Sequential gene regulatory events leading to glucocorticoid-evoked apoptosis of CEM human leukemic cells:interactions of MAPK, MYC and glucocorticoid pathways

Abstract

Gene expression responses to glucocorticoid (GC) in the hours preceding onset of apoptosis were compared in three clones of human acute lymphoblastic leukemia CEM cells. Between 2 and 20h, all three clones showed increasing numbers of responding genes. Each clone had many unique responses, but the two responsive clones showed a group of responding genes in common, different from the resistant clone. MYC levels and the balance of activities between the three major groups of MAPKs are known important regulators of glucocorticoid-driven apoptosis in several lymphoid cell systems. Common to the two sensitive clones were changed transcript levels from genes that decrease amounts or activity of anti-apoptotic ERK/MAPK1 and JNK2/MAPK9, or of genes that increase activity of pro-apoptotic p38/MAPK14. Down-regulation of MYC and several MYC-regulated genes relevant to MAPKs also occurred in both sensitive clones. Transcriptomine comparisons revealed probable NOTCH-GC crosstalk in these cells.

Fig 1

Total genes increased ≥1.5 fold with p≤0.05 after treating cells with Dex continually for the stated times.

Fig 2

Time course of regulation up (A) or down (B) of selected genes.

Fig 3

Growth of GR/Myc network; pre-apoptotic interactions over time in Dex-dependent CEM clones. Red indicates increase; green, decrease

Fig 4

CaN inhibitors enhance CEM cell kill by Dex.. C7-14 cells were exposed to Dex, FK506, CsA, or combinations of the three for 48 hr. This figure shows the effects after 48 hr, when they were most striking. Between each pair of bars, p was ≤ 0.008. Further data in Supp Fig 2 and Table 3.

Fig 5

Immunoblots from extracts of 3 clones of CEM cells showing a) basal levels of JNK1/2 protein, total (apo-) and phosphorylated/activated (Phospho-) and b) PYK2 protein levels at control/basal state (C) and after Dex treatment (D).

Fig 6

Interactions of GR, MYC and MAPK pathways, from analysis of time course of altered gene regulation in CEM cells. GC (triangle) enters cell and activates GR, which by up- or down-regulating many genes, causes lowered activity and or amounts of antiapoptotic ERK and JNK and enhanced activity of proapoptotic p38. In forward feedback, p38 phosphorylates a specific ser of GR, which further enhances GR activity. GR down-regulates Myc which reduces transcription of JNK and further affects MAPKs via downregulation of various (yellow) other Myc-dependent genes. All arrows indicate direct or indirect regulation. Large right-angle arrows, up or down regulation.. Red indicates increased activity/amount; green, decreased.