AP-1 protein induction during monopoiesis favors C/EBP: AP-1 heterodimers over C/EBP homodimerization and stimulates FosB transcription

摘要

AP-1 proteins heterodimerize via their LZ domains to bind TGACGTCA or TGACTCA, whereas C/EBPs dimerize to bind ATTGCGCAAT. We demonstrate that intact C/EBP?± also heterodimerizes with c-Jun or c-Fos to bind a hybrid DNA element, TGACGCAA, or more weakly to TGATGCAA. A 2:1 ratio of c-Jun:C/EBP?± or c-Fos:C/EBP?± was sufficient for preferential binding. Semiquantitative Western blot analysis indicates that the summation of c-Jun, JunB, and c-Fos levels in differentiating myeloid cells is similar to or exceeds the entirety of C/EBP?± and C/EBP?2, indicating the feasibility of heterodimer formation. Induction of AP-1 proteins during monocytic differentiation favored formation of C/EBP:AP-1 heterodimers, with C/EBP?± homodimers more evident during granulopoiesis. Approximately 350 human and 300 murine genes contain the TGACGCAA motif between a€“2 kb and +1 kb of their transcription start sites. We focused on the murine Fosb promoter, which contains a C/EBP:AP-1 cis element at a€“56 and a€“253, with the hFOSB gene containing an identical site at a€“253 and a 1-bp mismatch at a€“56. C/EBP?±:AP-1 heterodimers bound either site preferentially in a gel-shift assay, C/EBP?±:c-Fos ER fusion proteins induced endogenous Fosb mRNA but not in the presence of CHX, C/EBP and AP-1 proteins bound the endogenous Fosb promoter, mutation of the a€“56 cis element reduced reporter activity fivefold, and endogenous FosB protein was expressed preferentially during monopoiesis versus granulopoiesis. Increased expression of Jun/Fos proteins elevates C/EBP:AP-1 heterodimer formation to potentially activate novel sets of genes during monopoiesis and potentially during other biologic processes.