Cystic fibrosis (CF) is a common recessive disorder caused by >1600 mutations in the CF transmembrane conductance regulator (CFTR) gene. About 13% of CFTR mutations are classified as “splicing mutations,” but for almost 40% of these, their role in affecting the pre-mRNA splicing of the gene is not yet defined. In this work, we describe a new splicing mutation detected in three unrelated Italian CF patients. By DNA analyses and mRNA studies, we identified the c.1002–1110_1113delTAAG mutation localized in intron 6b of the CFTR gene. At the mRNA level, this mutation creates an aberrant inclusion of a sequence of 101 nucleotides between exons 6b and 7. This sequence corresponds to a portion of intron 6b and resembles a cryptic exon because it is characterized by an upstream ag and a downstream gt sequence, which are most probably recognized as 5′- and 3′-splice sites by the spliceosome. Through functional analysis of this splicing defect, we show that this mutation abolishes the interaction of the splicing regulatory protein heterogeneous nuclear ribonucleoprotein A2/B1 with an intronic splicing regulatory element and creates a new recognition motif for the SRp75 splicing factor, causing activation of the cryptic exon. Our results show that the c.1002–1110_1113delTAAG mutation creates a new intronic splicing regulatory element in intron 6b of the CFTR gene exclusively recognized by SRp75. Cystic fibrosis (CF) is a common recessive disorder caused by >1600 mutations in the CF transmembrane conductance regulator (CFTR) gene. About 13% of CFTR mutations are classified as “splicing mutations,” but for almost 40% of these, their role in affecting the pre-mRNA splicing of the gene is not yet defined. In this work, we describe a new splicing mutation detected in three unrelated Italian CF patients. By DNA analyses and mRNA studies, we identified the c.1002–1110_1113delTAAG mutation localized in intron 6b of the CFTR gene. At the mRNA level, this mutation creates an aberrant inclusion of a sequence of 101 nucleotides between exons 6b and 7. This sequence corresponds to a portion of intron 6b and resembles a cryptic exon because it is characterized by an upstream ag and a downstream gt sequence, which are most probably recognized as 5′- and 3′-splice sites by the spliceosome. Through functional analysis of this splicing defect, we show that this mutation abolishes the interaction of the splicing regulatory protein heterogeneous nuclear ribonucleoprotein A2/B1 with an intronic splicing regulatory element and creates a new recognition motif for the SRp75 splicing factor, causing activation of the cryptic exon. Our results show that the c.1002–1110_1113delTAAG mutation creates a new intronic splicing regulatory element in intron 6b of the CFTR gene exclusively recognized by SRp75. Cystic fibrosis (CF 2The abbreviations used are: CFcystic fibrosisCFTRCF transmembrane conductance regulatorhnRNPheterogeneous nuclear ribonucleoproteinHPLChigh pressure liquid chromatographyRTreverse transcriptionMALDImatrix-assisted laser desorption ionizationsiRNAsmall interfering RNAWTwild-type. 2The abbreviations used are: CFcystic fibrosisCFTRCF transmembrane conductance regulatorhnRNPheterogeneous nuclear ribonucleoproteinHPLChigh pressure liquid chromatographyRTreverse transcriptionMALDImatrix-assisted laser desorption ionizationsiRNAsmall interfering RNAWTwild-type.; MIM entry #219700) is the most frequent severe autosomic recessive disorder in the European population (1Welsh M.J. Tsui L.C. Boat T.F. Beaudet A.I. Cystic Fibrosis: The Metabolic and Molecular Bases of Inherited Disease.in: Scriver C.R. Beaudet A.I. Sly W.S. Valle D. McGraw-Hill Book Co., New York1995: 3799-3876Google Scholar). Indeed, CF affects ∼1 in 2500 births, and ∼1 in 25 individuals are heterozygotes, with marked regional variations (2Estivill X. Bancells C. Ramos C. Hum. Mutat. 1997; 10: 135-154Crossref PubMed Scopus (267) Google Scholar). CF is caused by mutations of the CF transmembrane conductance regulator (CFTR or ABCC7; MIM #602421) gene, which is also involved in a broad spectrum of phenotypes, including male infertility due to congenital bilateral absence of the vas deferens (3Chillón M. Casals T. Mercier B. Brassas L. Lissens W. Silber S. Romey M.C. Ruiz-Romero J. Verlingue C. Claustres M. N. Engl. J. Med. 1995; 332: 1475-1480Crossref PubMed Scopus (808) Google Scholar, 4Costes B. Girodon E. Ghanem N. Flori E. Jardin A. Soufir J.C. Goossens M. Eur. J. Hum. Genet. 1995; 3: 285-293Crossref PubMed Scopus (138) Google Scholar), disseminated bronchiectasis (5Pignatti P.F. Bombieri C. Marigo C. Benetazzo M. Luisetti M. Hum. Mol. Genet. 1995; 4: 635-639Crossref PubMed Scopus (152) Google Scholar, 6Girodon E. Cazeneuve C. Lebargy F. Chinet T. Costes B. Ghanem N. Martin J. Lemay S. Scheid P. Housset B. Bignon J. Goossens M. Eur. J. Hum. Genet. 1997; 5: 149-155Crossref PubMed Scopus (150) Google Scholar), and chronic pancreatitis (7Cohn J.A. Friedman K.J. Noone P.G. Knowles M.R. Silverman L.M. Jowel P.S. N. Engl. J. Med. 1998; 339: 653-658Crossref PubMed Scopus (802) Google Scholar, 8Sharer N. Schwarz M. Malone G. Howarth A. Painter J. Super M. Brazanga J. N. Engl. J. Med. 1998; 339: 645-652Crossref PubMed Scopus (795) Google Scholar). The mutational spectrum of this disease is made up of >1600 different mutations, 98% of which consist of point mutations or microdeletions/insertions (Cystic Fibrosis Mutation Database). A significant fraction (∼13%) of CFTR mutations are classified as “splicing mutations,” but for almost 40% of these, their role in affecting the pre-mRNA splicing of the gene is not yet defined.Splicing mutations act by directly disrupting splice sites or by creating new ones. Varied levels of alternative splicing have been detected for some of the splicing mutations (9Zielenski J. Patrizio P. Corey M. Handelin B. Markiewicz D. Asch R. Tsui L.C. Am. J. Hum. Genet. 1995; 57: 958-960PubMed Google Scholar, 10Chiba-Falek O. Kerem E. Shoshani T. Aviram M. Augarten A. Bentur L. Tal A. Tullis E. Rahat A. Kerem B. Genomics. 1998; 53: 276-283Crossref PubMed Scopus (72) Google Scholar, 11Chiba-Falek O. Parad R.B. Kerem E. Kerem B. Am. J. Respir. Crit. Care Med. 1999; 159: 1998-2002Crossref PubMed Scopus (32) Google Scholar). These differences are most likely due to different utilization of intragenic splicing elements by trans-acting splicing factors (12Nissim-Rafinia M. Chiba-Falek O. Sharon G. Boss A. Kerem B. Hum. Mol. Genet. 2000; 9: 1771-1778Crossref PubMed Scopus (56) Google Scholar, 13Pagani F. Buratti E. Stuani C. Romano M. Zuccato E. Niksic M. Giglio L. Faraguna D. Baralle F.E. J. Biol. Chem. 2000; 275: 21041-21047Abstract Full Text Full Text PDF PubMed Scopus (152) Google Scholar). cis-Acting elements required for splicing in mammalian cells include short and poorly conserved consensus sequences, which specify the 5′-splice site, branch site, and 3′-splice site. The 3′-splice site is preceded by a polypyrimidine tract of variable length.The branch point is typically located 18–40 nucleotides upstream of the polypyrimidine tract. These splicing signals are essential but do not contain sufficient information to specify correct splicing patterns, and they alone cannot be distinguished from numerous surrounding cryptic splice sites of similar strength (14Cartegni L. Chew S.L. Krainer A.R. Nat. Rev. Genet. 2002; 3: 285-298Crossref PubMed Scopus (1726) Google Scholar). Their efficient recognition, as well as their regulated usage, requires sequences within exons and introns called enhancers and silencers. Pre-mRNA splicing requires the interaction of these critical cis-acting sequences with splicing regulating factors that form the spliceosome. The spliceosome is a highly complex RNA/protein machine consisting of five small nuclear RNAs and >100 proteins such as the SR (serine/arginine-rich) proteins and heterogeneous nuclear ribonucleoproteins (hnRNPs), allowing the fine-tuning of the splicing process (15Graveley B.R. RNA. 2000; 6: 1197-1211Crossref PubMed Scopus (875) Google Scholar, 16Nilsen T.W. BioEssays. 2003; 25: 1147-1149Crossref PubMed Scopus (281) Google Scholar, 17Zhou Z. Licklider L.J. Gygi S.P. Reed R. Nature. 2002; 419: 182-185Crossref PubMed Scopus (717) Google Scholar).In classical splicing defects, the cis-acting elements are mutated, and the effect on splicing can be predicted on the basis of genomic DNA sequence analysis. However, the effect of mutations in non-obvious regulatory elements is more difficult to evaluate directly from genome scanning methodologies. Patient CFTR mRNA analysis represents a good tool for the identification and definition of the pathogenic role of sequence variations not yet defined and splicing defects causing alternative products.In this work, we describe three unrelated Italian patients affected by CF whose CFTR mRNA contains an insertion of 101 nucleotides between exons 6b and 7 corresponding to a portion of intron 6b. This sequence resembles a cryptic exon because it is characterized by an upstream ag and a downstream gt sequence, which are most probably recognized as 5′- and 3′-splice sites by the spliceosome. We assume that this anomalous transcript is caused by a mutation affecting an intronic splicing regulatory element that either abolishes or creates specific protein-binding domains, causing an alteration of the intronic splicing process. To elucidate the mechanisms involved in the aberrant splicing, we carried out molecular analysis of intron 6b as well as functional studies.DISCUSSIONCF is a common recessive disorder caused by >1600 mutations in the CFTR gene. Despite the extensive mutation characterization work around the world, there is still a substantial fraction of patients with typical clinical diagnosis of CF but unknown mutations in the CFTR gene (28Groman J.D. Meyer M.E. Wilmott R.W. Zeitlin P.L. Cutting G.R. N. Engl. J. Med. 2002; 347: 401-407Crossref PubMed Scopus (158) Google Scholar). About 13% of CFTR mutations are termed as affecting the pre-mRNA splicing, but for almost 40% of these, their role in affecting the pre-mRNA splicing is not yet defined. Genotype screening in the CFTR gene frequently sequence variations whose with the disease is In the effect of an is difficult to this CFTR mRNA analysis represents a good tool for the identification and definition of the pathogenic role of sequence variations not yet defined and splicing defects causing alternative products.In this we have a new splicing mutation detected in three unrelated Italian CF patients. The molecular characterization of these patients carried out either the DNA or The splicing abolishes the interaction of A2/B1 with an intronic splicing regulatory element and creates a new recognition motif for the SRp75 splicing factor, causing the activation of a cryptic exon. To this is the that the SR splicing protein SRp75 been directly in the of a cryptic exon in the CFTR proteins (15Graveley B.R. RNA. 2000; 6: 1197-1211Crossref PubMed Scopus (875) Google are typically involved in of splicing, splicing by with or intronic splicing In is most frequently by G. N. Nat. Rev. Mol. Biol. 2002; 3: PubMed Scopus Google Scholar), which by sequences as or intronic splicing silencers. have been identified as splicing these, the protein been The splicing identified in of exon Mol. Biol. 1995; Scopus Google Scholar, A. A. Mol. Biol. 1995; Scopus (150) Google Scholar). This splicing to and mutations disrupting the splicing and exon the in the genome of sequences between and nucleotides in with 5′- and 3′-splice sites either their inclusion not to be a frequent the pre-mRNA that of these contain defects in their good with sequences Mol. Biol. 2000; PubMed Scopus Google Scholar). In with three splicing have been to be in these with their surrounding intronic Mol. Biol. 2000; PubMed Scopus Google Scholar). This is in with a that that DNA is in sequences that can splicing M. G. L. R. N. R. PubMed Scopus Google intronic sequences to be to exons because most of the inclusion from a This mutation the of a or site, by the of or site However, mechanisms that the inclusion of mutations also the or of splicing regulatory are in the A S. S. R. Am. J. Hum. Genet. 2002; Full Text Full Text PDF PubMed Scopus Google and F. Buratti E. Stuani C. R. T. Baralle F.E. Nat. Genet. 2002; PubMed Scopus Google in which the of splicing regulatory elements been to In the gene, the inclusion of a is the of a in intron that a small nuclear site located within the sequence This is called the intronic splicing and results in the activation of an upstream cryptic site and of a downstream cryptic splicing mutation affecting the CFTR gene in this work to the splicing in the gene by F. Buratti E. Stuani C. R. T. Baralle F.E. Nat. Genet. 2002; PubMed Scopus Google Scholar). In results that the c.1002–1110_1113delTAAG mutation creates a new intronic splicing regulatory element in intron 6b of the CFTR gene recognized by the splicing SRp75. This as that the inclusion the splicing have that the affecting splicing due to mutation is not and cannot be from in The of an intronic splicing be a common in and to be to the in CF patients and to new Cystic fibrosis (CF 2The abbreviations used are: CFcystic fibrosisCFTRCF transmembrane conductance regulatorhnRNPheterogeneous nuclear ribonucleoproteinHPLChigh pressure liquid chromatographyRTreverse transcriptionMALDImatrix-assisted laser desorption ionizationsiRNAsmall interfering RNAWTwild-type. 2The abbreviations used are: CFcystic fibrosisCFTRCF transmembrane conductance regulatorhnRNPheterogeneous nuclear ribonucleoproteinHPLChigh pressure liquid chromatographyRTreverse transcriptionMALDImatrix-assisted laser desorption ionizationsiRNAsmall interfering RNAWTwild-type.; MIM entry #219700) is the most frequent severe autosomic recessive disorder in the European population (1Welsh M.J. Tsui L.C. Boat T.F. Beaudet A.I. Cystic Fibrosis: The Metabolic and Molecular Bases of Inherited Disease.in: Scriver C.R. Beaudet A.I. Sly W.S. Valle D. McGraw-Hill Book Co., New York1995: 3799-3876Google Scholar). Indeed, CF affects ∼1 in 2500 births, and ∼1 in 25 individuals are heterozygotes, with marked regional variations (2Estivill X. Bancells C. Ramos C. Hum. Mutat. 1997; 10: 135-154Crossref PubMed Scopus (267) Google Scholar). CF is caused by mutations of the CF transmembrane conductance regulator (CFTR or ABCC7; MIM #602421) gene, which is also involved in a broad spectrum of phenotypes, including male infertility due to congenital bilateral absence of the vas deferens (3Chillón M. Casals T. Mercier B. Brassas L. Lissens W. Silber S. Romey M.C. Ruiz-Romero J. Verlingue C. Claustres M. N. Engl. J. Med. 1995; 332: 1475-1480Crossref PubMed Scopus (808) Google Scholar, 4Costes B. Girodon E. Ghanem N. Flori E. Jardin A. Soufir J.C. Goossens M. Eur. J. Hum. Genet. 1995; 3: 285-293Crossref PubMed Scopus (138) Google Scholar), disseminated bronchiectasis (5Pignatti P.F. Bombieri C. Marigo C. Benetazzo M. Luisetti M. Hum. Mol. Genet. 1995; 4: 635-639Crossref PubMed Scopus (152) Google Scholar, 6Girodon E. Cazeneuve C. Lebargy F. Chinet T. Costes B. Ghanem N. Martin J. Lemay S. Scheid P. Housset B. Bignon J. Goossens M. Eur. J. Hum. Genet. 1997; 5: 149-155Crossref PubMed Scopus (150) Google Scholar), and chronic pancreatitis (7Cohn J.A. Friedman K.J. Noone P.G. Knowles M.R. Silverman L.M. Jowel P.S. N. Engl. J. Med. 1998; 339: 653-658Crossref PubMed Scopus (802) Google Scholar, 8Sharer N. Schwarz M. Malone G. Howarth A. Painter J. Super M. Brazanga J. N. Engl. J. Med. 1998; 339: 645-652Crossref PubMed Scopus (795) Google Scholar). The mutational spectrum of this disease is made up of >1600 different mutations, 98% of which consist of point mutations or microdeletions/insertions (Cystic Fibrosis Mutation Database). A significant fraction (∼13%) of CFTR mutations are classified as “splicing mutations,” but for almost 40% of these, their role in affecting the pre-mRNA splicing of the gene is not yet defined. fibrosis CF transmembrane conductance regulator heterogeneous nuclear ribonucleoprotein pressure liquid laser desorption small interfering fibrosis CF transmembrane conductance regulator heterogeneous nuclear ribonucleoprotein pressure liquid laser desorption small interfering mutations act by directly disrupting splice sites or by creating new ones. Varied levels of alternative splicing have been detected for some of the splicing mutations (9Zielenski J. Patrizio P. Corey M. Handelin B. Markiewicz D. Asch R. Tsui L.C. Am. J. Hum. Genet. 1995; 57: 958-960PubMed Google Scholar, 10Chiba-Falek O. Kerem E. Shoshani T. Aviram M. Augarten A. Bentur L. Tal A. Tullis E. Rahat A. Kerem B. Genomics. 1998; 53: 276-283Crossref PubMed Scopus (72) Google Scholar, 11Chiba-Falek O. Parad R.B. Kerem E. Kerem B. Am. J. Respir. Crit. Care Med. 1999; 159: 1998-2002Crossref PubMed Scopus (32) Google Scholar). These differences are most likely due to different utilization of intragenic splicing elements by trans-acting splicing factors (12Nissim-Rafinia M. Chiba-Falek O. Sharon G. Boss A. Kerem B. Hum. Mol. Genet. 2000; 9: 1771-1778Crossref PubMed Scopus (56) Google Scholar, 13Pagani F. Buratti E. Stuani C. Romano M. Zuccato E. Niksic M. Giglio L. Faraguna D. Baralle F.E. J. Biol. Chem. 2000; 275: 21041-21047Abstract Full Text Full Text PDF PubMed Scopus (152) Google Scholar). cis-Acting elements required for splicing in mammalian cells include short and poorly conserved consensus sequences, which specify the 5′-splice site, branch site, and 3′-splice site. The 3′-splice site is preceded by a polypyrimidine tract of variable The branch point is typically located 18–40 nucleotides upstream of the polypyrimidine tract. These splicing signals are essential but do not contain sufficient information to specify correct splicing patterns, and they alone cannot be distinguished from numerous surrounding cryptic splice sites of similar strength (14Cartegni L. Chew S.L. Krainer A.R. Nat. Rev. Genet. 2002; 3: 285-298Crossref PubMed Scopus (1726) Google Scholar). Their efficient recognition, as well as their regulated usage, requires sequences within exons and introns called enhancers and silencers. Pre-mRNA splicing requires the interaction of these critical cis-acting sequences with splicing regulating factors that form the spliceosome. The spliceosome is a highly complex RNA/protein machine consisting of five small nuclear RNAs and >100 proteins such as the SR (serine/arginine-rich) proteins and heterogeneous nuclear ribonucleoproteins (hnRNPs), allowing the fine-tuning of the splicing process (15Graveley B.R. RNA. 2000; 6: 1197-1211Crossref PubMed Scopus (875) Google Scholar, 16Nilsen T.W. BioEssays. 2003; 25: 1147-1149Crossref PubMed Scopus (281) Google Scholar, 17Zhou Z. Licklider L.J. Gygi S.P. Reed R. Nature. 2002; 419: 182-185Crossref PubMed Scopus (717) Google Scholar). In classical splicing defects, the cis-acting elements are mutated, and the effect on splicing can be predicted on the basis of genomic DNA sequence analysis. However, the effect of mutations in non-obvious regulatory elements is more difficult to evaluate directly from genome scanning methodologies. Patient CFTR mRNA analysis represents a good tool for the identification and definition of the pathogenic role of sequence variations not yet defined and splicing defects causing alternative In this work, we describe three unrelated Italian patients affected by CF whose CFTR mRNA contains an insertion of 101 nucleotides between exons 6b and 7 corresponding to a portion of intron 6b. This sequence resembles a cryptic exon because it is characterized by an upstream ag and a downstream gt sequence, which are most probably recognized as 5′- and 3′-splice sites by the spliceosome. We assume that this anomalous transcript is caused by a mutation affecting an intronic splicing regulatory element that either abolishes or creates specific protein-binding domains, causing an alteration of the intronic splicing process. To elucidate the mechanisms involved in the aberrant splicing, we carried out molecular analysis of intron 6b as well as functional is a common recessive disorder caused by >1600 mutations in the CFTR gene. Despite the extensive mutation characterization work around the world, there is still a substantial fraction of patients with typical clinical diagnosis of CF but unknown mutations in the CFTR gene (28Groman J.D. Meyer M.E. Wilmott R.W. Zeitlin P.L. Cutting G.R. N. Engl. J. Med. 2002; 347: 401-407Crossref PubMed Scopus (158) Google Scholar). About 13% of CFTR mutations are termed as affecting the pre-mRNA splicing, but for almost 40% of these, their role in affecting the pre-mRNA splicing is not yet defined. Genotype screening in the CFTR gene frequently sequence variations whose with the disease is In the effect of an is difficult to this CFTR mRNA analysis represents a good tool for the identification and definition of the pathogenic role of sequence variations not yet defined and splicing defects causing alternative products.In this we have a new splicing mutation detected in three unrelated Italian CF patients. The molecular characterization of these patients carried out either the DNA or The splicing abolishes the interaction of A2/B1 with an intronic splicing regulatory element and creates a new recognition motif for the SRp75 splicing factor, causing the activation of a cryptic exon. To this is the that the SR splicing protein SRp75 been directly in the of a cryptic exon in the CFTR proteins (15Graveley B.R. RNA. 2000; 6: 1197-1211Crossref PubMed Scopus (875) Google are typically involved in of splicing, splicing by with or intronic splicing In is most frequently by G. N. Nat. Rev. Mol. Biol. 2002; 3: PubMed Scopus Google Scholar), which by sequences as or intronic splicing silencers. have been identified as splicing these, the protein been The splicing identified in of exon Mol. Biol. 1995; Scopus Google Scholar, A. A. Mol. Biol. 1995; Scopus (150) Google Scholar). This splicing to and mutations disrupting the splicing and exon the in the genome of sequences between and nucleotides in with 5′- and 3′-splice sites either their inclusion not to be a frequent the pre-mRNA that of these contain defects in their good with sequences Mol. Biol. 2000; PubMed Scopus Google Scholar). In with three splicing have been to be in these with their surrounding intronic Mol. Biol. 2000; PubMed Scopus Google Scholar). This is in with a that that DNA is in sequences that can splicing M. G. L. R. N. R. PubMed Scopus Google intronic sequences to be to exons because most of the inclusion from a This mutation the of a or site, by the of or site However, mechanisms that the inclusion of mutations also the or of splicing regulatory are in the A S. S. R. Am. J. Hum. Genet. 2002; Full Text Full Text PDF PubMed Scopus Google and F. Buratti E. Stuani C. R. T. Baralle F.E. Nat. Genet. 2002; PubMed Scopus Google in which the of splicing regulatory elements been to In the gene, the inclusion of a is the of a in intron that a small nuclear site located within the sequence This is called the intronic splicing and results in the activation of an upstream cryptic site and of a downstream cryptic splicing mutation affecting the CFTR gene in this work to the splicing in the gene by F. Buratti E. Stuani C. R. T. Baralle F.E. Nat. Genet. 2002; PubMed Scopus Google Scholar). In results that the c.1002–1110_1113delTAAG mutation creates a new intronic splicing regulatory element in intron 6b of the CFTR gene recognized by the splicing SRp75. This as that the inclusion the splicing have that the affecting splicing due to mutation is not and cannot be from in The of an intronic splicing be a common in and to be to the in CF patients and to new CF is a common recessive disorder caused by >1600 mutations in the CFTR gene. Despite the extensive mutation characterization work around the world, there is still a substantial fraction of patients with typical clinical diagnosis of CF but unknown mutations in the CFTR gene (28Groman J.D. Meyer M.E. Wilmott R.W. Zeitlin P.L. Cutting G.R. N. Engl. J. Med. 2002; 347: 401-407Crossref PubMed Scopus (158) Google Scholar). About 13% of CFTR mutations are termed as affecting the pre-mRNA splicing, but for almost 40% of these, their role in affecting the pre-mRNA splicing is not yet defined. Genotype screening in the CFTR gene frequently sequence variations whose with the disease is In the effect of an is difficult to this CFTR mRNA analysis represents a good tool for the identification and definition of the pathogenic role of sequence variations not yet defined and splicing defects causing alternative In this we have a new splicing mutation detected in three unrelated Italian CF patients. The molecular characterization of these patients carried out either the DNA or The splicing abolishes the interaction of A2/B1 with an intronic splicing regulatory element and creates a new recognition motif for the SRp75 splicing factor, causing the activation of a cryptic exon. To this is the that the SR splicing protein SRp75 been directly in the of a cryptic exon in the CFTR SR proteins (15Graveley B.R. RNA. 2000; 6: 1197-1211Crossref PubMed Scopus (875) Google are typically involved in of splicing, splicing by with or intronic splicing In is most frequently by G. N. Nat. Rev. Mol. Biol. 2002; 3: PubMed Scopus Google Scholar), which by sequences as or intronic splicing silencers. have been identified as splicing these, the protein been The splicing identified in of exon Mol. Biol. 1995; Scopus Google Scholar, A. A. Mol. Biol. 1995; Scopus (150) Google Scholar). This splicing to and mutations disrupting the splicing and exon Despite the in the genome of sequences between and nucleotides in with 5′- and 3′-splice sites either their inclusion not to be a frequent the pre-mRNA that of these contain defects in their good with sequences Mol. Biol. 2000; PubMed Scopus Google Scholar). In with three splicing have been to be in these with their surrounding intronic Mol. Biol. 2000; PubMed Scopus Google Scholar). This is in with a that that DNA is in sequences that can splicing M. G. L. R. N. R. PubMed Scopus Google Scholar). intronic sequences to be to exons because most of the inclusion from a This mutation the of a or site, by the of or site However, mechanisms that the inclusion of mutations also the or of splicing regulatory are in the A S. S. R. Am. J. Hum. Genet. 2002; Full Text Full Text PDF PubMed Scopus Google and F. Buratti E. Stuani C. R. T. Baralle F.E. Nat. Genet. 2002; PubMed Scopus Google in which the of splicing regulatory elements been to In the gene, the inclusion of a is the of a in intron that a small nuclear site located within the sequence This is called the intronic splicing and results in the activation of an upstream cryptic site and of a downstream cryptic site. The splicing mutation affecting the CFTR gene in this work to the splicing in the gene by F. Buratti E. Stuani C. R. T. Baralle F.E. Nat. Genet. 2002; PubMed Scopus Google Scholar). In results that the c.1002–1110_1113delTAAG mutation creates a new intronic splicing regulatory element in intron 6b of the CFTR gene recognized by the splicing SRp75. This as that the inclusion the splicing process. We have that the affecting splicing due to mutation is not and cannot be from in The of an intronic splicing be a common in and to be to the in CF patients and to new We G. and for clinical
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