Fig. 2.

Poly-GR peptides are detected as (peri)nuclear puncta in zebrafish embryos at 1-4 dpf. (A) Immunofluorescence staining for poly-GR (red) in 2 dpf old wild-type AB embryos after injection with 10 pg poly-GR or 400 pg mCherry only. N=30 per group. C9FTD patient frontal cortex sections were used as a positive control. Poly-GR peptides were detected as nuclear or perinuclear dots in all poly-GR-injected fish (arrows). Nuclei were stained with Hoechst. Scale bars: 20 µm. (B) Poly-GR is detected in postmortem frozen brain samples of C9ORF72 FTD/ALS cases (n=7) but not in postmortem frozen brain samples of FTD patients with mutations in progranulin (GRN; n=2), Valosin-containing protein (VCP; n=2), microtubule-associated protein tau (MAPT, n=1) or in brain samples of non-demented controls (n=5) (mean±s.e.m.). One-way ANOVA (P=0.0024) with post Tukey's test indicating a difference between the C9ORF72 FTD/ALS group and all other groups. (C) ELISA for the detection of poly-GR shows a signal of 10-20 pg peptide in 1-4 dpf wild-type AB embryos injected with 10 pg RNA encoding poly-GR. Two-way ANOVA (P=0.0003) with post Bonferroni test indicating that all timepoints were significantly different from mCherry-only-injected fish. n=90 fish per group per timepoint divided over three independent experiments (30 fish per group×three experiments per timepoint). *P<0.05, ***P<0.0001.