A novel variant in the 3′ UTR of human SCN1A gene from a patient with Dravet syndrome decreases mRNA stability mediated by GAPDH’s binding

Abstract

Mutations in the SCN1A gene-encoding voltage-gated sodium channel α-I subunit (Nav1.1) cause various spectrum of epilepsies including Dravet syndrome (DS), a severe and intractable form. A large number of SCN1A mutations identified from the DS patients lead to the loss of function or truncation of Nav1.1 that result in a haploinsufficiency effects, indicating that the exact expression level of SCN1A should be essential to maintain normal brain function. In this study, we have identified five variants c.*1025T>C, c.*1031A>T, c.*1739C>T, c.*1794C>T and c.*1961C>T in the SCN1A 3' UTR in the patients with DS. The c.*1025T>C, c.*1031A>T and c.*1794C>T are conserved among different species. Of all the five variants, only c.*1794C>T is a novel variant and alters the predicted secondary structure of the 3' UTR. We also show that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) only binds to the 3' UTR sequence containing the mutation allele 1794U but not the wild-type allele 1794C, indicating that the mutation allele forms a new GAPDH-binding site. Functional analyses show that the variant negatively regulates the reporter gene expression by affecting the mRNA stability that is mediated by GAPDH's binding, and this phenomenon could be reversed by shRNA-induced GAPDH knockdown. These findings suggest that GAPDH and the 3'-UTR variant are involved in regulating SCN1A expression at post-transcriptional level, which may provide an important clue for further investigating on the relationship between 3'-UTR variants and SCN1A-related diseases.

The sweet side ofRNAregulation: glyceraldehyde‐3‐phosphate dehydrogenase as a noncanonicalRNA‐binding protein

“…The C1794U mutation in the 3′ UTR of SCN1A mRNA was shown to affect protein expression and mRNA stability, possibly via changes in predicted mRNA secondary structures. GAPDH was shown to selectively bind to the U‐rich mutant 3′ UTR and to negatively regulate protein expression and mRNA stability …”

“…Third, GAPDH sequesters Rheb and prevents mTORc1 activation . Finally, in ketogenic diets, GAPDH may be free from glycolysis and able to destabilize the SCN1A mRNA to reduce translation of the sodium transporter NaV 1.1, and alleviate conditions linked to Dravet syndrome …”

Mosaicism of de novo pathogenic SCN1A variants in epilepsy is a frequent phenomenon that correlates with variable phenotypes

“…These variants are virtually always associated with severe phenotypes [8][9][10] ; 97.7% of genomic rearrangements and splice site, nonsense, and frameshift SCN1A variants are mainly associated with Dravet syndrome. 13,[17][18][19][20] Moreover, parental mosaicism for the pathogenic SCN1A variant has been well recognized in cases were mosaic parents of Dravet children show a mild epilepsy phenotype. Functional studies have shown varying degrees of loss of function through a lack of sodium current when pathogenic variants are located in critical regions of the gene (voltage sensor and/or ion-pore regions).…”

Is RNA Binding in the Rossmann Fold Essential for GAPDH Intranuclear Functions?

“…GAPDH-RNA binding alters mRNA stability (Bonafe et al, 2005;Ikeda et al, 2012;Zeng et al, 2014). Several reports suggest that GAPDH binding to RNA may play a role in nucleocytoplasmic transport of RNA (Singh and Green, 1993;Muller et al, 1992;Hamilton et al, 1993).…”

“…Three types of polynucleotide-GAPDH interaction have been described, so far: (i) GAPDH binding to RNA, mostly represented by AU-Rich Elements (ARE) in the 3'-UTR of mRNA (Nagy and Rigby, 1995;Seidler, 2013) (ii) formation of multiprotein-RNA complexes, with GAPDH and RNA being important structural components (Mazurek et al, 1996;Carlile et al, 1998) (iii) GAPDH binding to single-stranded DNA at telomeric sequences near the chromosomal ends (Sundararaj et al, 2004;Demarse et al, 2009). GAPDH-RNA binding affects mRNA stability and template activity (Bonafe et al, 2005;Rodriguez-Pascual et al, 2008;Backlund et al, 2009;Kondo et al, 2011;Ikeda et al, 2012;Zeng et al, 2014). RNA binding activity of GAPDH is attributed to the Rossmann fold which forms NAD + -binding center located at the Nterminus of GAPDH polypeptide (Rossman, 1981).…”