Figure 3.
qRT-PCR confirms cooperative gene regulation by EVI1 and ATRA in U937 cells. RNA from U937_vec and U937_EVI1 cells that had been treated with solvent or ATRA for 24 h was reverse transcribed and subjected to qRT-PCR for the indicated genes. Gene expression relative to the housekeeping gene B2M and to solvent treated U937_vec cells was calculated using the ΔΔCt method.88 Results represent means + SEs from at least 3 independent experiments; all of these were also independent of the microarray experiments. In the absence of ATRA, TGM2 mRNA levels were below the detection limit. Significance was calculated using Student's 2-tailed t-test (*, P < 0.05; **, P < 0.01; ***, P < 0.001). Even though results for GDF15 and IL1B were not significant due to variation in induction factors, the expression levels of all genes were reproducibly substantially higher in ATRA treated EVI1 expressing cells as compared to all other conditions. Please note the differences in scale between the different panels.