Early B Cell Factor Is an Activator of the B Lymphoid Kinase Promoter in Early B Cell Development

Abstract

AbstractEarly B cell factor (EBF) is a transcription factor suggested to be involved in the transcriptional control of several B cell restricted genes. EBF is also essential for B lymphocyte development because mice carrying a homologous disruption of the EBF gene lack mature B lymphocytes. This makes the identification of genetic targets for EBF important for the understanding of early B cell development. Examination of the nucleotide sequence of the B lymphoid kinase (Blk) promoter suggested the presence of an EBF binding site, and in vivo footprinting analysis showed that the site was protected from methylation in a pre-B cell line. EMSA indicated that recombinant and cellular EBF interact physically with this site; furthermore, transient transfections indicated that ectopic expression of EBF in nonlymphoid HeLa cells activate a Blk promoter-controlled reporter construct 9-fold. The defined EBF binding site was also important for the function of the Blk promoter in pre-B cells, because transient transfections of a reporter construct under the control of an EBF site-mutated Blk promoter displayed only 20–30% of the activity of the wild-type promoter. Furthermore, transient transfections in HeLa cells proposed that EBF and B cell-specific activator protein were able to cooperate in the activation of a Blk promoter-controlled reporter construct. These data indicate that EBF plays an important role in the regulation of the Blk promoter in early B cell development and that EBF and BSAP are capable to act in cooperation to induce a target gene.