<named-content content-type="genus-species">Trypanosoma brucei</named-content> RRM1 Is a Nuclear RNA-Binding Protein and Modulator of Chromatin Structure

ABSTRACT TbRRM1 of Trypanosoma brucei is a nucleoprotein that was previously identified in a search for splicing factors in T. brucei. We show that TbRRM1 associates with mRNAs and with the auxiliary splicing factor polypyrimidine tract-binding protein 2, but not with components of the core spliceos...

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Autores principales:	Arunasalam Naguleswaran, Kapila Gunasekera, Bernd Schimanski, Manfred Heller, Andrew Hemphill, Torsten Ochsenreiter, Isabel Roditi
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2015
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/da0b481d64e14f18bf26068751de0ed7
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spelling	oai:doaj.org-article:da0b481d64e14f18bf26068751de0ed72021-11-15T15:41:33Z<named-content content-type="genus-species">Trypanosoma brucei</named-content> RRM1 Is a Nuclear RNA-Binding Protein and Modulator of Chromatin Structure10.1128/mBio.00114-152150-7511https://doaj.org/article/da0b481d64e14f18bf26068751de0ed72015-05-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00114-15https://doaj.org/toc/2150-7511ABSTRACT TbRRM1 of Trypanosoma brucei is a nucleoprotein that was previously identified in a search for splicing factors in T. brucei. We show that TbRRM1 associates with mRNAs and with the auxiliary splicing factor polypyrimidine tract-binding protein 2, but not with components of the core spliceosome. TbRRM1 also interacts with several retrotransposon hot spot (RHS) proteins and histones. RNA immunoprecipitation of a tagged form of TbRRM1 from procyclic (insect) form trypanosomes identified ca. 1,500 transcripts that were enriched and 3,000 transcripts that were underrepresented compared to cellular mRNA. Enriched transcripts encoded RNA-binding proteins, including TbRRM1 itself, several RHS transcripts, mRNAs with long coding regions, and a high proportion of stage-regulated mRNAs that are more highly expressed in bloodstream forms. Transcripts encoding ribosomal proteins, other factors involved in translation, and procyclic-specific transcripts were underrepresented. Knockdown of TbRRM1 by RNA interference caused widespread changes in mRNA abundance, but these changes did not correlate with the binding of the protein to transcripts, and most splice sites were unchanged, negating a general role for TbRRM1 in splice site selection. When changes in mRNA abundance were mapped across the genome, regions with many downregulated mRNAs were identified. Two regions were analyzed by chromatin immunoprecipitation, both of which exhibited increases in nucleosome occupancy upon TbRRM1 depletion. In addition, subjecting cells to heat shock resulted in translocation of TbRRM1 to the cytoplasm and compaction of chromatin, consistent with a second role for TbRRM1 in modulating chromatin structure. IMPORTANCE Trypanosoma brucei, the parasite that causes human sleeping sickness, is transmitted by tsetse flies. The parasite progresses through different life cycle stages in its two hosts, altering its pattern of gene expression in the process. In trypanosomes, protein-coding genes are organized as polycistronic units that are processed into monocistronic mRNAs. Since genes in the same unit can be regulated independently of each other, it is believed that gene regulation is essentially posttranscriptional. In this study, we investigated the role of a nuclear RNA-binding protein, TbRRM1, in the insect stage of the parasite. We found that TbRRM1 binds nuclear mRNAs and also affects chromatin status. Reduction of nuclear TbRRM1 by RNA interference or heat shock resulted in chromatin compaction. We propose that TbRRM1 regulates RNA polymerase II-driven gene expression both cotranscriptionally, by facilitating transcription and efficient splicing, and posttranscriptionally, via its interaction with nuclear mRNAs.Arunasalam NaguleswaranKapila GunasekeraBernd SchimanskiManfred HellerAndrew HemphillTorsten OchsenreiterIsabel RoditiAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 6, Iss 2 (2015)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Arunasalam Naguleswaran Kapila Gunasekera Bernd Schimanski Manfred Heller Andrew Hemphill Torsten Ochsenreiter Isabel Roditi <named-content content-type="genus-species">Trypanosoma brucei</named-content> RRM1 Is a Nuclear RNA-Binding Protein and Modulator of Chromatin Structure
description	ABSTRACT TbRRM1 of Trypanosoma brucei is a nucleoprotein that was previously identified in a search for splicing factors in T. brucei. We show that TbRRM1 associates with mRNAs and with the auxiliary splicing factor polypyrimidine tract-binding protein 2, but not with components of the core spliceosome. TbRRM1 also interacts with several retrotransposon hot spot (RHS) proteins and histones. RNA immunoprecipitation of a tagged form of TbRRM1 from procyclic (insect) form trypanosomes identified ca. 1,500 transcripts that were enriched and 3,000 transcripts that were underrepresented compared to cellular mRNA. Enriched transcripts encoded RNA-binding proteins, including TbRRM1 itself, several RHS transcripts, mRNAs with long coding regions, and a high proportion of stage-regulated mRNAs that are more highly expressed in bloodstream forms. Transcripts encoding ribosomal proteins, other factors involved in translation, and procyclic-specific transcripts were underrepresented. Knockdown of TbRRM1 by RNA interference caused widespread changes in mRNA abundance, but these changes did not correlate with the binding of the protein to transcripts, and most splice sites were unchanged, negating a general role for TbRRM1 in splice site selection. When changes in mRNA abundance were mapped across the genome, regions with many downregulated mRNAs were identified. Two regions were analyzed by chromatin immunoprecipitation, both of which exhibited increases in nucleosome occupancy upon TbRRM1 depletion. In addition, subjecting cells to heat shock resulted in translocation of TbRRM1 to the cytoplasm and compaction of chromatin, consistent with a second role for TbRRM1 in modulating chromatin structure. IMPORTANCE Trypanosoma brucei, the parasite that causes human sleeping sickness, is transmitted by tsetse flies. The parasite progresses through different life cycle stages in its two hosts, altering its pattern of gene expression in the process. In trypanosomes, protein-coding genes are organized as polycistronic units that are processed into monocistronic mRNAs. Since genes in the same unit can be regulated independently of each other, it is believed that gene regulation is essentially posttranscriptional. In this study, we investigated the role of a nuclear RNA-binding protein, TbRRM1, in the insect stage of the parasite. We found that TbRRM1 binds nuclear mRNAs and also affects chromatin status. Reduction of nuclear TbRRM1 by RNA interference or heat shock resulted in chromatin compaction. We propose that TbRRM1 regulates RNA polymerase II-driven gene expression both cotranscriptionally, by facilitating transcription and efficient splicing, and posttranscriptionally, via its interaction with nuclear mRNAs.
format	article
author	Arunasalam Naguleswaran Kapila Gunasekera Bernd Schimanski Manfred Heller Andrew Hemphill Torsten Ochsenreiter Isabel Roditi
author_facet	Arunasalam Naguleswaran Kapila Gunasekera Bernd Schimanski Manfred Heller Andrew Hemphill Torsten Ochsenreiter Isabel Roditi
author_sort	Arunasalam Naguleswaran
title	<named-content content-type="genus-species">Trypanosoma brucei</named-content> RRM1 Is a Nuclear RNA-Binding Protein and Modulator of Chromatin Structure
title_short	<named-content content-type="genus-species">Trypanosoma brucei</named-content> RRM1 Is a Nuclear RNA-Binding Protein and Modulator of Chromatin Structure
title_full	<named-content content-type="genus-species">Trypanosoma brucei</named-content> RRM1 Is a Nuclear RNA-Binding Protein and Modulator of Chromatin Structure
title_fullStr	<named-content content-type="genus-species">Trypanosoma brucei</named-content> RRM1 Is a Nuclear RNA-Binding Protein and Modulator of Chromatin Structure
title_full_unstemmed	<named-content content-type="genus-species">Trypanosoma brucei</named-content> RRM1 Is a Nuclear RNA-Binding Protein and Modulator of Chromatin Structure
title_sort	<named-content content-type="genus-species">trypanosoma brucei</named-content> rrm1 is a nuclear rna-binding protein and modulator of chromatin structure
publisher	American Society for Microbiology
publishDate	2015
url	https://doaj.org/article/da0b481d64e14f18bf26068751de0ed7
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<named-content content-type="genus-species">Trypanosoma brucei</named-content> RRM1 Is a Nuclear RNA-Binding Protein and Modulator of Chromatin Structure

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