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. 2020 Oct 29;183(3):636–649.e18. doi: 10.1016/j.cell.2020.09.020

Figure 1.

Figure 1

Inflammatory Signaling from TDP-43 Is Dependent on cGAS/STING

(A) Vector alone (Ctrl) or plasmids encoding FLAG-tagged wild-type (WT) TDP-43 and the ALS mutation (Q331K) were transiently overexpressed in mouse embryonic fibroblasts (MEFs) genetically deficient for different innate immune sensors. Expression of Ifnb1 and Tnf was measured by qPCR after 72 h and was ablated only when cGAS or STING were genetically deficient.

(B) Inducible TDP-43 constructs (WT or Q331K) were transduced into WT or STING CRISPR knockout (KO) THP-1 cells. 72 h after Dox induction, qPCR for IFNB1 and TNF was performed.

(C) TDP-43-overexpressing THP-1 cells as in (B) were subjected to western blot analysis of inflammatory signaling pathways related to type I IFN and NF-κB.

(D and E) The cGAS inhibitor RU.521 and STING inhibitor H-151 prevent IFNB1 and TNF induction from TDP-43-overexpressing THP-1 cells used in (B) and (E) iPSC-derived motor neurons from ALS patients compared with healthy controls (RU.521, 10 μM; H-151, 1 μM). DMSO was used as a solvent control (0).

(F and G) Quantification of cGAMP by ELISA from (F) cell lysates of iPSC-derived motor neurons from healthy controls and ALS patients and from (G) post-mortem spinal cord samples of patients with sporadic ALS (n = 16) or MS (n = 12).

Data are mean ± SEM, pooled from 3–5 independent experiments ([A], [B], and [D]–[F]) or representative of 3 independent experiments (C). The p values were calculated using two-way ANOVA to Ctrl in (A), (B), and (D) or unpaired t test between healthy control and ALS patient iPSC-MN lines (G298S, M337V, and A382T) in (E)–(G). ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. See also Figure S1 and Table S1.