Figure 4

Figure 4 mRNA injections of SMN1 hypomorphic variants associated with SMA type II and III only partially rescue the phenotype of smn^−/− zebrafish, supporting the method’s ability to detect hypomorphic variants.

(A) Details of the SMN1 variants selected for further analysis. Eight additional variants were evaluated: two associated with SMA type II, four with SMA type III, one additional known pathogenic variant (path 2), and one additional known non-pathogenic variant (non-path 2). All variants were selected to be specifically associated with patients carrying a single SMN2 copy, providing a uniform genetic background and avoiding confounding effects related to SMN2 dosage. (B) Morphological malformations consistent with SMA-hallmarks were observed in smn^−/− embryos injected with all six SMN1 type II/III variants, although these appeared later or in milder forms than in mock-injected negative controls or embryos injected with SMA Type I pathogenic variants SMN1 c.549del (path 1) and SMN1 c43C>T (p.Gln15*) (path 2). In contrast, mRNA encoding non-pathogenic SMN1 variants, SMN1 c.462 A > G (non-path 1) and SMN1 c.84 C > T (non-path 2) as well as the two VUS (861VUS and 855VUS), fully rescued the smn^−/− phenotype, showing no difference from the wt-SMN1 mRNA positive control. These results provide further evidence of functional SMN protein expression and confirm the non-pathogenic nature of the tested VUS. Empty boxes indicate all larvae are dead. Scale bar, 500 µm. Source data are available online for this figure.