Functional collaboration between HLH transcription factors in B cell development

Abstract: The cells in B cell development can be divided into several subgroups or fractions, e.g. early-pro-B-, pro-B, large-pre-B-, small-pre-B-, immature and mature B cells, where the stages reflect the maturity degree of the cells. This characterization is based on the expression of intracellular markers such as the recombination activating genes 1 and 2(Rag 1 and Rag 2), the terminal deoxynucleotidyl transferase gene (TdT) and the expression of surface markers such as the components of the pre-B and B cell receptor (pre-BCR and BCR) as well as the rearrangement status of the heavy- and light chain genes. The expression of these genes is strongly regulated and controlled, otherwise it will lead to disruptions in the B cell development. One important way of maintaining this is by the interaction of proteins denoted transcription factors (TFs) and the genes regulatory sequences, i.e. promoter and enhancer sequences. This interaction, where the TFs help to stabilize and activate the transcriptional machinery, is called transcriptional regulation. Several TFs have been shown to play a crucial role in this regulation process during B cell development, since disruption of genes encoding these TFs causes developmental blocks and disruptions. Two of these factors are the Early B cell Factor (EBF) and E47. Using the mouse EBF cDNA as a probe, we managed to clone the human homologue in a human pre-B cell cDNA library. This made possible a study of the role of this protein during the human B cell development. In this thesis we suggest that EBF and E47 have conserved target sequences and function between man and mouse. This evolutionary conservation reflects their importance during B cell development. Furthermore, there is an interaction during transcription between the two TFs that in many cases result in a synergistic cooperation, reflected in the activation degree of the gene in question. Several target genes are components of the pre-BCR and positioned in different chromosomes, suggesting that EBF is a pleiotropic activator of genes encoding the pre-BCR. Also, we were able to clone a promoter region upstream the EBF gene. This sequence contain binding sites for both E47 and EBF, suggesting that E47 is a key regulator in the transcription of EBF and that EBF auto-regulates it self in a loop. This also suggests that E47 is placed upstream of EBF in the hierarchy of transcriptional factors that participates in the transcriptional regulation of B cell development.