(A) ATRX promotor binding at Chek1 loci in
mNPC cells (ChIP-seq, n=3 from Danussi et al.,
2018). (B) mGBM cells with ATRXKO
(“NPA”) show reduced ATRX and H3.3 binding at Chek1 gene loci
(“1” through “4” from A) compared to
mGBM cell controls without ATRXKO (“NP”), n=3.
(C) Chek1 expression is downregulated in mNPC
cells with ATRX loss. (D) Western blot of U251 ATRXEV
and U251 ATRXKO cells with and without 4 Gy IR. ATR pathway proteins
are marked in purple. (E) Western blot of U251 ATRXKO
cells with isogenic Chk1 overexpression or empty vector (n=3 replicates for C
and D). (F-G) Incucyte live cell imaging analysis of U251
ATRXKOChk1OE cells incorporated with the FastFUCCI
reporter plasmid show a gradual return (more than 1.5X slower) to cycling after
4 Gy IR. (Mean ± SEM for triplicate experiments are shown.
*P≤ 0.05, **P≤ 0.01, and
***P≤0.001 using Welch’s t-test). For
additional data, see also Figure S4.