Figure 2.

MTX induces expression of genes whose protein products promote apoptosis. A, Jurkat cells were cultured with MTXfor 48 hr., transcript levels of genes whose protein products are known to influence apoptosis were determined [Sup. Figure 1]. Results are expressed as fold increase in transcript levels over untreated cells after normalization to GAPDH. B, JNK1-DN, JNK2-DN or ‘empty vector’ plasmids with a GFP expression plasmid were introduced into Jurkat cells by nucleofection. Left panel: c-Jun levels in, black line, untreated cells, blue line, MTX treated cells, red line, MTX treated cells transfected with JNK1-DN. Middle panel: c-Jun levels in, black line, untreated cells, blue line, MTX treated cells, red line, MTX treated cells transfected with JNK2-DN. Right panel: Mean changes in expression levels of c-Jun, Fos, PUMA and TRAILR1 in MTX treated Jurkat cells determined by flow cytometry expressed as fold change relative to untreated Jurkat cells ± S.D. C, Inhibition of MTX-mediated increases in c-Jun and Fos protein levels by specific JNK inhibitors, BI-78D3 or pepJIP1 relative to untreated Jurkat cells ± S.D. * P < 0.05 relative to protein levels in MTX-treated Jurkat cells.