, after erythroid differentiation induction, or in HeLa cells. Our analyses by ribonuclease protection assays and reverse transcription-coupled quantitative PCR show that β-globin pre-mRNAs carrying NMD-competent PTCs, but not those containing a NMD-resistant PTC, exhibit a significant decrease in their steady-state levels relatively to the wild-type or to a missense-mutated β-globin pre-mRNA. On the contrary, in HeLa cells, human β-globin pre-mRNAs carrying NMD-competent PTCs accumulate at normal levels. Functional analyses of these pre-mRNAs in MEL cells demonstrate that their low steady-state levels do not reflect significantly lower pre-mRNA stabilities when compared to the normal control. Furthermore, our results also provide evidence that the relative splicing efficiencies of intron 1 and 2 are unaffected. This set of data highlights potential nuclear pathways that might be promoter- and/or cell line-specific, which recognize the NMD-sensitive transcripts as abnormal. These specialized nuclear pathway(s) may be superimposed on the general NMD mechanism.Public Library of ScienceRepositório Científico do Instituto Nacional de SaúdeMorgado, AnaAlmeida, FátimaSilva, Ana LuísaRomão, Luísa2012-07-10T12:23:53Z2012-06-042012-06-04T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfhttp://hdl.handle.net/10400.18/891engPLoS One. 2012;7(6):e38505. Epub 2012 Jun 41932-6203ESSN: 1932-6203info:eu-repo/semantics/openAccessreponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãoinstacron:RCAAP2023-07-20T15:38:26Zoai:repositorio.insa.pt:10400.18/891Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-19T18:36:04.672209Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãofalse |
dc.title.none.fl_str_mv |
Unspliced Precursors of NMD-Sensitive β-Globin Transcripts Exhibit Decreased Steady-State Levels in Erythroid Cells |
title |
Unspliced Precursors of NMD-Sensitive β-Globin Transcripts Exhibit Decreased Steady-State Levels in Erythroid Cells |
spellingShingle |
Unspliced Precursors of NMD-Sensitive β-Globin Transcripts Exhibit Decreased Steady-State Levels in Erythroid Cells Morgado, Ana Doenças Genéticas Genómica Funcional e Estrutural |
title_short |
Unspliced Precursors of NMD-Sensitive β-Globin Transcripts Exhibit Decreased Steady-State Levels in Erythroid Cells |
title_full |
Unspliced Precursors of NMD-Sensitive β-Globin Transcripts Exhibit Decreased Steady-State Levels in Erythroid Cells |
title_fullStr |
Unspliced Precursors of NMD-Sensitive β-Globin Transcripts Exhibit Decreased Steady-State Levels in Erythroid Cells |
title_full_unstemmed |
Unspliced Precursors of NMD-Sensitive β-Globin Transcripts Exhibit Decreased Steady-State Levels in Erythroid Cells |
title_sort |
Unspliced Precursors of NMD-Sensitive β-Globin Transcripts Exhibit Decreased Steady-State Levels in Erythroid Cells |
author |
Morgado, Ana |
author_facet |
Morgado, Ana Almeida, Fátima Silva, Ana Luísa Romão, Luísa |
author_role |
author |
author2 |
Almeida, Fátima Silva, Ana Luísa Romão, Luísa |
author2_role |
author author author |
dc.contributor.none.fl_str_mv |
Repositório Científico do Instituto Nacional de Saúde |
dc.contributor.author.fl_str_mv |
Morgado, Ana Almeida, Fátima Silva, Ana Luísa Romão, Luísa |
dc.subject.por.fl_str_mv |
Doenças Genéticas Genómica Funcional e Estrutural |
topic |
Doenças Genéticas Genómica Funcional e Estrutural |
description |
Nonsense-mediated mRNA decay (NMD) is a quality control mechanism that detects and rapidly degrades mRNAs carrying premature translation-termination codons (PTCs). Mammalian NMD depends on both splicing and translation, and requires recognition of the premature stop codon by the cytoplasmic ribosomes. Surprisingly, some published data have suggested that nonsense codons may also affect the nuclear metabolism of the nonsense-mutated transcripts. To determine if nonsense codons could influence nuclear events, we have directly assessed the steady-state levels of the unspliced transcripts of wild-type and PTC-containing human β-globin genes stably transfected in mouse erythroleukemia (MEL) cells, after erythroid differentiation induction, or in HeLa cells. Our analyses by ribonuclease protection assays and reverse transcription-coupled quantitative PCR show that β-globin pre-mRNAs carrying NMD-competent PTCs, but not those containing a NMD-resistant PTC, exhibit a significant decrease in their steady-state levels relatively to the wild-type or to a missense-mutated β-globin pre-mRNA. On the contrary, in HeLa cells, human β-globin pre-mRNAs carrying NMD-competent PTCs accumulate at normal levels. Functional analyses of these pre-mRNAs in MEL cells demonstrate that their low steady-state levels do not reflect significantly lower pre-mRNA stabilities when compared to the normal control. Furthermore, our results also provide evidence that the relative splicing efficiencies of intron 1 and 2 are unaffected. This set of data highlights potential nuclear pathways that might be promoter- and/or cell line-specific, which recognize the NMD-sensitive transcripts as abnormal. These specialized nuclear pathway(s) may be superimposed on the general NMD mechanism. |
publishDate |
2012 |
dc.date.none.fl_str_mv |
2012-07-10T12:23:53Z 2012-06-04 2012-06-04T00:00:00Z |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/article |
format |
article |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
http://hdl.handle.net/10400.18/891 |
url |
http://hdl.handle.net/10400.18/891 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
PLoS One. 2012;7(6):e38505. Epub 2012 Jun 4 1932-6203 ESSN: 1932-6203 |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
application/pdf |
dc.publisher.none.fl_str_mv |
Public Library of Science |
publisher.none.fl_str_mv |
Public Library of Science |
dc.source.none.fl_str_mv |
reponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação instacron:RCAAP |
instname_str |
Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
instacron_str |
RCAAP |
institution |
RCAAP |
reponame_str |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
collection |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
repository.name.fl_str_mv |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
repository.mail.fl_str_mv |
|
_version_ |
1799132090335232000 |