Skip to main content
. 2022 Jan 24;28(1):104–116. doi: 10.1038/s41591-021-01615-z

Extended Data Fig. 3. Decreased lifespan of mutant FUS heterozygous animals and absence of NMJ denervation and MN loss in newborn P517L/P517L mice.

Extended Data Fig. 3

(a) Box (median, 25th and 75th percentiles) and whiskers (2.5th and 97.5th percentiles) plot of birth weights of mice with the indicated combinations of Fus WT, P517L, Δ14, and null-knockout (KO) alleles. *P < 0.05, **P < 0.01 and ***P < 0.001, using one-way ANOVA with Tukey’s post hoc test. N = 13–183 animals per genotype as indicated in Table 1. (b) Selected data reproduced from (a) to illustrate partial functionality and dose-dependent toxicity of mutant FUS. Increased birth weight of P517L/KO compared to KO/KO animals demonstrates that mutant FUS protein is able to partially rescue the null phenotype and thus is functional. Comparison of P517L/P517L versus P517L/KO animals demonstrates that further addition of mutant FUS protein decreases birth weight, consistent with dose-dependent toxicity of mutant FUS protein. Statistical significance was assessed in (a) as a part of a full analysis of all genotype groups. (c) Kaplan-Meier survival curves for successfully weaned WT/WT, P517L/WT, Δ14/WT, and WT/KO mice. Both heterozygous mutants but not WT/KO have significantly decreased median survival age compared to wild type controls. All possible pairwise comparisons were performed using Log-rank (Mantel-Cox) test and the resulting p-values were adjusted for multiple comparison using the Bonferroni correction. (d) NMJ staining of tibialis anterior (TA) muscle of newborn WT/WT (left) and homozygous P517L/P517L(right) animals using antibodies to pre-synaptic synaptophysin (green) and alpha-bungarotoxin (magenta, post-synaptic). Scale bar= 100 µm. (e) Quantification of innervated NMJs in the WT/WT (black) and homozygous P517L (dark red) animals. N = 3 for WT/WT and 4 for P517L/P517L groups respectively. (f) Immunofluorescence staining of lumbar level 4–5 (L4-L5) MNs in WT/WT (left) and homozygous P517L/ P517L (right) animals using anti-ChAT and anti-P517L antibodies (right inset panel). Scale bar=25 µm for the left and middle panels, 100 µm for the right panel. (g) Quantification of ChAT-positive neurons in lumbar levels 4–5 (L4-L5) in newborn WT/WT (black) and homozygous P517L/ P517L (dark red) animals. N = 3 animals per genotype. For e and g, statistical significance was assessed using Welch’s t-test. Error bars represent SD.