Allele-specific silencing of a dominant SETX mutation in familial amyotrophic lateral sclerosis type 4
Summary: Amyotrophic lateral sclerosis 4 (ALS4) is an autosomal dominant motor neuron disease that is molecularly characterized by reduced R-loop levels and caused by pathogenic variants in senataxin (SETX). SETX encodes an RNA/DNA helicase that resolves three-stranded nucleic acid structures called...
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Elsevier
2025-07-01
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| Series: | HGG Advances |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2666247725000387 |
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| author | Audrey Winkelsas Athena Apfel Brian Johnson George Harmison Kimberly Diaz Perez Dongjun Li Vivian G. Cheung Christopher Grunseich |
| author_facet | Audrey Winkelsas Athena Apfel Brian Johnson George Harmison Kimberly Diaz Perez Dongjun Li Vivian G. Cheung Christopher Grunseich |
| author_sort | Audrey Winkelsas |
| collection | DOAJ |
| description | Summary: Amyotrophic lateral sclerosis 4 (ALS4) is an autosomal dominant motor neuron disease that is molecularly characterized by reduced R-loop levels and caused by pathogenic variants in senataxin (SETX). SETX encodes an RNA/DNA helicase that resolves three-stranded nucleic acid structures called R-loops. Currently, there are no disease-modifying therapies available for ALS4. Given that SETX is haplosufficient, removing the product of the mutated allele presents a potential therapeutic strategy. We designed a series of siRNAs to selectively target the RNA transcript from the ALS4 allele containing the c.1166T>C mutation (p.Leu389Ser). Transfection of HEK293 cells with siRNA and plasmids encoding either wild-type or mutant (Leu389Ser) epitope-tagged SETX revealed that three siRNAs specifically reduced mutant SETX protein levels while having minimal effect on the wild-type SETX protein. In ALS4 primary fibroblasts, siRNA treatment silenced the endogenous mutant SETX allele while sparing the wild-type allele and restored R-loop levels in patient cells. Our findings demonstrate that mutant SETX, differing from wild-type by a single nucleotide, can be effectively and specifically silenced by RNA interference. |
| format | Article |
| id | doaj-art-35ddd5b001804ef38c9b1fc5cc34ceea |
| institution | OA Journals |
| issn | 2666-2477 |
| language | English |
| publishDate | 2025-07-01 |
| publisher | Elsevier |
| record_format | Article |
| series | HGG Advances |
| spelling | doaj-art-35ddd5b001804ef38c9b1fc5cc34ceea2025-08-20T02:12:16ZengElsevierHGG Advances2666-24772025-07-016310043510.1016/j.xhgg.2025.100435Allele-specific silencing of a dominant SETX mutation in familial amyotrophic lateral sclerosis type 4Audrey Winkelsas0Athena Apfel1Brian Johnson2George Harmison3Kimberly Diaz Perez4Dongjun Li5Vivian G. Cheung6Christopher Grunseich7National Institute of Neurological Disorders and Stroke, National Institutes of Health, 35 Convent Dr., Bethesda, MD 20892, USANational Institute of Neurological Disorders and Stroke, National Institutes of Health, 35 Convent Dr., Bethesda, MD 20892, USANational Institute of Neurological Disorders and Stroke, National Institutes of Health, 35 Convent Dr., Bethesda, MD 20892, USANational Institute of Neurological Disorders and Stroke, National Institutes of Health, 35 Convent Dr., Bethesda, MD 20892, USADepartment of Molecular Biology, Cell Biology, and Biochemistry, Brown University, Providence, RI 02912, USADepartment of Pediatrics and Life Sciences Institute, University of Michigan, 210 Washtenaw, Ann Arbor, MI 48109, USADepartment of Pediatrics and Life Sciences Institute, University of Michigan, 210 Washtenaw, Ann Arbor, MI 48109, USANational Institute of Neurological Disorders and Stroke, National Institutes of Health, 35 Convent Dr., Bethesda, MD 20892, USA; Corresponding authorSummary: Amyotrophic lateral sclerosis 4 (ALS4) is an autosomal dominant motor neuron disease that is molecularly characterized by reduced R-loop levels and caused by pathogenic variants in senataxin (SETX). SETX encodes an RNA/DNA helicase that resolves three-stranded nucleic acid structures called R-loops. Currently, there are no disease-modifying therapies available for ALS4. Given that SETX is haplosufficient, removing the product of the mutated allele presents a potential therapeutic strategy. We designed a series of siRNAs to selectively target the RNA transcript from the ALS4 allele containing the c.1166T>C mutation (p.Leu389Ser). Transfection of HEK293 cells with siRNA and plasmids encoding either wild-type or mutant (Leu389Ser) epitope-tagged SETX revealed that three siRNAs specifically reduced mutant SETX protein levels while having minimal effect on the wild-type SETX protein. In ALS4 primary fibroblasts, siRNA treatment silenced the endogenous mutant SETX allele while sparing the wild-type allele and restored R-loop levels in patient cells. Our findings demonstrate that mutant SETX, differing from wild-type by a single nucleotide, can be effectively and specifically silenced by RNA interference.http://www.sciencedirect.com/science/article/pii/S2666247725000387siRNAsenataxinamyotrophic lateral sclerosismotor neuron disease |
| spellingShingle | Audrey Winkelsas Athena Apfel Brian Johnson George Harmison Kimberly Diaz Perez Dongjun Li Vivian G. Cheung Christopher Grunseich Allele-specific silencing of a dominant SETX mutation in familial amyotrophic lateral sclerosis type 4 HGG Advances siRNA senataxin amyotrophic lateral sclerosis motor neuron disease |
| title | Allele-specific silencing of a dominant SETX mutation in familial amyotrophic lateral sclerosis type 4 |
| title_full | Allele-specific silencing of a dominant SETX mutation in familial amyotrophic lateral sclerosis type 4 |
| title_fullStr | Allele-specific silencing of a dominant SETX mutation in familial amyotrophic lateral sclerosis type 4 |
| title_full_unstemmed | Allele-specific silencing of a dominant SETX mutation in familial amyotrophic lateral sclerosis type 4 |
| title_short | Allele-specific silencing of a dominant SETX mutation in familial amyotrophic lateral sclerosis type 4 |
| title_sort | allele specific silencing of a dominant setx mutation in familial amyotrophic lateral sclerosis type 4 |
| topic | siRNA senataxin amyotrophic lateral sclerosis motor neuron disease |
| url | http://www.sciencedirect.com/science/article/pii/S2666247725000387 |
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