Regulating PCCA gene expression by modulation of pseudoexon splicing patterns to rescue enzyme activity in propionic acidemia
Pseudoexons are nonfunctional intronic sequences that can be activated by deep-intronic sequence variation. Activation increases pseudoexon inclusion in mRNA and interferes with normal gene expression. The PCCA c.1285-1416A>G variation activates a pseudoexon and causes the severe metabolic disord...
| Main Authors: | , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2024-03-01
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| Series: | Molecular Therapy: Nucleic Acids |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2162253123003190 |
| _version_ | 1797378787939188736 |
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| author | Ulrika Simone Spangsberg Petersen Maja Dembic Ainhoa Martínez-Pizarro Eva Richard Lise Lolle Holm Jesper Foged Havelund Thomas Koed Doktor Martin Røssel Larsen Nils J. Færgeman Lourdes Ruiz Desviat Brage Storstein Andresen |
| author_facet | Ulrika Simone Spangsberg Petersen Maja Dembic Ainhoa Martínez-Pizarro Eva Richard Lise Lolle Holm Jesper Foged Havelund Thomas Koed Doktor Martin Røssel Larsen Nils J. Færgeman Lourdes Ruiz Desviat Brage Storstein Andresen |
| author_sort | Ulrika Simone Spangsberg Petersen |
| collection | DOAJ |
| description | Pseudoexons are nonfunctional intronic sequences that can be activated by deep-intronic sequence variation. Activation increases pseudoexon inclusion in mRNA and interferes with normal gene expression. The PCCA c.1285-1416A>G variation activates a pseudoexon and causes the severe metabolic disorder propionic acidemia by deficiency of the propionyl-CoA carboxylase enzyme encoded by PCCA and PCCB. We characterized this pathogenic pseudoexon activation event in detail and identified hnRNP A1 to be important for normal repression. The PCCA c.1285-1416A>G variation disrupts an hnRNP A1-binding splicing silencer and simultaneously creates a splicing enhancer. We demonstrate that blocking this region of regulation with splice-switching antisense oligonucleotides restores normal splicing and rescues enzyme activity in patient fibroblasts and in a cellular model created by CRISPR gene editing. Interestingly, the PCCA pseudoexon offers an unexploited potential to upregulate gene expression because healthy tissues show relatively high inclusion levels. By blocking inclusion of the nonactivated wild-type pseudoexon, we can increase both PCCA and PCCB protein levels, which increases the activity of the heterododecameric enzyme. Surprisingly, we can increase enzyme activity from residual levels in not only patient fibroblasts harboring PCCA missense variants but also those harboring PCCB missense variants. This is a potential treatment strategy for propionic acidemia. |
| first_indexed | 2024-03-08T20:12:40Z |
| format | Article |
| id | doaj.art-ff8b723093454117929845437197aef9 |
| institution | Directory Open Access Journal |
| issn | 2162-2531 |
| language | English |
| last_indexed | 2024-03-08T20:12:40Z |
| publishDate | 2024-03-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Molecular Therapy: Nucleic Acids |
| spelling | doaj.art-ff8b723093454117929845437197aef92023-12-23T05:20:33ZengElsevierMolecular Therapy: Nucleic Acids2162-25312024-03-01351102101Regulating PCCA gene expression by modulation of pseudoexon splicing patterns to rescue enzyme activity in propionic acidemiaUlrika Simone Spangsberg Petersen0Maja Dembic1Ainhoa Martínez-Pizarro2Eva Richard3Lise Lolle Holm4Jesper Foged Havelund5Thomas Koed Doktor6Martin Røssel Larsen7Nils J. Færgeman8Lourdes Ruiz Desviat9Brage Storstein Andresen10Department of Biochemistry and Molecular Biology and the Villum Center for Bioanalytical Sciences, University of Southern Denmark, 5230 Odense M, DenmarkDepartment of Biochemistry and Molecular Biology and the Villum Center for Bioanalytical Sciences, University of Southern Denmark, 5230 Odense M, Denmark; Department of Clinical Genetics, Odense University Hospital, 5000 Odense C, Denmark; Department of Mathematics and Computer Science, University of Southern Denmark, 5230 Odense M, DenmarkCentro de Biología Molecular Severo Ochoa, UAM-CSIC, CEDEM, CIBERER, IdiPaz, Universidad Autónoma de Madrid, 28049 Madrid, SpainCentro de Biología Molecular Severo Ochoa, UAM-CSIC, CEDEM, CIBERER, IdiPaz, Universidad Autónoma de Madrid, 28049 Madrid, SpainDepartment of Biochemistry and Molecular Biology and the Villum Center for Bioanalytical Sciences, University of Southern Denmark, 5230 Odense M, DenmarkDepartment of Biochemistry and Molecular Biology and the Villum Center for Bioanalytical Sciences, University of Southern Denmark, 5230 Odense M, DenmarkDepartment of Biochemistry and Molecular Biology and the Villum Center for Bioanalytical Sciences, University of Southern Denmark, 5230 Odense M, DenmarkDepartment of Biochemistry and Molecular Biology and the Villum Center for Bioanalytical Sciences, University of Southern Denmark, 5230 Odense M, DenmarkDepartment of Biochemistry and Molecular Biology and the Villum Center for Bioanalytical Sciences, University of Southern Denmark, 5230 Odense M, DenmarkCentro de Biología Molecular Severo Ochoa, UAM-CSIC, CEDEM, CIBERER, IdiPaz, Universidad Autónoma de Madrid, 28049 Madrid, SpainDepartment of Biochemistry and Molecular Biology and the Villum Center for Bioanalytical Sciences, University of Southern Denmark, 5230 Odense M, Denmark; Corresponding author: Professor Brage Storstein Andresen, Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, 5230 Odense M, Denmark.Pseudoexons are nonfunctional intronic sequences that can be activated by deep-intronic sequence variation. Activation increases pseudoexon inclusion in mRNA and interferes with normal gene expression. The PCCA c.1285-1416A>G variation activates a pseudoexon and causes the severe metabolic disorder propionic acidemia by deficiency of the propionyl-CoA carboxylase enzyme encoded by PCCA and PCCB. We characterized this pathogenic pseudoexon activation event in detail and identified hnRNP A1 to be important for normal repression. The PCCA c.1285-1416A>G variation disrupts an hnRNP A1-binding splicing silencer and simultaneously creates a splicing enhancer. We demonstrate that blocking this region of regulation with splice-switching antisense oligonucleotides restores normal splicing and rescues enzyme activity in patient fibroblasts and in a cellular model created by CRISPR gene editing. Interestingly, the PCCA pseudoexon offers an unexploited potential to upregulate gene expression because healthy tissues show relatively high inclusion levels. By blocking inclusion of the nonactivated wild-type pseudoexon, we can increase both PCCA and PCCB protein levels, which increases the activity of the heterododecameric enzyme. Surprisingly, we can increase enzyme activity from residual levels in not only patient fibroblasts harboring PCCA missense variants but also those harboring PCCB missense variants. This is a potential treatment strategy for propionic acidemia.http://www.sciencedirect.com/science/article/pii/S2162253123003190MT: Oligonucleotides: Therapies and Applicationspre-mRNA splicingcryptic splicingpseudoexonsplice-switching antisense oligonucleotidesPCCA |
| spellingShingle | Ulrika Simone Spangsberg Petersen Maja Dembic Ainhoa Martínez-Pizarro Eva Richard Lise Lolle Holm Jesper Foged Havelund Thomas Koed Doktor Martin Røssel Larsen Nils J. Færgeman Lourdes Ruiz Desviat Brage Storstein Andresen Regulating PCCA gene expression by modulation of pseudoexon splicing patterns to rescue enzyme activity in propionic acidemia Molecular Therapy: Nucleic Acids MT: Oligonucleotides: Therapies and Applications pre-mRNA splicing cryptic splicing pseudoexon splice-switching antisense oligonucleotides PCCA |
| title | Regulating PCCA gene expression by modulation of pseudoexon splicing patterns to rescue enzyme activity in propionic acidemia |
| title_full | Regulating PCCA gene expression by modulation of pseudoexon splicing patterns to rescue enzyme activity in propionic acidemia |
| title_fullStr | Regulating PCCA gene expression by modulation of pseudoexon splicing patterns to rescue enzyme activity in propionic acidemia |
| title_full_unstemmed | Regulating PCCA gene expression by modulation of pseudoexon splicing patterns to rescue enzyme activity in propionic acidemia |
| title_short | Regulating PCCA gene expression by modulation of pseudoexon splicing patterns to rescue enzyme activity in propionic acidemia |
| title_sort | regulating pcca gene expression by modulation of pseudoexon splicing patterns to rescue enzyme activity in propionic acidemia |
| topic | MT: Oligonucleotides: Therapies and Applications pre-mRNA splicing cryptic splicing pseudoexon splice-switching antisense oligonucleotides PCCA |
| url | http://www.sciencedirect.com/science/article/pii/S2162253123003190 |
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