Fig. 2

Age-dependent accumulation of endogenous FUS and ALS-linked mutant FUS. a Proposed model for FUS auto-regulation. FUS protein binds to the intron 6–7 of FUS mRNA, which results in retention of the introns. Retained introns contain premature stop codon that triggers nonsense-mediated decay to lower the FUS mRNA. Black arrow denotes the PCR primers for exon 3/4 and 12/14. b, c Relative mRNA expression of mouse FUS in the hippocampus of non-transgenic, wild-type FUS (WT-FUS) and R514G-FUS mice at 3 and 12 months of age. b Expression levels of mouse FUS mRNA in WT-FUS and R514G-FUS mice are approximately 51% and 78% to that of non-transgenic mice, respectively, at 3 months of age using exon 3/4 primers (n = 3, per genotype). Similarly, 60% and 76% of mouse FUS mRNA in WT-FUS and R514G-FUS mice using exon 12/14 primers. c Expression levels of mouse FUS mRNA in WT-FUS and R514G-FUS mice are approximately 63% and 72% to that of non-transgenic mice, respectively, at 12 months of age using exon 3/4 primers (n = 3, for non-transgenic and WT-FUS mice, n = 6 for R514G mice). Similarly, 68% and 68% of mouse FUS mRNA in WT-FUS and R514G-FUS mice using exon 12/14 primers. d–g Immunoblot of FUS and tubulin using total hippocampal lysates from 3 to 12 month old non-transgenic, WT-FUS and R514G-FUS mice. FUS was probed with a monoclonal antibody (clone 4H11), which recognizes human and mouse FUS with similar affinity. d Human FUS transgene (red arrow) expressed in a level similar to the endogenous FUS (blue arrow) and the total FUS level is comparable among the non-transgenic, WT-FUS and R514G-FUS mice at 3 months of age. Tubulin was used as a loading control. Ponceaus S staining was also used to ensure similar loading. e Quantification of mouse FUS proteins in non-transgenic, WT-FUS and R514G-FUS mice at 3 months of age. f Accumulation of endogenous FUS (red arrow) and R514G-FUS (blue arrow) transgenes at 12 months of age. Tubulin was used as a loading control. Ponceaus S staining was also used to ensure similar loading. g Quantification of mouse FUS proteins in non-transgenic, WT-FUS and R514G-FUS mice at 12 months of age. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001