Extended Data Fig. 9: CRISPR KO of MAVS, RIG-I, and PKR and higher primates have increased Ku expression.
From: Ku limits RNA-induced innate immunity to allow Alu expansion in primates
a, The validation of PKR CRISPR KO. PKR is an IFN inducible gene. The poly I:C treatment induced maximal PKR expression and activation only in parental cells and ensured a clean CRISPR KO. b, Western blotting analyses of Ku in human, non-human primates, and non-primate cell lines. c-e, The mRNA expression levels of XRCC5(c), XRCC6(d), and ADAR1(e) were analyzed across species. Human and mouse data came from our previous studies, while other species’ data were from public datasets. The bars represent means and SEMs from n = 26, 25, 17, 18, 22 and 20 (c-d) and n = 36 and 20 (e) biologically independent samples (n values are in the same order as plotted). f-h, AlphaFold3 modeling of core human Ku (XRCC5 aa 1–536, pink/purple; XRCC6 aa 36–521, yellow/orange) bound to asSL1 RNA of Alu (70nt, silver) or perfectly paired dsDNA (58nt, gold). In each model, light-shaded proteins (pink and yellow) represent Ku bound to asSL1 RNA, while darker-shaded proteins (purple and orange) represent Ku bound to dsDNA. The top view (f) and front-side view (g) are presented in overlay format. The inset shows that base U19 and U21 of asSL1 flip out in the Ku-bound model but not in the absence of Ku. Panel (h) compares the structures of Ku-bound dsDNA (top, Ku hidden), Ku-bound asSL1 RNA (middle, Ku hidden), and free asSL1 RNA. The hairpin diagram was generated using the IDT Oligo Analysis Tool. The U19 and U21 bases of asSL1 flip out to accommodate Ku binding. See Methods for further details.
