A broad analysis of splicing regulation in yeast using a large library of synthetic introns.

RNA splicing is a key process in eukaryotic gene expression, in which an intron is spliced out of a pre-mRNA molecule to eventually produce a mature mRNA. Most intron-containing genes are constitutively spliced, hence efficient splicing of an intron is crucial for efficient regulation of gene expres...

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Autores principales:	Dvir Schirman, Zohar Yakhini, Yitzhak Pilpel, Orna Dahan
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2021
Materias:	Genetics QH426-470
Acceso en línea:	https://doaj.org/article/5ba94e676d774c43b1a50c724747047d
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spelling	oai:doaj.org-article:5ba94e676d774c43b1a50c724747047d2021-12-02T20:03:19ZA broad analysis of splicing regulation in yeast using a large library of synthetic introns.1553-73901553-740410.1371/journal.pgen.1009805https://doaj.org/article/5ba94e676d774c43b1a50c724747047d2021-09-01T00:00:00Zhttps://doi.org/10.1371/journal.pgen.1009805https://doaj.org/toc/1553-7390https://doaj.org/toc/1553-7404RNA splicing is a key process in eukaryotic gene expression, in which an intron is spliced out of a pre-mRNA molecule to eventually produce a mature mRNA. Most intron-containing genes are constitutively spliced, hence efficient splicing of an intron is crucial for efficient regulation of gene expression. Here we use a large synthetic oligo library of ~20,000 variants to explore how different intronic sequence features affect splicing efficiency and mRNA expression levels in S. cerevisiae. Introns are defined by three functional sites, the 5' donor site, the branch site, and the 3' acceptor site. Using a combinatorial design of synthetic introns, we demonstrate how non-consensus splice site sequences in each of these sites affect splicing efficiency. We then show that S. cerevisiae splicing machinery tends to select alternative 3' splice sites downstream of the original site, and we suggest that this tendency created a selective pressure, leading to the avoidance of cryptic splice site motifs near introns' 3' ends. We further use natural intronic sequences from other yeast species, whose splicing machineries have diverged to various extents, to show how intron architectures in the various species have been adapted to the organism's splicing machinery. We suggest that the observed tendency for cryptic splicing is a result of a loss of a specific splicing factor, U2AF1. Lastly, we show that synthetic sequences containing two introns give rise to alternative RNA isoforms in S. cerevisiae, demonstrating that merely a synthetic fusion of two introns might be suffice to facilitate alternative splicing in yeast. Our study reveals novel mechanisms by which introns are shaped in evolution to allow cells to regulate their transcriptome. In addition, it provides a valuable resource to study the regulation of constitutive and alternative splicing in a model organism.Dvir SchirmanZohar YakhiniYitzhak PilpelOrna DahanPublic Library of Science (PLoS)articleGeneticsQH426-470ENPLoS Genetics, Vol 17, Iss 9, p e1009805 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Genetics QH426-470
spellingShingle	Genetics QH426-470 Dvir Schirman Zohar Yakhini Yitzhak Pilpel Orna Dahan A broad analysis of splicing regulation in yeast using a large library of synthetic introns.
description	RNA splicing is a key process in eukaryotic gene expression, in which an intron is spliced out of a pre-mRNA molecule to eventually produce a mature mRNA. Most intron-containing genes are constitutively spliced, hence efficient splicing of an intron is crucial for efficient regulation of gene expression. Here we use a large synthetic oligo library of ~20,000 variants to explore how different intronic sequence features affect splicing efficiency and mRNA expression levels in S. cerevisiae. Introns are defined by three functional sites, the 5' donor site, the branch site, and the 3' acceptor site. Using a combinatorial design of synthetic introns, we demonstrate how non-consensus splice site sequences in each of these sites affect splicing efficiency. We then show that S. cerevisiae splicing machinery tends to select alternative 3' splice sites downstream of the original site, and we suggest that this tendency created a selective pressure, leading to the avoidance of cryptic splice site motifs near introns' 3' ends. We further use natural intronic sequences from other yeast species, whose splicing machineries have diverged to various extents, to show how intron architectures in the various species have been adapted to the organism's splicing machinery. We suggest that the observed tendency for cryptic splicing is a result of a loss of a specific splicing factor, U2AF1. Lastly, we show that synthetic sequences containing two introns give rise to alternative RNA isoforms in S. cerevisiae, demonstrating that merely a synthetic fusion of two introns might be suffice to facilitate alternative splicing in yeast. Our study reveals novel mechanisms by which introns are shaped in evolution to allow cells to regulate their transcriptome. In addition, it provides a valuable resource to study the regulation of constitutive and alternative splicing in a model organism.
format	article
author	Dvir Schirman Zohar Yakhini Yitzhak Pilpel Orna Dahan
author_facet	Dvir Schirman Zohar Yakhini Yitzhak Pilpel Orna Dahan
author_sort	Dvir Schirman
title	A broad analysis of splicing regulation in yeast using a large library of synthetic introns.
title_short	A broad analysis of splicing regulation in yeast using a large library of synthetic introns.
title_full	A broad analysis of splicing regulation in yeast using a large library of synthetic introns.
title_fullStr	A broad analysis of splicing regulation in yeast using a large library of synthetic introns.
title_full_unstemmed	A broad analysis of splicing regulation in yeast using a large library of synthetic introns.
title_sort	broad analysis of splicing regulation in yeast using a large library of synthetic introns.
publisher	Public Library of Science (PLoS)
publishDate	2021
url	https://doaj.org/article/5ba94e676d774c43b1a50c724747047d
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A broad analysis of splicing regulation in yeast using a large library of synthetic introns.

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