Fig. 3: TDP-43 knockdown corresponds to R6/2 transcriptional changes. | Nature Neuroscience

Fig. 3: TDP-43 knockdown corresponds to R6/2 transcriptional changes.

From: Aberrant splicing in Huntington’s disease accompanies disrupted TDP-43 activity and altered m6A RNA modification

Fig. 3

a, Reproducible R6/2 HD and NT IDR TDP-43 peaks were centered and plotted on all TDP-43 binding sites, compared to Down DEGs, Up DEGs and cassette exons. b, TDP-43 mRNA level by qPCR normalized to cyclophilin as a percentage of PBS control. CTX, cortex; SC, spinal cord; STR, striatum. n = 5. Black bar represents the mean. c, Heatmap showing clustering of 3mos HD R6/2, NT, TDP-43 ASO treated and control PBS treated on TDP-43 KD-dependent DGE changes. Gradient scale represents z-scores of normalized gene counts. d, Schematic of iPSC differentiation into MSNs with TDP-43 KD by siRNA. D, day; NPC, neural progenitor cell; NSC, neural stem cell. e, Left: western blot for TDP-43 protein levels after treatment of MSNs with TDP-43 siRNA. n = 3 differentiation replicates per condition. Right: bar graph plots TDP-43 intensity normalized to Revert total protein stain. Data are presented as mean values ± s.e.m. Statistical significance was determined by two-way ANOVA with Sidak’s multiple comparisons test (18Q: P < 0.0001, 95% CI: 6.230–10.20; 50Q: P < 0.0001, 95% CI: 6.084–10.06). IB, immunoblot; M, marker. f, Venn diagram showing the overlap of DEGs between HTT-18Q MSNs scramble control versus TDP-43 siRNA and HTT-18Q MSNs versus mHTT-50Q. g, Example of key gene expression changes anticipated from TDP-43 KD. n = 3 differentiation replicates per condition. Statistical significance was determined by unpaired two-tailed t-test. Data are presented as mean values ± s.e.m. (TDP-43 KD versus Ctrl STMN2: P < 0.0001, t = 24.41, d.f. = 4, F = 2.639, 95% CI: −768.6 to −611.6; 18Q versus 50Q STMN2: P = 0.0001, t = 14.49, d.f. = 4, F = 2.344, 95% CI: −496.4 to −336.7; TDP-43 KD versus Ctrl UNC13B: P = 0.0076, t = 4.988, d.f. = 4, F = 2.598, 95% CI: −64.21 to −18.29; 18Q versus 50Q UNC13B: P = 0.0132, t = 4.242, d.f. = 4, F = 2.249, 95% CI: −36.78 to −7.682; TDP-43 KD versus Ctrl CAMK2B: P = 0.0006, t = 10.04, d.f. = 4, F = 5.482, 95% CI: −47.26 to −26.79; 18Q versus 50Q CAMK2B: P = 0.0287, t = 3.345, d.f. = 4, F = 4.953, 95% CI: −29.00 to −2.693). Ctrl, control.

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