Exon First Nucleotide Mutations in Splicing: Evaluation of In Silico Prediction Tools
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{"title"=>"Exon first nucleotide mutations in splicing: Evaluation of in silico prediction tools", "type"=>"journal", "authors"=>[{"first_name"=>"Lucie", "last_name"=>"Grodecká", "scopus_author_id"=>"35758982100"}, {"first_name"=>"Pavla", "last_name"=>"Lockerová", "scopus_author_id"=>"56068071700"}, {"first_name"=>"Barbora", "last_name"=>"Ravčuková", "scopus_author_id"=>"12770897800"}, {"first_name"=>"Emanuele", "last_name"=>"Buratti", "scopus_author_id"=>"7003676483"}, {"first_name"=>"Francisco E.", "last_name"=>"Baralle", "scopus_author_id"=>"7006772526"}, {"first_name"=>"Ladislav", "last_name"=>"Dušek", "scopus_author_id"=>"8559629900"}, {"first_name"=>"Tomáš", "last_name"=>"Freiberger", "scopus_author_id"=>"55885407200"}], "year"=>2014, "source"=>"PLoS ONE", "identifiers"=>{"sgr"=>"84896116359", "pmid"=>"24586880", "scopus"=>"2-s2.0-84896116359", "issn"=>"19326203", "pui"=>"372608164", "doi"=>"10.1371/journal.pone.0089570"}, "id"=>"ef7ce68d-5dac-3011-9c34-ee4dcc5ab922", "abstract"=>"Mutations in the first nucleotide of exons (E(+1)) mostly affect pre-mRNA splicing when found in AG-dependent 3' splice sites, whereas AG-independent splice sites are more resistant. The AG-dependency, however, may be difficult to assess just from primary sequence data as it depends on the quality of the polypyrimidine tract. For this reason, in silico prediction tools are commonly used to score 3' splice sites. In this study, we have assessed the ability of sequence features and in silico prediction tools to discriminate between the splicing-affecting and non-affecting E(+1) variants. For this purpose, we newly tested 16 substitutions in vitro and derived other variants from literature. Surprisingly, we found that in the presence of the substituting nucleotide, the quality of the polypyrimidine tract alone was not conclusive about its splicing fate. Rather, it was the identity of the substituting nucleotide that markedly influenced it. Among the computational tools tested, the best performance was achieved using the Maximum Entropy Model and Position-Specific Scoring Matrix. As a result of this study, we have now established preliminary discriminative cut-off values showing sensitivity up to 95% and specificity up to 90%. This is expected to improve our ability to detect splicing-affecting variants in a clinical genetic setting.", "link"=>"http://www.mendeley.com/research/exon-first-nucleotide-mutations-splicing-evaluation-silico-prediction-tools", "reader_count"=>13, "reader_count_by_academic_status"=>{"Researcher"=>6, "Student > Doctoral Student"=>1, "Student > Ph. D. Student"=>1, "Student > Postgraduate"=>1, "Student > Master"=>3, "Other"=>1}, "reader_count_by_user_role"=>{"Researcher"=>6, "Student > Doctoral Student"=>1, "Student > Ph. D. Student"=>1, "Student > Postgraduate"=>1, "Student > Master"=>3, "Other"=>1}, "reader_count_by_subject_area"=>{"Unspecified"=>1, "Biochemistry, Genetics and Molecular Biology"=>5, "Mathematics"=>2, "Agricultural and Biological Sciences"=>4, "Medicine and Dentistry"=>1}, "reader_count_by_subdiscipline"=>{"Medicine and Dentistry"=>{"Medicine and Dentistry"=>1}, "Agricultural and Biological Sciences"=>{"Agricultural and Biological Sciences"=>4}, "Biochemistry, Genetics and Molecular Biology"=>{"Biochemistry, Genetics and Molecular Biology"=>5}, "Mathematics"=>{"Mathematics"=>2}, "Unspecified"=>{"Unspecified"=>1}}, "reader_count_by_country"=>{"Italy"=>1}, "group_count"=>0}

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Figshare

  • {"files"=>["https://ndownloader.figshare.com/files/1394564", "https://ndownloader.figshare.com/files/1394565", "https://ndownloader.figshare.com/files/1394566", "https://ndownloader.figshare.com/files/1394567", "https://ndownloader.figshare.com/files/1394568", "https://ndownloader.figshare.com/files/1394569"], "description"=>"<div><p>Mutations in the first nucleotide of exons (E<sup>+1</sup>) mostly affect pre-mRNA splicing when found in AG-dependent 3′ splice sites, whereas AG-independent splice sites are more resistant. The AG-dependency, however, may be difficult to assess just from primary sequence data as it depends on the quality of the polypyrimidine tract. For this reason, <i>in silico</i> prediction tools are commonly used to score 3′ splice sites. In this study, we have assessed the ability of sequence features and <i>in silico</i> prediction tools to discriminate between the splicing-affecting and non-affecting E<sup>+1</sup> variants. For this purpose, we newly tested 16 substitutions <i>in vitro</i> and derived other variants from literature. Surprisingly, we found that in the presence of the substituting nucleotide, the quality of the polypyrimidine tract alone was not conclusive about its splicing fate. Rather, it was the identity of the substituting nucleotide that markedly influenced it. Among the computational tools tested, the best performance was achieved using the Maximum Entropy Model and Position-Specific Scoring Matrix. As a result of this study, we have now established preliminary discriminative cut-off values showing sensitivity up to 95% and specificity up to 90%. This is expected to improve our ability to detect splicing-affecting variants in a clinical genetic setting.</p></div>", "links"=>[], "tags"=>["Computational biology", "Molecular genetics", "gene expression", "genetics", "Gene splicing", "Molecular cell biology", "RNA processing", "exon", "nucleotide", "mutations", "tools"], "article_id"=>941339, "categories"=>["Biological Sciences"], "users"=>["Lucie Grodecká", "Pavla Lockerová", "Barbora Ravčuková", "Emanuele Buratti", "Francisco E. Baralle", "Ladislav Dušek", "Tomas Freiberger"], "doi"=>["https://dx.doi.org/10.1371/journal.pone.0089570.s001", "https://dx.doi.org/10.1371/journal.pone.0089570.s002", "https://dx.doi.org/10.1371/journal.pone.0089570.s003", "https://dx.doi.org/10.1371/journal.pone.0089570.s004", "https://dx.doi.org/10.1371/journal.pone.0089570.s005", "https://dx.doi.org/10.1371/journal.pone.0089570.s006"], "stats"=>{"downloads"=>29, "page_views"=>9, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/Exon_First_Nucleotide_Mutations_in_Splicing_Evaluation_of_In_Silico_Prediction_Tools/941339", "title"=>"Exon First Nucleotide Mutations in Splicing: Evaluation of <i>In Silico</i> Prediction Tools", "pos_in_sequence"=>0, "defined_type"=>4, "published_date"=>"2014-02-21 02:59:01"}
  • {"files"=>["https://ndownloader.figshare.com/files/1394546"], "description"=>"a<p>The cut-off values were proposed according to the predicted values obtained using the test set of naturally occurring G<sup>+1</sup> mutations (as used in <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0089570#pone-0089570-t001\" target=\"_blank\">Tables 1</a> and <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0089570#pone-0089570-t002\" target=\"_blank\">2</a>). <sup>b</sup>The sensitivity and specificity calculated using Fu-mut set of G<sup>+1</sup> mutations, containing artificially mutated PPTs. <sup>c</sup>The sensitivity and specificity calculated using the original test set of G<sup>+1</sup> mutations. The predicted scores and percentiles below the cut-off values and the differences between the predicted values for wild type and mutant sequences above the cut-off values are supposed to pertain to variants prone to affect splicing.</p>", "links"=>[], "tags"=>["Computational biology", "Molecular genetics", "gene expression", "genetics", "Gene splicing", "Molecular cell biology", "RNA processing", "cut-off", "tools", "discriminate", "ag-dependent", "ag-independent"], "article_id"=>941321, "categories"=>["Biological Sciences"], "users"=>["Lucie Grodecká", "Pavla Lockerová", "Barbora Ravčuková", "Emanuele Buratti", "Francisco E. Baralle", "Ladislav Dušek", "Tomas Freiberger"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0089570.t003", "stats"=>{"downloads"=>0, "page_views"=>2, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Proposed_cut_off_values_for_the_in_silico_tools_that_discriminate_AG_dependent_3_ss_from_AG_independent_3_ss_/941321", "title"=>"Proposed cut-off values for the <i>in silico</i> tools that discriminate AG-dependent 3′ss from AG-independent 3′ss.", "pos_in_sequence"=>0, "defined_type"=>3, "published_date"=>"2014-02-21 02:59:01"}
  • {"files"=>["https://ndownloader.figshare.com/files/1394544"], "description"=>"a<p>Each combined prediction was considered positive if two or more of the three predicted values exceeded the herein proposed cut-off values of the individual tools. <sup>b</sup>The sensitivity and specificity was calculated using Fu-mut set of G<sup>+1</sup> mutations, containing artificially mutated PPTs. <sup>c</sup>The sensitivity and specificity calculated using combined sets: test set and borderline set of G<sup>+1</sup> mutations. Py25 = number of pyrimidines in the 25 nucleotides upstream from splice site; ME s.d. = difference between wild type and mutant sequence scores predicted by MaxEnt program; ME p.d. = difference between wild type and mutant sequence percentiles predicted by MaxEnt program; PSSM s.d.: accordingly.</p>", "links"=>[], "tags"=>["Computational biology", "Molecular genetics", "gene expression", "genetics", "Gene splicing", "Molecular cell biology", "RNA processing", "predictions"], "article_id"=>941319, "categories"=>["Biological Sciences"], "users"=>["Lucie Grodecká", "Pavla Lockerová", "Barbora Ravčuková", "Emanuele Buratti", "Francisco E. Baralle", "Ladislav Dušek", "Tomas Freiberger"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0089570.t004", "stats"=>{"downloads"=>0, "page_views"=>10, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Results_of_combined_predictions_in_discrimination_of_G_1_dependent_3_8242_ss_from_G_1_independent_3_8242_ss_/941319", "title"=>"Results of combined predictions in discrimination of G<sup>+1</sup>-dependent 3′ss from G<sup>+1</sup>-independent 3′ss.", "pos_in_sequence"=>0, "defined_type"=>3, "published_date"=>"2014-02-21 02:59:01"}
  • {"files"=>["https://ndownloader.figshare.com/files/1394545"], "description"=>"<p>Computational predictions for experimentally confirmed test set of splicing-affecting and non-affecting G<sup>+1</sup> mutations were subjected to statistical analysis using the Mann-Whitney test (see Table S1 <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0089570#pone.0089570.s006\" target=\"_blank\">File S1</a> for details). Significant differences (p<0.05) are marked in bold. Diff. = difference, M-W test = Mann-Whitney test, perc. = percentile.</p>", "links"=>[], "tags"=>["Computational biology", "Molecular genetics", "gene expression", "genetics", "Gene splicing", "Molecular cell biology", "RNA processing", "ranges", "mutations", "instruments", "evaluating"], "article_id"=>941320, "categories"=>["Biological Sciences"], "users"=>["Lucie Grodecká", "Pavla Lockerová", "Barbora Ravčuková", "Emanuele Buratti", "Francisco E. Baralle", "Ladislav Dušek", "Tomas Freiberger"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0089570.t001", "stats"=>{"downloads"=>1, "page_views"=>8, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Predicted_value_ranges_in_the_test_set_of_G_1_mutations_obtained_from_the_instruments_evaluating_the_overall_strength_of_the_3_8242_splice_site_/941320", "title"=>"Predicted value ranges in the test set of G<sup>+1</sup> mutations obtained from the instruments evaluating the overall strength of the 3′splice site.", "pos_in_sequence"=>0, "defined_type"=>3, "published_date"=>"2014-02-21 02:59:01"}
  • {"files"=>["https://ndownloader.figshare.com/files/1394543"], "description"=>"<p>The boxplots show the values, median and interquartile range of the values predicted for the test set sequences. The values of scores are shown in the original units of the tools, differences are counted as ratios of the absolute score (or percentile) difference to the wild type score (or percentile). Other values are simply numbers of nucleotides. Asterisks indicate statistically significant differences between the two sets of values (*at p<0.05; **at p<0.01; ***at p<0.001). Abbreviations: Ab = group of sequence variants that lead to aberrant splicing; diff. = difference; Non = group of sequence variants that do not disrupt the process of splicing; nt = nucleotide(s); perc. = percentile; Py in 25 nt = number of pyrimidines in 25 nucleotides upstream of acceptor splice site.</p>", "links"=>[], "tags"=>["Computational biology", "Molecular genetics", "gene expression", "genetics", "Gene splicing", "Molecular cell biology", "RNA processing", "predictions", "ag-dependent", "ag-independent", "splice"], "article_id"=>941318, "categories"=>["Biological Sciences"], "users"=>["Lucie Grodecká", "Pavla Lockerová", "Barbora Ravčuková", "Emanuele Buratti", "Francisco E. Baralle", "Ladislav Dušek", "Tomas Freiberger"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0089570.g004", "stats"=>{"downloads"=>1, "page_views"=>8, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Selected_results_of_in_silico_predictions_and_other_parameters_of_AG_dependent_and_AG_independent_splice_sites_/941318", "title"=>"Selected results of <i>in silico</i> predictions and other parameters of AG-dependent and AG-independent splice sites.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2014-02-21 02:59:01"}
  • {"files"=>["https://ndownloader.figshare.com/files/1394540"], "description"=>"<p>The sequences are ordered according to the length of their longest PPS. Exons whose splicing was shown to depend on intact E<sup>+1</sup> position are underlined. The PPS are singly underlined and other polypyrimidine stretches are dashed underlined. Sites of mutations are showed in bold. Seq. = sequence. (A) Sequences of the test set. (B) Sequences of the borderline set.</p>", "links"=>[], "tags"=>["Computational biology", "Molecular genetics", "gene expression", "genetics", "Gene splicing", "Molecular cell biology", "RNA processing"], "article_id"=>941315, "categories"=>["Biological Sciences"], "users"=>["Lucie Grodecká", "Pavla Lockerová", "Barbora Ravčuková", "Emanuele Buratti", "Francisco E. Baralle", "Ladislav Dušek", "Tomas Freiberger"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0089570.g002", "stats"=>{"downloads"=>0, "page_views"=>3, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Analyzed_sequences_/941315", "title"=>"Analyzed sequences.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2014-02-21 02:59:01"}
  • {"files"=>["https://ndownloader.figshare.com/files/1394539"], "description"=>"<p>(A) Schematic sequences of mutated acceptor splice sites. Introns are shown in lower-case, and exons are shown in capital letters. Mutated nucleotides are bold and underlined. The PPS are singly underlined and other polypyrimidine stretches are dashed underlined. (B) RT-PCR of minigenes transfected into HeLa cells. cDNA bands originating from O13 mutated minigene are numbered as follows: 1) cryptic 3′ss utilization 81 nt upstream of the authentic splice site (the aberrant exon starts at c.1350-81G), 2) normally spliced RNA, 3) skipping of mutated exon. (C) RT-PCR from RNA extracted from patients’ blood. P = patient’s sample, HC = healthy control sample. The O13 cDNA bands are numbered as in (B).</p>", "links"=>[], "tags"=>["Computational biology", "Molecular genetics", "gene expression", "genetics", "Gene splicing", "Molecular cell biology", "RNA processing", "mutations", "o13", "hk08"], "article_id"=>941314, "categories"=>["Biological Sciences"], "users"=>["Lucie Grodecká", "Pavla Lockerová", "Barbora Ravčuková", "Emanuele Buratti", "Francisco E. Baralle", "Ladislav Dušek", "Tomas Freiberger"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0089570.g001", "stats"=>{"downloads"=>0, "page_views"=>13, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Analysis_of_the_two_E_1_mutations_of_the_BTK_gene_O13_c_1482G_T_and_HK08_c_1883G_A_/941314", "title"=>"Analysis of the two E<sup>+1</sup> mutations of the <i>BTK</i> gene, O13 (c.1482G>T) and HK08 (c.1883G>A).", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2014-02-21 02:59:01"}
  • {"files"=>["https://ndownloader.figshare.com/files/1394547"], "description"=>"<p>The BS and PPT parameters were predicted using prediction programs provided by Kol et al. <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0089570#pone.0089570-Kol1\" target=\"_blank\">[10]</a> or Schwartz et al. <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0089570#pone.0089570-Schwartz1\" target=\"_blank\">[11]</a> using the Sroogle engine. Computational predictions and other sequence parameters for an experimentally confirmed test set of splicing-affecting and non-affecting G<sup>+1</sup> mutations were subjected to statistical analysis using the Mann-Whitney test (see Table S1 <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0089570#pone.0089570.s006\" target=\"_blank\">File S1</a> for details). Significant differences (p<0.05) are marked in bold. dist. = distance, BS = branch point site, M-W test = Mann-Whitney test, perc. = percentile, PPS = the longest uninterrupted polypyrimidine stretch Py = number of pyrimidines (in 25 or 50 nt upstream from 3′ss).</p>", "links"=>[], "tags"=>["Computational biology", "Molecular genetics", "gene expression", "genetics", "Gene splicing", "Molecular cell biology", "RNA processing", "ranges", "describing", "intronic"], "article_id"=>941322, "categories"=>["Biological Sciences"], "users"=>["Lucie Grodecká", "Pavla Lockerová", "Barbora Ravčuková", "Emanuele Buratti", "Francisco E. Baralle", "Ladislav Dušek", "Tomas Freiberger"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0089570.t002", "stats"=>{"downloads"=>0, "page_views"=>6, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Comparison_of_value_ranges_describing_particular_intronic_parameters_in_the_test_set_of_G_1_mutations_/941322", "title"=>"Comparison of value ranges describing particular intronic parameters in the test set of G<sup>+1</sup> mutations.", "pos_in_sequence"=>0, "defined_type"=>3, "published_date"=>"2014-02-21 02:59:01"}
  • {"files"=>["https://ndownloader.figshare.com/files/1394541"], "description"=>"<p>RT-PCR analysis of the literature-derived E<sup>+1</sup> variations. The splicing affecting sequences are underlined. (A) The test set sequences. cDNA bands originating from <i>BTK</i> exon 10 mutated minigene are numbered as follows: 1) cryptic 3′ss utilization 31 nt upstream of the authentic splice site (the aberrant exon starts at c.840-31G), 2) normally spliced RNA. (B) The borderline set sequences.</p>", "links"=>[], "tags"=>["Computational biology", "Molecular genetics", "gene expression", "genetics", "Gene splicing", "Molecular cell biology", "RNA processing", "splicing", "minigene"], "article_id"=>941316, "categories"=>["Biological Sciences"], "users"=>["Lucie Grodecká", "Pavla Lockerová", "Barbora Ravčuková", "Emanuele Buratti", "Francisco E. Baralle", "Ladislav Dušek", "Tomas Freiberger"], "doi"=>"https://dx.doi.org/10.1371/journal.pone.0089570.g003", "stats"=>{"downloads"=>1, "page_views"=>8, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Results_of_the_splicing_minigene_analyses_/941316", "title"=>"Results of the splicing minigene analyses.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2014-02-21 02:59:01"}

PMC Usage Stats | Further Information

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Relative Metric

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