Fig. 5.

Myc inhibits and cyclin-dependent kinase 18 (CDK18) promotes ATR signaling in poly(ADP-ribose) polymerase inhibitor-treated glioblastoma stem-like cells (GSCs). a Representative western blots (n = 3) showing olaparib-induced DNA damage response (DDR) in GSCs with knockdown (sh) or overexpression of MYC and MYCN. Cells treated with mock or olaparib (Ola, 10 μM) for 24 and 48 h and with (+) or without (−) doxycycline (Dox). Same membranes and β-Actin control as in Fig. 2d. b Quantification of p-Chk1 levels determined from a and two additional independent experiments. Normalized to β-Actin. c (upper) Representative western blot (n = 3) showing kinetics (hours indicated) of p-Chk1 in olaparib-treated (10 μM) MGG4-shMYC or MGG4-shCon, with Dox. (lower) Quantification of p-Chk1 levels normalized to Vinculin from (upper) and two additional independent experiments. d (upper) Representative western blots (n = 3) showing olaparib-induced DDR in MGG4 with CDK18 overexpression or BT74 with CDK18 knockdown after treatment for 48 h with Dox. (lower) Quantification of p-Chk1 levels from (upper) and two additional independent experiments. With (+) or without (−) olaparib. e (Upper) Representative western blots (n = 3) showing olaparib-induced DDR in MGG4 with knockdown of MYC alone (MGG4-shMYC-shCon) and with CDK18 (MGG4-shMYC-shCDK18) or in BT74 overexpressing MYC alone (BT74-MYC-Con) and with CDK18 (BT74-MYC-CDK18), treated as in d. (lower) Quantification of p-Chk1 from (upper) and two additional independent experiments, normalized to β-Actin. *p < 0.05, **p < 0.01 and ***p < 0.001, t test. Mean ± SEM