Analysis of a nuclear localization signal in the p14 splicing factor in Trypanosoma cruzi

There are only a few reported nuclear localization signals (NLS) in trypanosomes despite intensive research on nuclear metabolic processes such as mRNA processing and transcription during the recent past. Moreover, there are only two reports for a monopartite (La protein) and bipartite (H2B histone,...

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Autores principales:	Westergaard, G.G., Bercovich, N., Reinert, M.D., Vazquez, M.P.
Formato:	JOUR
Materias:	Gateway vector NLS P14 PTREX Splicing factor Trypanosoma cell protein green fluorescent protein protein p14 protein SF3b155 unclassified drug nuclear localization signal nuclear protein protozoal protein recombinant protein RNA binding protein amino acid mapping protein RNA amino acid sequence article carboxy terminal sequence cell nucleus complex formation controlled study genetic transfection molecular recognition molecular size nonhuman nuclear import peptide mapping polymerase chain reaction protein domain protein motif RNA splicing Trypanosoma cruzi yeast enzymology genetics metabolism protein transport Amino Acid Motifs Green Fluorescent Proteins Nuclear Localization Signals Nuclear Proteins Protein Transport Protozoan Proteins Recombinant Proteins RNA-Binding Proteins Trypanosoma brucei
Acceso en línea:	http://hdl.handle.net/20.500.12110/paper_00207519_v40_n9_p1029_Westergaard
Aporte de:	Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) de Universidad de Buenos Aires

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spelling	todo:paper_00207519_v40_n9_p1029_Westergaard2023-10-03T14:18:35Z Analysis of a nuclear localization signal in the p14 splicing factor in Trypanosoma cruzi Westergaard, G.G. Bercovich, N. Reinert, M.D. Vazquez, M.P. Gateway vector NLS P14 PTREX Splicing factor Trypanosoma cell protein green fluorescent protein protein p14 protein SF3b155 unclassified drug nuclear localization signal nuclear protein protozoal protein recombinant protein RNA binding protein amino acid mapping protein RNA amino acid sequence article carboxy terminal sequence cell nucleus complex formation controlled study genetic transfection molecular recognition molecular size nonhuman nuclear import nuclear localization signal peptide mapping polymerase chain reaction protein domain protein motif RNA splicing Trypanosoma cruzi yeast enzymology genetics metabolism protein transport Trypanosoma cruzi Amino Acid Motifs Green Fluorescent Proteins Nuclear Localization Signals Nuclear Proteins Protein Transport Protozoan Proteins Recombinant Proteins RNA-Binding Proteins Trypanosoma cruzi Trypanosoma Trypanosoma brucei Trypanosoma cruzi Amino Acid Motifs Green Fluorescent Proteins Nuclear Localization Signals Nuclear Proteins Protein Transport Protozoan Proteins Recombinant Proteins RNA-Binding Proteins Trypanosoma cruzi There are only a few reported nuclear localization signals (NLS) in trypanosomes despite intensive research on nuclear metabolic processes such as mRNA processing and transcription during the recent past. Moreover, there are only two reports for a monopartite (La protein) and bipartite (H2B histone, ESAG8) NLS in Trypanosoma brucei. We decided to investigate a NLS in Trypanosoma cruzi by selecting p14, a small RNA recognition motif (RRM) containing protein involved in the splicing process in the nucleus. Its small size (117 amino acids), and an optimized streamlined workflow for analysis in T. cruzi, allowed us to define a region of basic amino acids (RRKRRR) located at the C-terminus that is necessary for nuclear localization. However, the NLS for p14 appeared to be more complex since the signature RRKRRR alone is necessary but not sufficient to direct heterologous proteins, such as GFP, to the nucleus. Since p14 interacts strongly with splicing factor SF3b155, a much larger protein, we designed a p14 variant unable to interact with it. The results allowed us to discard the notion that p14 is entering the nucleus, or is retained within, as the sole consequence of being part of a larger complex. Extensive mapping showed that all of the information for nuclear import resides within the small p14 protein in a bipartite NLS composed of the signature RRKRRR and a region of the RRM domain. Thus, NLS definition in T. cruzi is more complex than previously described. © 2010 Australian Society for Parasitology Inc. Fil:Westergaard, G.G. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Bercovich, N. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Vazquez, M.P. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. JOUR info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by/2.5/ar http://hdl.handle.net/20.500.12110/paper_00207519_v40_n9_p1029_Westergaard
institution	Universidad de Buenos Aires
institution_str	I-28
repository_str	R-134
collection	Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA)
topic	Gateway vector NLS P14 PTREX Splicing factor Trypanosoma cell protein green fluorescent protein protein p14 protein SF3b155 unclassified drug nuclear localization signal nuclear protein protozoal protein recombinant protein RNA binding protein amino acid mapping protein RNA amino acid sequence article carboxy terminal sequence cell nucleus complex formation controlled study genetic transfection molecular recognition molecular size nonhuman nuclear import nuclear localization signal peptide mapping polymerase chain reaction protein domain protein motif RNA splicing Trypanosoma cruzi yeast enzymology genetics metabolism protein transport Trypanosoma cruzi Amino Acid Motifs Green Fluorescent Proteins Nuclear Localization Signals Nuclear Proteins Protein Transport Protozoan Proteins Recombinant Proteins RNA-Binding Proteins Trypanosoma cruzi Trypanosoma Trypanosoma brucei Trypanosoma cruzi Amino Acid Motifs Green Fluorescent Proteins Nuclear Localization Signals Nuclear Proteins Protein Transport Protozoan Proteins Recombinant Proteins RNA-Binding Proteins Trypanosoma cruzi
spellingShingle	Gateway vector NLS P14 PTREX Splicing factor Trypanosoma cell protein green fluorescent protein protein p14 protein SF3b155 unclassified drug nuclear localization signal nuclear protein protozoal protein recombinant protein RNA binding protein amino acid mapping protein RNA amino acid sequence article carboxy terminal sequence cell nucleus complex formation controlled study genetic transfection molecular recognition molecular size nonhuman nuclear import nuclear localization signal peptide mapping polymerase chain reaction protein domain protein motif RNA splicing Trypanosoma cruzi yeast enzymology genetics metabolism protein transport Trypanosoma cruzi Amino Acid Motifs Green Fluorescent Proteins Nuclear Localization Signals Nuclear Proteins Protein Transport Protozoan Proteins Recombinant Proteins RNA-Binding Proteins Trypanosoma cruzi Trypanosoma Trypanosoma brucei Trypanosoma cruzi Amino Acid Motifs Green Fluorescent Proteins Nuclear Localization Signals Nuclear Proteins Protein Transport Protozoan Proteins Recombinant Proteins RNA-Binding Proteins Trypanosoma cruzi Westergaard, G.G. Bercovich, N. Reinert, M.D. Vazquez, M.P. Analysis of a nuclear localization signal in the p14 splicing factor in Trypanosoma cruzi
topic_facet	Gateway vector NLS P14 PTREX Splicing factor Trypanosoma cell protein green fluorescent protein protein p14 protein SF3b155 unclassified drug nuclear localization signal nuclear protein protozoal protein recombinant protein RNA binding protein amino acid mapping protein RNA amino acid sequence article carboxy terminal sequence cell nucleus complex formation controlled study genetic transfection molecular recognition molecular size nonhuman nuclear import nuclear localization signal peptide mapping polymerase chain reaction protein domain protein motif RNA splicing Trypanosoma cruzi yeast enzymology genetics metabolism protein transport Trypanosoma cruzi Amino Acid Motifs Green Fluorescent Proteins Nuclear Localization Signals Nuclear Proteins Protein Transport Protozoan Proteins Recombinant Proteins RNA-Binding Proteins Trypanosoma cruzi Trypanosoma Trypanosoma brucei Trypanosoma cruzi Amino Acid Motifs Green Fluorescent Proteins Nuclear Localization Signals Nuclear Proteins Protein Transport Protozoan Proteins Recombinant Proteins RNA-Binding Proteins Trypanosoma cruzi
description	There are only a few reported nuclear localization signals (NLS) in trypanosomes despite intensive research on nuclear metabolic processes such as mRNA processing and transcription during the recent past. Moreover, there are only two reports for a monopartite (La protein) and bipartite (H2B histone, ESAG8) NLS in Trypanosoma brucei. We decided to investigate a NLS in Trypanosoma cruzi by selecting p14, a small RNA recognition motif (RRM) containing protein involved in the splicing process in the nucleus. Its small size (117 amino acids), and an optimized streamlined workflow for analysis in T. cruzi, allowed us to define a region of basic amino acids (RRKRRR) located at the C-terminus that is necessary for nuclear localization. However, the NLS for p14 appeared to be more complex since the signature RRKRRR alone is necessary but not sufficient to direct heterologous proteins, such as GFP, to the nucleus. Since p14 interacts strongly with splicing factor SF3b155, a much larger protein, we designed a p14 variant unable to interact with it. The results allowed us to discard the notion that p14 is entering the nucleus, or is retained within, as the sole consequence of being part of a larger complex. Extensive mapping showed that all of the information for nuclear import resides within the small p14 protein in a bipartite NLS composed of the signature RRKRRR and a region of the RRM domain. Thus, NLS definition in T. cruzi is more complex than previously described. © 2010 Australian Society for Parasitology Inc.
format	JOUR
author	Westergaard, G.G. Bercovich, N. Reinert, M.D. Vazquez, M.P.
author_facet	Westergaard, G.G. Bercovich, N. Reinert, M.D. Vazquez, M.P.
author_sort	Westergaard, G.G.
title	Analysis of a nuclear localization signal in the p14 splicing factor in Trypanosoma cruzi
title_short	Analysis of a nuclear localization signal in the p14 splicing factor in Trypanosoma cruzi
title_full	Analysis of a nuclear localization signal in the p14 splicing factor in Trypanosoma cruzi
title_fullStr	Analysis of a nuclear localization signal in the p14 splicing factor in Trypanosoma cruzi
title_full_unstemmed	Analysis of a nuclear localization signal in the p14 splicing factor in Trypanosoma cruzi
title_sort	analysis of a nuclear localization signal in the p14 splicing factor in trypanosoma cruzi
url	http://hdl.handle.net/20.500.12110/paper_00207519_v40_n9_p1029_Westergaard
work_keys_str_mv	AT westergaardgg analysisofanuclearlocalizationsignalinthep14splicingfactorintrypanosomacruzi AT bercovichn analysisofanuclearlocalizationsignalinthep14splicingfactorintrypanosomacruzi AT reinertmd analysisofanuclearlocalizationsignalinthep14splicingfactorintrypanosomacruzi AT vazquezmp analysisofanuclearlocalizationsignalinthep14splicingfactorintrypanosomacruzi
_version_	1807322631278428160

Analysis of a nuclear localization signal in the p14 splicing factor in Trypanosoma cruzi

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