Fig. 2: Rescue construct of Munc13-1+SynPhy3’UTR drives axonal localization of Munc13-1 mRNAs in cultured Smn KO motoneurons.

a Scheme of three viral rescue constructs expressing Munc13-1 with synonymous 3’UTR (wtMunc13-1), the Synaptophysin (SynPhy) 3’UTR, or no 3’UTR (Δ3’UTR). b qRT-PCR reveals that the 3’UTR of Munc13-1 (***P = 0.0003, ***P = 0.0006 from n = 7 biological replicates) and the 3’UTR of SynPhy (*P = 0.05, *P = 0.0286 from n = 3 biological replicates) drive Munc13-1 transcripts into distal axons in wt motoneurons, while Munc13-1 construct lacking the 3’UTR does not (*P = 0.0143 from n = 4 biological replicates). c Immunoblot shows increased Munc13-1 protein levels in total lysates from motoneurons transduced with viral constructs shown in (a, b) (representative of n = 3 biological replicates). d Representative images of smFISH in the soma, axon, and axonal growth cones of cultured motoneurons. Nuclei are stained with DAPI. e Munc13-1 mRNAs are upregulated in the soma of Rescue and RescueΔ3’UTR-transduced Smn KO motoneurons (***P = 0.0002, ****P < 0.0001; n = 134–186 cells from n = 2 biological replicates). f, g Munc13-1 mRNA levels are elevated in axons (f) *P = 0.0401, ****P < 0.0001; n = 88–124 cells from n = 3 biological replicates) and axonal growth cones (g) *P = 0.035, *P = 0.028, **P = 0.0011, **P = 0.002; n = 13–27 cells from n = 3 biological replicates) of Rescue, but not RescueΔ3’UTR-transduced Smn KO motoneurons. One-tailed Mann-Whitney U test in b and One-way ANOVA with Dunn’s post-test in (e–g). Bars represent mean ± SEM. Source data are provided as a Source Data file.