The regulation of megakaryocytic differentiation is poorly understood. Using K562 cells, which can partly recapitulate the process in response to phorbol 12-myristate 13-acetate (PMA), we performed microarray-based gene expression profiling to identify genes that play significant roles in megakaryopoiesis. Here, we describe the function of FosB, an AP-1 transcription factor. FosB is induced in PMA treated K562 cells in a sustained manner and forms an active AP-1 protein-DNA complex. Down-regulation of FosB with specific shRNAs inhibited the induction of CD41, a specific cell surface marker of megakaryocytes. We also show that activation of the PKC-MEK-ERK signaling pathway is required for induction of FosB and CD41. Finally,we cross-examined the microarray data in conjunction with gene function annotation data to identify additional target genes of FosB. We define 3 genes, INHBA, CD9, and ITGA2B as regulatory targets of FosB and show that CD9, in particular, is a direct target of FosB.