A super minigene with a short promoter and truncated introns recapitulates essential features of transcription and splicing regulation of the SMN1 and SMN2 genes.
Nucleic Acids Res
; 52(7): 3547-3571, 2024 Apr 24.
Article
em En
| MEDLINE
| ID: mdl-38214229
ABSTRACT
Here we report a Survival Motor Neuron 2 (SMN2) super minigene, SMN2Sup, encompassing its own promoter, all exons, their flanking intronic sequences and the entire 3'-untranslated region. We confirm that the pre-mRNA generated from SMN2Sup undergoes splicing to produce a translation-competent mRNA. We demonstrate that mRNA generated from SMN2Sup produces more SMN than an identical mRNA generated from a cDNA clone. We uncover that overexpression of SMN triggers skipping of exon 3 of SMN1/SMN2. We define the minimal promoter and regulatory elements associated with the initiation and elongation of transcription of SMN2. The shortened introns within SMN2Sup preserved the ability of camptothecin, a transcription elongation inhibitor, to induce skipping of exons 3 and 7 of SMN2. We show that intron 1-retained transcripts undergo nonsense-mediated decay. We demonstrate that splicing factor SRSF3 and DNA/RNA helicase DHX9 regulate splicing of multiple exons in the context of both SMN2Sup and endogenous SMN1/SMN2. Prevention of SMN2 exon 7 skipping has implications for the treatment of spinal muscular atrophy (SMA). We validate the utility of the super minigene in monitoring SMN levels upon splicing correction. Finally, we demonstrate how the super minigene could be employed to capture the cell type-specific effects of a pathogenic SMN1 mutation.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Transcrição Gênica
/
Íntrons
/
Éxons
/
Regiões Promotoras Genéticas
/
Proteína 1 de Sobrevivência do Neurônio Motor
/
Proteína 2 de Sobrevivência do Neurônio Motor
Limite:
Humans
Idioma:
En
Revista:
Nucleic Acids Res
Ano de publicação:
2024
Tipo de documento:
Article
País de afiliação:
Estados Unidos