MODULATION OF MESSENGER-RNA EXPRESSION OF A NOVEL HUMAN MYELOID-SELECTIVE CCAAT ENHANCER BINDING-PROTEIN GENE (C/EBP-EPSILON)/

Human C/EBP epsilon is a newly cloned gene coding for a CCAAT/enhancer binding protein that may be involved in the regulation of myeloid dif ferentiation. Our studies showed that levels of C/EBP epsilon mRNA wer e markedly increased in NB4 cells (promyelocytic leukemia line), becau se they were induced by 9-cis retinoic acid (9-cis RA) to differentiat e towards granulocytes. Accumulation of C/EBP epsilon mRNA occurred as early as 1 hour after exposure of NB4 cells to 9-cis RA (5 x 10(-7) m ol/L); and at 48 hours, levels were increased by 5.1-fold. Dose-respon se studies showed that 10(-7) to 10(-6) mol/L 9-cis RA (12 hours) resu lted in peak levels of C/EBP epsilon mRNA; but even 10(-10) mol/L 9-ci s RA increased levels of these transcripts. NB4 cells pulse-exposed (3 0 minutes) to all-trans retinoic acid (ATRA), washed, and cultured (3 days) with either dimethylsulfoxide (DMSO) or hexamethylene bisacetami de (HMBA) had a prominent increase in levels of C/EBP epsilon mRNA and an increase in granulocytic differentiation, but exposure to either D MSO or HMBA alone had no effect on base levels of C/EBP epsilon and di d not induce differentiation. Macrophage-differentiation of NB4 reduce d levels of C/EBP epsilon mRNA. Nuclear run-off assays and half-life s tudies showed that accumulation of C/EBP epsilon mRNA by 9-cis RA was due to enhanced transcription. Furthermore, this C/EBP epsilon mRNA ac cumulation did not require synthesis of new protein factors because 9- cis RA induced C/EBP epsilon mRNA accumulation in the absence of new p rotein synthesis. ATRA also induced expression of C/EBP epsilon protei n in NB4 cells, as shown by Western blotting. In contrast to the incre ase of C/EBP epsilon in 9-cis RA-mediated granulocytic differentiation , the DMSO-induced differentiation of HL-60 cells down the granulocyti c pathway was associated with an initial reduction of C/EBP epsilon mR NA levels. In summary, we have discovered that expression of C/EBP eps ilon mRNA is markedly enhanced as the NB4 promyelocytes are induced by retinoids to differentiate towards granulocytes. This induction of C/ EBP epsilon mRNA expression is transcriptionally mediated and occurs i n the absence of synthesis of additional protein factors. We suspect t hat the C/EBP epsilon promoter/enhancer contains a retinoic acid-respo nse element that is directly stimulated by retinoids. (C) 1997 by The American Society of Hematology.