Fig. 5

Induction of ISR-mediated cell survival under nutrient-restricted conditions through KD of Sp1, direct SSBs induced by H₂O₂, or direct DSBs induced by ionising radiation. a (left) Representative Western blot of pATM, Sp1, and XRCC1 levels after siSp1. (right) Quantification of XRCC1 levels shown on the left, n = 5. b, c Representative Western blot of b Sp1, Tubulin, γH2AX, and Histone H3, or c Sp1, ATF4, and peIF2α after siSp1. Normalised levels of ATF4 and peIF2α are indicated. d–e Phase-contrast images of cells treated with siCon (d) or siSp1 (e), grown in a medium containing 1% FCS. f Quantification of d and e, n = 4. g Alkaline comet assay of cells treated with 25 μM H₂O₂. h Neutral comet assay of cells pre-treated with 25 μM or 50 μM H₂O₂ for three consecutive days, or 4 Gy IR 15 min prior to harvesting, respectively. n = 4. i, j Representative Western blots of i γH2AX and Histone H3 or j XRCC1, ATF4, and peIF2α, in cells treated as in h. Normalised levels of XRCC1, ATF4, and peIF2α are indicated. k–m Phase-contrast images of cells treated as in h prior to seeding into a medium containing 1% FCS. n Quantification of k–m. n = 4. o Western blot of pATM, Tubulin, ATF4, and peIF2α in control cells or cells treated with 0.5, 1, or 2 Gy of IR on three consecutive days. Normalised levels of ATF4 and peIF2α are indicated. p–s Phase-contrast images of control cells (p) or cells treated as in o prior to seeding into a medium containing 1% FCS. t Quantification of p–s, n = 4