scholarly journals Interpretation of mRNA splicing mutations in genetic disease: review of the literature and guidelines for information-theoretical analysis

F1000Research ◽  
2014 ◽  
Vol 3 ◽  
pp. 282 ◽  
Author(s):  
Natasha G. Caminsky ◽  
Eliseos J. Mucaki ◽  
Peter K. Rogan
Keyword(s):  
Splice Site ◽  
Genetic Disease ◽  
Mrna Splicing ◽  
Regulatory Sequences ◽  
Splice Sites ◽  
Review Of The Literature ◽  
Cryptic Splice Site ◽  
Broad Perspective ◽  
Genomic Variants ◽  
Splicing Mutations

The interpretation of genomic variants has become one of the paramount challenges in the post-genome sequencing era. In this review we summarize nearly 20 years of research on the applications of information theory (IT) to interpret coding and non-coding mutations that alter mRNA splicing in rare and common diseases. We compile and summarize the spectrum of published variants analyzed by IT, to provide a broad perspective of the distribution of deleterious natural and cryptic splice site variants detected, as well as those affecting splicing regulatory sequences. Results for natural splice site mutations can be interrogated dynamically with Splicing Mutation Calculator, a companion software program that computes changes in information content for any splice site substitution, linked to corresponding publications containing these mutations. The accuracy of IT-based analysis was assessed in the context of experimentally validated mutations. Because splice site information quantifies binding affinity, IT-based analyses can discern the differences between variants that account for the observed reduced (leaky) versus abolished mRNA splicing. We extend this principle by comparing predicted mutations in natural, cryptic, and regulatory splice sites with observed deleterious phenotypic and benign effects. Our analysis of 1727 variants revealed a number of general principles useful for ensuring portability of these analyses and accurate input and interpretation of mutations. We offer guidelines for optimal use of IT software for interpretation of mRNA splicing mutations.

scholarly journals Interpretation of mRNA splicing mutations in genetic disease: review of the literature and guidelines for information-theoretical analysis

F1000Research ◽  
2015 ◽  
Vol 3 ◽  
pp. 282 ◽  
Author(s):  
Natasha G. Caminsky ◽  
Eliseos J. Mucaki ◽  
Peter K. Rogan
Keyword(s):  
Splice Site ◽  
Genetic Disease ◽  
Mrna Splicing ◽  
Regulatory Sequences ◽  
Splice Sites ◽  
Review Of The Literature ◽  
Cryptic Splice Site ◽  
Broad Perspective ◽  
Genomic Variants ◽  
Splicing Mutations

The interpretation of genomic variants has become one of the paramount challenges in the post-genome sequencing era. In this review we summarize nearly 20 years of research on the applications of information theory (IT) to interpret coding and non-coding mutations that alter mRNA splicing in rare and common diseases. We compile and summarize the spectrum of published variants analyzed by IT, to provide a broad perspective of the distribution of deleterious natural and cryptic splice site variants detected, as well as those affecting splicing regulatory sequences. Results for natural splice site mutations can be interrogated dynamically with Splicing Mutation Calculator, a companion software program that computes changes in information content for any splice site substitution, linked to corresponding publications containing these mutations. The accuracy of IT-based analysis was assessed in the context of experimentally validated mutations. Because splice site information quantifies binding affinity, IT-based analyses can discern the differences between variants that account for the observed reduced (leaky) versus abolished mRNA splicing. We extend this principle by comparing predicted mutations in natural, cryptic, and regulatory splice sites with observed deleterious phenotypic and benign effects. Our analysis of 1727 variants revealed a number of general principles useful for ensuring portability of these analyses and accurate input and interpretation of mutations. We offer guidelines for optimal use of IT software for interpretation of mRNA splicing mutations.

scholarly journals Background splicing and genetic disease

2020 ◽  
Author(s):  
Diana Alexieva ◽  
Yi Long ◽  
Rupa Sarkar ◽  
Hansraj Dhayan ◽  
Emmanuel Bruet ◽  
...  
Keyword(s):  
Splice Site ◽  
Genetic Disease ◽  
Exon Skipping ◽  
Splice Sites ◽  
Cryptic Splice Site ◽  
Aberrant Splicing ◽  
Low Level ◽  
Splicing Mutations ◽  
Brca1 Brca2 ◽  
Rna Splice Sites

Abstract We report that low level background splicing by normal genes can be used to predict the likely effect of splicing mutations upon cryptic splice site activation and exon skipping, with emphasis on the DBASS databases, BRCA1, BRCA2 and DMD. In addition we show that background RNA splice sites are also involved in pseudoexon formation, recursive splicing and aberrant splicing in cancer. We discuss how background splicing information might inform splicing therapy.

scholarly journals U1 small nuclear RNAs with altered specificity can be stably expressed in mammalian cells and promote permanent changes in pre-mRNA splicing.

1993 ◽  
Vol 13 (5) ◽  
pp. 2666-2676 ◽  
Author(s):  
J B Cohen ◽  
S D Broz ◽  
A D Levinson
Keyword(s):  
Splice Site ◽  
Mammalian Cells ◽  
Mrna Processing ◽  
Mrna Splicing ◽  
Small Nuclear Rna ◽  
Splice Sites ◽  
Gene Complementation ◽  
Small Nuclear Rnas ◽  
Nuclear Rna ◽  
Mutant Genes

Pre-mRNA 5' splice site activity depends, at least in part, on base complementarity to U1 small nuclear RNA. In transient coexpression assays, defective 5' splice sites can regain activity in the presence of U1 carrying compensatory changes, but it is unclear whether such mutant U1 RNAs can be permanently expressed in mammalian cells. We have explored this issue to determine whether U1 small nuclear RNAs with altered specificity may be of value to rescue targeted mutant genes or alter pre-mRNA processing profiles. This effort was initiated following our observation that U1 with specificity for a splice site associated with an alternative H-ras exon substantially reduced the synthesis of the potentially oncogenic p21ras protein in transient assays. We describe the development of a mammalian complementation system that selects for removal of a splicing-defective intron placed within a drug resistance gene. Complementation was observed in proportion to the degree of complementarity between transfected mutant U1 genes and different defective splice sites, and all cells selected in this manner were found to express mutant U1 RNA. In addition, these cells showed specific activation of defective splice sites presented by an unlinked reporter gene. We discuss the prospects of this approach to permanently alter the expression of targeted genes in mammalian cells.

Spontaneous splicing mutations at the dihydrofolate reductase locus in Chinese hamster ovary cells

1986 ◽  
Vol 6 (6) ◽  
pp. 1926-1935
Author(s):  
P J Mitchell ◽  
G Urlaub ◽  
L Chasin
Keyword(s):  
Amino Acid ◽  
Dihydrofolate Reductase ◽  
Splice Site ◽  
Chinese Hamster Ovary ◽  
Chinese Hamster Ovary Cells ◽  
Chinese Hamster ◽  
Splice Sites ◽  
Ovary Cells ◽  
Splicing Mutations ◽  
Intron 4

We isolated and characterized three spontaneous mutants of Chinese hamster ovary cells that were deficient in dihydrofolate reductase activity. All three mutants contained no detectable enzyme activity and produced dihydrofolate reductase mRNA species that were shorter than those of the wild type by about 120 bases. Six exons are normally represented in this mRNA; exon 5 was missing in all three mutant mRNAs. Nuclease S1 analysis of the three mutants indicated that during the processing of the mutant RNA, exon 4 was spliced to exon 6. The three mutant genes were cloned, and the regions around exons 4 and 5 were sequenced. In one mutant, the GT dinucleotide at the 5' end of intron 5 had changed to CT. In a second mutant, the first base in exon 5 had changed from G to T. In a revertant of this mutant, this base was further mutated to A, a return to a purine. Approximately 25% of the mRNA molecules in the revertant were spliced correctly to produce an enzyme with one presumed amino acid change. In the third mutant, the AG at the 3' end of intron 4 had changed to AA. A mutation that partially reversed the mutant phenotype had changed the dinucleotide at the 5' end of intron 4 from GT to AT. The splicing pattern in this revertant was consistent with the use of cryptic donor and acceptor splice sites close to the original sites to produce an mRNA with three base changes and a protein with two amino acid changes. These mutations argue against a scanning model for the selection of splice site pairs and suggest that only a single splice site need be inactivated to bring about efficient exon skipping (a regulatory mechanism for some genes). The fact that all three mutants analyzed exhibited exon 5 splicing mutations indicates that these splice sites are hot spots for spontaneous mutation.

scholarly journals Splice site mutations are a common cause of X-linked chronic granulomatous disease

Blood ◽  
1992 ◽  
Vol 80 (6) ◽  
pp. 1553-1558 ◽  
Author(s):  
M de Boer ◽  
BG Bolscher ◽  
MC Dinauer ◽  
SH Orkin ◽  
CI Smith ◽  
...  
Keyword(s):  
Chronic Granulomatous Disease ◽  
Splice Site ◽  
Messenger Rna ◽  
Mrna Splicing ◽  
Molecular Defect ◽  
Granulomatous Disease ◽  
Splice Sites ◽  
Nucleotide Substitutions ◽  
Common Cause ◽  
Donor Splice

Chronic granulomatous disease (CGD) is characterized by the absence of a respiratory burst in activated phagocytes. Defects in at least four different genes lead to CGD. Patients with the X-linked form of CGD have mutations in the gene for the beta-subunit of cytochrome b558 (gp91-phox). We studied the molecular defect in four patients with X- linked CGD. In a fifth family, we studied the mother of a patient with X-linked CGD who had died before our investigations. Gp91-phox messenger RNA (mRNA) was reverse transcribed into cDNA and the coding region was amplified by polymerase chain reaction into three fragments. Sequence analysis showed the absence of the exon 7, 5, 3, and 2 sequences in patients 1, 2, 3, and 4, respectively. In carrier 5, we found both normal cDNA and cDNA that lacked 57 3′-nucleotides of exon 6. We analyzed the splice sites of the flanking introns of the missing exons. In patients 1, 2, and 3, we found single nucleotide substitutions within the first five positions of the down-stream 5′ donor splice sites. In patient 4, a similar substitution was found at position -1 of the 3′ acceptor splice site of intron 1. In carrier 5, no mutation was found in the exon 6-intron 6 boundary sequence. Instead, a single substitution was observed in exon 6 (C----A at nucleotide 633) that created a new donor splice site. Apparently, mRNA splicing occurs preferentially at this newly created splice site. We conclude that the absence of the exon sequences in the gp91-phox mRNA of these patients is due to splicing errors. Of 30 European X-linked CGD patients studied by us so far, five appear to be caused by mutations that affect correct mRNA splicing. Thus, such mutations appear to be a common cause of X-linked CGD.

scholarly journals Pan-cancer repository of validated natural and cryptic mRNA splicing mutations

F1000Research ◽  
2019 ◽  
Vol 7 ◽  
pp. 1908 ◽  
Author(s):  
Ben C. Shirley ◽  
Eliseos J. Mucaki ◽  
Peter K. Rogan
Keyword(s):  
Splice Junction ◽  
Cancer Genome ◽  
Mrna Splicing ◽  
The Cancer Genome Atlas ◽  
Splice Sites ◽  
Global Alliance ◽  
Multiple Tumor ◽  
Link Type ◽  
Splicing Mutations ◽  
Single Nucleotide Polymorphism Database

We present a major public resource of mRNA splicing mutations validated according to multiple lines of evidence of abnormal gene expression. Likely mutations present in all tumor types reported in the Cancer Genome Atlas (TCGA) and the International Cancer Genome Consortium (ICGC) were identified based on the comparative strengths of splice sites in tumor versus normal genomes, and then validated by respectively comparing counts of splice junction spanning and abundance of transcript reads in RNA-Seq data from matched tissues and tumors lacking these mutations. The comprehensive resource features 341,486 of these validated mutations, the majority of which (69.9%) are not present in the Single Nucleotide Polymorphism Database (dbSNP 150). There are 131,347 unique mutations which weaken or abolish natural splice sites, and 222,071 mutations which strengthen cryptic splice sites (11,932 affect both simultaneously). 28,812 novel or rare flagged variants (with <1% population frequency in dbSNP) were observed in multiple tumor tissue types. Single variants or chromosome ranges can be queried using a Global Alliance for Genomics and Health (GA4GH)-compliant, web-based Beacon “Validated Splicing Mutations” either separately or in aggregate alongside other Beacons through the public Beacon Network, as well as through our website.

U1 small nuclear RNAs with altered specificity can be stably expressed in mammalian cells and promote permanent changes in pre-mRNA splicing

1993 ◽  
Vol 13 (5) ◽  
pp. 2666-2676
Author(s):  
J B Cohen ◽  
S D Broz ◽  
A D Levinson
Keyword(s):  
Splice Site ◽  
Mammalian Cells ◽  
Mrna Processing ◽  
Mrna Splicing ◽  
Small Nuclear Rna ◽  
Splice Sites ◽  
Gene Complementation ◽  
Small Nuclear Rnas ◽  
Nuclear Rna ◽  
Mutant Genes

Pre-mRNA 5' splice site activity depends, at least in part, on base complementarity to U1 small nuclear RNA. In transient coexpression assays, defective 5' splice sites can regain activity in the presence of U1 carrying compensatory changes, but it is unclear whether such mutant U1 RNAs can be permanently expressed in mammalian cells. We have explored this issue to determine whether U1 small nuclear RNAs with altered specificity may be of value to rescue targeted mutant genes or alter pre-mRNA processing profiles. This effort was initiated following our observation that U1 with specificity for a splice site associated with an alternative H-ras exon substantially reduced the synthesis of the potentially oncogenic p21ras protein in transient assays. We describe the development of a mammalian complementation system that selects for removal of a splicing-defective intron placed within a drug resistance gene. Complementation was observed in proportion to the degree of complementarity between transfected mutant U1 genes and different defective splice sites, and all cells selected in this manner were found to express mutant U1 RNA. In addition, these cells showed specific activation of defective splice sites presented by an unlinked reporter gene. We discuss the prospects of this approach to permanently alter the expression of targeted genes in mammalian cells.

Virus deletion mutants that affect a 3' splice site in the E3 transcription unit of adenovirus 2

1985 ◽  
Vol 5 (9) ◽  
pp. 2405-2413
Author(s):  
B M Bhat ◽  
H A Brady ◽  
W S Wold
Keyword(s):  
Steady State ◽  
Splice Site ◽  
Transcription Unit ◽  
Mrna Splicing ◽  
Deletion Mutants ◽  
Splice Sites ◽  
Rich Region ◽  
Specific Sequence ◽  
Exon Deletion

Five viable virus mutants were constructed with deletions near a 3' splice site located at nucleotide 2157 in the E3 transcription unit of adenovirus 2. The mutants were examined for splicing activity at the 2157 3' splice site in vivo by nuclease-gel analysis of steady-state cytoplasmic mRNA. Splicing was not prevented by an exon deletion (dl719) that leaves 16 5'-proximal exon nucleotides intact or by intron deletions that leave 34 (dl717, dl712) or 18 (dl716) 3'-proximal intron nucleotides intact. The sequences deleted in one of these intron mutants (dl716) include the putative branchpoint site used in lariat formation during splicing. Thus, a surrogate branchpoint site apparently can be used for splicing. Another intron mutant (dl714) has a deletion that leaves 15 3'-proximal intron nucleotides intact; remarkably, this deletion virtually abolished splicing, even though the deletion is only 3 nucleotides closer to the splice site than is the deletion in dl716 which splices normally. The three nucleotides deleted in dl714 that are retained by dl716 are the sequence TGT. The TGT sequence is located on the 5' boundary of the pyrimidine-rich region upstream of the nucleotide 2157 3' splice site. Such pyrimidine-rich regions are ubiquitous at 3' splice sites. Most likely, the TGT is required for splicing at the nucleotide 2157 3' splice site. The TGT may be important because of its specific sequence or because it forms the 5' boundary of the pyrimidine-rich region.

scholarly journals Pan-cancer repository of validated natural and cryptic mRNA splicing mutations

F1000Research ◽  
2018 ◽  
Vol 7 ◽  
pp. 1908 ◽  
Author(s):  
Ben C. Shirley ◽  
Eliseos J. Mucaki ◽  
Peter K. Rogan
Keyword(s):  
Splice Junction ◽  
Mrna Splicing ◽  
The Cancer Genome Atlas ◽  
Splice Sites ◽  
Global Alliance ◽  
Multiple Tumor ◽  
Link Type ◽  
Splicing Mutations ◽  
Public Resource ◽  
Single Nucleotide Polymorphism Database

We present a major public resource of mRNA splicing mutations validated according to multiple lines of evidence of abnormal gene expression. Likely mutations present in all tumor types reported in the Cancer Genome Atlas (TCGA) were identified based on the comparative strengths of splice sites in tumor versus normal genomes, and then validated by respectively comparing counts of splice junction spanning and abundance of transcript reads in RNA-Seq data from matched tissues and tumors lacking these mutations. The comprehensive resource features 351,423 of these validated mutations, the majority of which (69.1%) are not present in the Single Nucleotide Polymorphism Database (dbSNP 150). There are 117,951 unique mutations which weaken or abolish natural splice sites, and 244,415 mutations which strengthen cryptic splice sites (10,943 affect both simultaneously). 27,803 novel or rare flagged variants (with <1% population frequency in dbSNP) were observed in multiple tumor tissue types. Single variants or chromosome ranges can be queried using a Global Alliance for Genomics and Health (GA4GH)-compliant, web-based Beacon “Validated Splicing Mutations” either separately or in aggregate alongside other Beacons through the public Beacon Network (http://www.beacon-network.org/#/search?beacon=cytognomix), as well as through our website (https://validsplicemut.cytognomix.com/).

scholarly journals Pan-Cancer Repository of Validated Natural and Cryptic mRNA Splicing Mutations

2018 ◽  
Author(s):  
Ben C. Shirley ◽  
Eliseos J. Mucaki ◽  
Peter K. Rogan
Keyword(s):  
Splice Junction ◽  
Mrna Splicing ◽  
The Cancer Genome Atlas ◽  
Splice Sites ◽  
Global Alliance ◽  
Multiple Tumor ◽  
Link Type ◽  
Splicing Mutations ◽  
Public Resource ◽  
Single Nucleotide Polymorphism Database

AbstractWe present a major public resource of mRNA splicing mutations validated according to multiple lines of evidence of abnormal gene expression. Likely mutations present in all tumor types reported in the Cancer Genome Atlas (TCGA) were identified based on the comparative strengths of splice sites in tumor versus normal genomes and then validated by respectively comparing counts of splice junction spanning and abundance of transcript reads in RNA-Seq data from matched tissues and tumors lacking these mutations. The comprehensive resource features 351,423 of these validated mutations, the majority of which (69.1%) are not featured in the Single Nucleotide Polymorphism Database (dbSNP 150). There are 117,951 unique mutations which weaken or abolish natural splice sites, and 244,415 mutations which strengthen cryptic splice sites (10,943 affect both simultaneously). 27,803 novel or rare flagged variants (with <1% population frequency in dbSNP) were observed in multiple tumor tissue types. Single variants or chromosome ranges can be queried using a Global Alliance for Genomics and Health (GA4GH)-compliant web Beacon, Validated Splicing Mutations, either separately or in aggregate alongside other beacons through the public Beacon Network (http://www.beacon-network.org/#/search?beacon=cytognomix), as well as through our website (https://validsplicemut.cytognomix.com/).

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