Influence of Polymerase II Processivity on Alternative Splicing Depends on Splice Site Strength

Transcription and pre-mRNA splicing are coordinated temporally and spatially, and both processes can influence each other. In particular, control of transcriptional elongation by RNA polymerase II has proved to be important for alternative splicing regulation. In this report we demonstrate that the...

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Autores principales:	Nogués, G., Muñoz, M.J., Kornblihtt, A.R.
Formato:	JOUR
Materias:	Cell culture Enzyme inhibition Genes Mutagenesis RNA Splice site strength Enzymes elongation factor fibronectin fibronectin extra domain I messenger RNA polypyrimidine tract binding protein RNA polymerase II unclassified drug article controlled study enzyme activation enzyme inhibition exon gene mutation human human cell molecular recognition nonhuman point mutation priority journal regulatory mechanism RNA splicing silencer element transcription regulation Alternative Splicing Binding Sites Cell Line Cell Line, Tumor Exons Herpes Simplex Virus Protein Vmw65 Humans Models, Biological Mutation Phosphorylation Plasmids Point Mutation Polymorphism, Genetic Protein Structure, Tertiary Pyrimidines RNA Polymerase II RNA Splicing RNA, Messenger Time Factors Transcription, Genetic Transfection
Acceso en línea:	http://hdl.handle.net/20.500.12110/paper_00219258_v278_n52_p52166_Nogues
Aporte de:	Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) de Universidad de Buenos Aires

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spelling	todo:paper_00219258_v278_n52_p52166_Nogues2023-10-03T14:23:04Z Influence of Polymerase II Processivity on Alternative Splicing Depends on Splice Site Strength Nogués, G. Muñoz, M.J. Kornblihtt, A.R. Cell culture Enzyme inhibition Genes Mutagenesis RNA Splice site strength Enzymes elongation factor fibronectin fibronectin extra domain I messenger RNA polypyrimidine tract binding protein RNA polymerase II unclassified drug article controlled study enzyme activation enzyme inhibition exon gene mutation human human cell molecular recognition nonhuman point mutation priority journal regulatory mechanism RNA splicing silencer element transcription regulation Alternative Splicing Binding Sites Cell Line Cell Line, Tumor Exons Herpes Simplex Virus Protein Vmw65 Humans Models, Biological Mutation Phosphorylation Plasmids Point Mutation Polymorphism, Genetic Protein Structure, Tertiary Pyrimidines RNA Polymerase II RNA Splicing RNA, Messenger Time Factors Transcription, Genetic Transfection Transcription and pre-mRNA splicing are coordinated temporally and spatially, and both processes can influence each other. In particular, control of transcriptional elongation by RNA polymerase II has proved to be important for alternative splicing regulation. In this report we demonstrate that the efficiency of exon recognition by the splicing machinery is crucial for the elongation control. Alternative splicing of the fibronectin extra domain I (EDI) is because the polypyrimidine tract of its 3'-splice site occurs suboptimal. By mutating the polypyrimidine tract of EDI in two different positions, individually or in combination, and by disrupting its exonic splicing silencer, we managed to generate minigenes with increasing degrees of exon recognition. Improvement of exon recognition is evidenced by independence from the splicing regulator SF2/ASF for inclusion. The mutated minigenes were used to transfect human cells in culture and study the responsiveness of EDI alternative splicing to activation or inhibition of pol II elongation. Our results revealed that responsiveness of exon skipping to elongation is inversely proportional to 3'-splice site strength, which means that the better the alternative exon is recognized by the splicing machinery, the less its degree of inclusion is affected by transcriptional elongation. JOUR info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by/2.5/ar http://hdl.handle.net/20.500.12110/paper_00219258_v278_n52_p52166_Nogues
institution	Universidad de Buenos Aires
institution_str	I-28
repository_str	R-134
collection	Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA)
topic	Cell culture Enzyme inhibition Genes Mutagenesis RNA Splice site strength Enzymes elongation factor fibronectin fibronectin extra domain I messenger RNA polypyrimidine tract binding protein RNA polymerase II unclassified drug article controlled study enzyme activation enzyme inhibition exon gene mutation human human cell molecular recognition nonhuman point mutation priority journal regulatory mechanism RNA splicing silencer element transcription regulation Alternative Splicing Binding Sites Cell Line Cell Line, Tumor Exons Herpes Simplex Virus Protein Vmw65 Humans Models, Biological Mutation Phosphorylation Plasmids Point Mutation Polymorphism, Genetic Protein Structure, Tertiary Pyrimidines RNA Polymerase II RNA Splicing RNA, Messenger Time Factors Transcription, Genetic Transfection
spellingShingle	Cell culture Enzyme inhibition Genes Mutagenesis RNA Splice site strength Enzymes elongation factor fibronectin fibronectin extra domain I messenger RNA polypyrimidine tract binding protein RNA polymerase II unclassified drug article controlled study enzyme activation enzyme inhibition exon gene mutation human human cell molecular recognition nonhuman point mutation priority journal regulatory mechanism RNA splicing silencer element transcription regulation Alternative Splicing Binding Sites Cell Line Cell Line, Tumor Exons Herpes Simplex Virus Protein Vmw65 Humans Models, Biological Mutation Phosphorylation Plasmids Point Mutation Polymorphism, Genetic Protein Structure, Tertiary Pyrimidines RNA Polymerase II RNA Splicing RNA, Messenger Time Factors Transcription, Genetic Transfection Nogués, G. Muñoz, M.J. Kornblihtt, A.R. Influence of Polymerase II Processivity on Alternative Splicing Depends on Splice Site Strength
topic_facet	Cell culture Enzyme inhibition Genes Mutagenesis RNA Splice site strength Enzymes elongation factor fibronectin fibronectin extra domain I messenger RNA polypyrimidine tract binding protein RNA polymerase II unclassified drug article controlled study enzyme activation enzyme inhibition exon gene mutation human human cell molecular recognition nonhuman point mutation priority journal regulatory mechanism RNA splicing silencer element transcription regulation Alternative Splicing Binding Sites Cell Line Cell Line, Tumor Exons Herpes Simplex Virus Protein Vmw65 Humans Models, Biological Mutation Phosphorylation Plasmids Point Mutation Polymorphism, Genetic Protein Structure, Tertiary Pyrimidines RNA Polymerase II RNA Splicing RNA, Messenger Time Factors Transcription, Genetic Transfection
description	Transcription and pre-mRNA splicing are coordinated temporally and spatially, and both processes can influence each other. In particular, control of transcriptional elongation by RNA polymerase II has proved to be important for alternative splicing regulation. In this report we demonstrate that the efficiency of exon recognition by the splicing machinery is crucial for the elongation control. Alternative splicing of the fibronectin extra domain I (EDI) is because the polypyrimidine tract of its 3'-splice site occurs suboptimal. By mutating the polypyrimidine tract of EDI in two different positions, individually or in combination, and by disrupting its exonic splicing silencer, we managed to generate minigenes with increasing degrees of exon recognition. Improvement of exon recognition is evidenced by independence from the splicing regulator SF2/ASF for inclusion. The mutated minigenes were used to transfect human cells in culture and study the responsiveness of EDI alternative splicing to activation or inhibition of pol II elongation. Our results revealed that responsiveness of exon skipping to elongation is inversely proportional to 3'-splice site strength, which means that the better the alternative exon is recognized by the splicing machinery, the less its degree of inclusion is affected by transcriptional elongation.
format	JOUR
author	Nogués, G. Muñoz, M.J. Kornblihtt, A.R.
author_facet	Nogués, G. Muñoz, M.J. Kornblihtt, A.R.
author_sort	Nogués, G.
title	Influence of Polymerase II Processivity on Alternative Splicing Depends on Splice Site Strength
title_short	Influence of Polymerase II Processivity on Alternative Splicing Depends on Splice Site Strength
title_full	Influence of Polymerase II Processivity on Alternative Splicing Depends on Splice Site Strength
title_fullStr	Influence of Polymerase II Processivity on Alternative Splicing Depends on Splice Site Strength
title_full_unstemmed	Influence of Polymerase II Processivity on Alternative Splicing Depends on Splice Site Strength
title_sort	influence of polymerase ii processivity on alternative splicing depends on splice site strength
url	http://hdl.handle.net/20.500.12110/paper_00219258_v278_n52_p52166_Nogues
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Influence of Polymerase II Processivity on Alternative Splicing Depends on Splice Site Strength

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