Developmental changes of GM-CSF gene inducibility in embryonal carcinoma cells

Murine embryonal carcinoma (EC) P19 cells, a tissue culture model of early embryonic development, failed to produce cytokines, such as interleukin-3 (IL-3), IL-4, granulocytemacrophage colony stimulating factor (GM-CSF) and interferon-beta (IFN-beta) at the mRNA level. Differentiation induced by retinoic acid (RA) released this repression to produce some cytokines. GM-CSF and IFN-beta genes were expressed in response to PMA/A23187, poly(I):poly(C), IL-1 alpha, forskolin, or LPS stimulation in differentiated P19 cells, whereas IL-3 and IL-4 genes were not expressed. To elucidate the mechanism of the GM-CSF gene induction after differentiation, we transfected a series of 5' deletion mutants of the mouse GM-CSF promoter fused to the bacterial CAT gene. The 740-bp fragment of the 5'-flanking region mediated the positive response. Deletion analysis revealed that the 5' boundary region of the DNA element required for activation lies between positions -95 and -84 and the region upstream of position -95 appears inhibitory. These results indicate that the maturation of the transcriptional machinery after differentiation results in the activation of the GM-CSF gene.