Fig 2: Anti-tau aggregation activity of compounds (E)-2f and 3a at 1, 3, 10, and 30 µM concentrations. (A) Tau proteins were stained with green fluorescence in HeLa cells transfected with the expression vector of tau 2N4R isoform carrying P301L mutation and incubated with the compounds. Microscopic observation was performed, and photos were shown on a 100 µm scale. Seed negative (seed-) and seed positive (seed+) indicate the absence and presence of fibrils of tau protein with P301L mutation, respectively. (B) Relative green spots per area (%) after exposure of Hela cells to compounds (E)-2f and 3a are compared to seed positive (seed+ = 100%). ImageJ counted more than 100 spots in the same area in each sample. *: denotes significant difference from control (seed+) at p < 0.05, **: denotes significant difference from control (seed+) at p < 0.01.

Fig 3: RfxCas13d-mediated lethality was not due to the loss of target gene function. a Western blot to measure SIK3-S expression level in brain lysates from Sik3-E5fl/fl and WT mice at 21 dpi. b Survival curve of Sik3-E5fl/fl (n=5) and WT (n=5) mice after injection of AAV-PHP.eB-hSYN-Cre. c Body weight change curve of Sik3-E5fl/fl (n=5) and WT (n=5) mice after injection of AAV-PHP.eB-hSYN-Cre. d, e RT-qPCR to measure the knockdown efficiency of Map2 (n=3) and Mapt (n=3) crRNAs in N2a cells. f Western blot to measure the knockdown efficiency of Rbfox3 crRNAs by knocking down overexpressed NeuN in HEK293T cells using RfxCas13d. g–i Transcriptome analysis in N2a cells 48 h after transfection of plasmids encoding RfxCas13d and crRNAs (n=3). j–l Body weight change curve of LSLRfxCas13dfl/fl mice after infection of PHP.eB carrying NT crRNA (n=5), Map2 crRNAs (n=5), Mapt crRNAs (n=5), or Rbfox3 crRNAs (n=5). m–o Survival curve of LSLRfxCas13dfl/fl mice after injection of AAV-PHP.eB carrying NT crRNA (n=5), Map2 crRNAs (n=5), Mapt crRNAs (n=5), or Rbfox3 crRNAs (n=5). p–r Western blot to measure MAP2 (at 6 dpi), Tau (at 25 dpi), or NeuN (at 50 dpi) expression level in brain lysates of LSLRfxCas13dfl/fl mice after injection of AAV-PHP.eB carrying NT crRNA, Map2 crRNAs, Mapt crRNAs or Rbfox3 crRNAs. d, e One-way ANOVA with Dunn’s multiple comparisons test. j–l Two-tail unpaired t test. b, m-o Log-rank test. Significance levels are noted as *P < 0.05, **P < 0.01, ****P < 0.0001 or ns (P > 0.05). All values are presented as mean ± SEM