Synergistic activation of the human choline acetyltransferase gene by c-Myb and C/EBPbeta.

ABSTRACT

To elucidate regulatory mechanisms at the transcriptional level of the human choline acetyltransferase gene (hChAT) we performed cotransfections assays in NG108-15 and SN56 cells using ChAT-CAT reporter plasmids with c-Myb and C/EBPbeta expression plasmids. The hChAT gene has several promoters, one of which (promoter P2 or M-type) is both c-Myb and C/EBPbeta inducible as 3-4-fold trans-activation was obtained in both cell lines when using either c-Myb or C/EBPbeta expression vectors alone. The simultaneous expression of c-Myb and C/EBPbeta in the absence or presence of NGFI-C (egr4) leads respectively to a 15-fold and 32-fold synergistic transcriptional activation of promoter P2. In the region upstream of exon M (P2) we identified a functional composite element including a c-Myb next to a C/EBP binding site. An oligonucleotide containing the composite element confers c-Myb and C/EBPbeta responsiveness to a heterologous promoter which is reduced after mutation of the c-Myb binding site. We also show that the coactivators CBP/p300 are required for c-Myb and C/EBPbeta trans-activation function and that RARalpha, RXRalpha and T3R have an inhibitory action on the synergistic transcriptional activity of c-Myb and C/EBPbeta and propose a model to explain the phenomena. Taken together, the results suggest that the synergistic effect of c-Myb and C/EBPbeta, previously observed in the hematopoietic system, functions equally in the neuronal system.