Phenotypic consequences of branch point substitutions
Phenotypic consequences of branch point substitutions
The branch point sequence (BPS) is a conserved splicing signal important for spliceosome assembly and lariat intron formation. BPS mutations may result in aberrant pre-mRNA splicing and genetic disorders, but their phenotypic consequences have been difficult to predict, largely due to a highly degenerate nature of the BPS consensus. Here, we have examined the splicing pattern of nine reporter pre-mRNAs that have previously been shown to give rise to human hereditary diseases as a result of single-nucleotide substitutions in the predicted BPS. Increased exon skipping and intron retention observed in vivo were recapitulated for each mutated pre-mRNA, but the reproducibility of cryptic splice site activation was lower. BP mutations in reporter pre-mRNAs frequently induced aberrant 3' splice sites and also activated a cryptic 5' splice site. Systematic mutagenesis of BP adenosines showed that in most pre-mRNAs, the expression of canonical transcripts was lower for BP transitions than BP transversions. Differential splicing outcome for transitions vs. transversions was abrogated or reduced if introns were truncated to 200 nt or less, suggesting that the nature of the BP residue is less critical for interactions across very short introns. Together, these results improve prediction of phenotypic consequences of point mutations upstream of splice acceptor sites and suggest that the overrepresentation of disease-causing adenosine-to-guanosine BP substitutions observed in Mendelian disorders is due to more profound defects of gene expression at the level of pre-mRNA splicing
mutagenesis, site-directed, rna splicing, gene expression, research, sequence analysis, phenotype, acid, introns, adenosine, polymorphism, regulatory sequences, rna, humans, metabolism, genetic predisposition to disease, cell line, disease, expression, rna splice sites, single nucleotide, point mutation, spliceosomes, genetics, ribonucleic acid, human, mutation, physiology, research support
803-813
Královiová, Jana
b3e0c1e7-05ed-445d-b3d9-ace11e3b4878
Lei, Haixin
24595b39-a4e6-4cfc-9354-5001b7f70d5a
Voechovský, Igor
7245de2f-8c9b-4034-8935-9a451d9b682e
August 2006
Královiová, Jana
b3e0c1e7-05ed-445d-b3d9-ace11e3b4878
Lei, Haixin
24595b39-a4e6-4cfc-9354-5001b7f70d5a
Voechovský, Igor
7245de2f-8c9b-4034-8935-9a451d9b682e
Královiová, Jana, Lei, Haixin and Voechovský, Igor
(2006)
Phenotypic consequences of branch point substitutions.
Human Mutation, 27 (8), .
(doi:10.1002/humu.20362).
Abstract
The branch point sequence (BPS) is a conserved splicing signal important for spliceosome assembly and lariat intron formation. BPS mutations may result in aberrant pre-mRNA splicing and genetic disorders, but their phenotypic consequences have been difficult to predict, largely due to a highly degenerate nature of the BPS consensus. Here, we have examined the splicing pattern of nine reporter pre-mRNAs that have previously been shown to give rise to human hereditary diseases as a result of single-nucleotide substitutions in the predicted BPS. Increased exon skipping and intron retention observed in vivo were recapitulated for each mutated pre-mRNA, but the reproducibility of cryptic splice site activation was lower. BP mutations in reporter pre-mRNAs frequently induced aberrant 3' splice sites and also activated a cryptic 5' splice site. Systematic mutagenesis of BP adenosines showed that in most pre-mRNAs, the expression of canonical transcripts was lower for BP transitions than BP transversions. Differential splicing outcome for transitions vs. transversions was abrogated or reduced if introns were truncated to 200 nt or less, suggesting that the nature of the BP residue is less critical for interactions across very short introns. Together, these results improve prediction of phenotypic consequences of point mutations upstream of splice acceptor sites and suggest that the overrepresentation of disease-causing adenosine-to-guanosine BP substitutions observed in Mendelian disorders is due to more profound defects of gene expression at the level of pre-mRNA splicing
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Published date: August 2006
Keywords:
mutagenesis, site-directed, rna splicing, gene expression, research, sequence analysis, phenotype, acid, introns, adenosine, polymorphism, regulatory sequences, rna, humans, metabolism, genetic predisposition to disease, cell line, disease, expression, rna splice sites, single nucleotide, point mutation, spliceosomes, genetics, ribonucleic acid, human, mutation, physiology, research support
Identifiers
Local EPrints ID: 59944
URI: http://eprints.soton.ac.uk/id/eprint/59944
ISSN: 1059-7794
PURE UUID: 6282c669-b755-4a7f-a2cc-468b673e7ad3
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Date deposited: 05 Sep 2008
Last modified: 16 Mar 2024 03:32
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Author:
Jana Královiová
Author:
Haixin Lei
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