Dissecting the roles of the different isoforms of the lymphoid-specific transcriptional coactivator OBF-1

Abstract

B cell differentiation depends on a highly regulated transcriptional machinery. The octamer sequence (ATGCAAAT) is a conserved DNA motif that confers transcriptional B cell specificity. This sequence is mainly found in immunoglobulin promoters, but also in some B cell specific genes. OBF-1, also known as Bob.1 or OCA-B, is a B lymphocytespecific transcription factor which coactivates Oct1 and Oct2 on such octamer sites. The main function of OBF-1 was identified in late stage B cells, as OBF-1 deficient mice exhibit a severe reduction of T cell-dependent immune response and a lack of germinal center formation in the spleen. However little is known about the role of OBF-1 in early B cells. The most relevant biological finding of this thesis illustrates the fact that OBF-1 expression level has to be tightly regulated to allow early B cell development. Most OBF1 target genes were identified in late B cells. This thesis proposes for the first time three direct OBF-1 target genes in early B cells, namely Id2, Id3, and EBF1. Indeed OBF-1 can bind their respective promoters. Aberrant OBF-1 expression led to increased levels of Id2 and Id3 transcripts, which resulted in differentiation blocks before and after B cell commitment. In fact Id2 and Id3 are known to be negative regulators of B cell differentiation probably by antagonizing E2A DNA binding. EBF1 is a transcription factor involved in B cell commitment and was reported to induce OBF-1 expression. EBF1 was upregulated in mice overexpressing OBF-1, therefore the activation of EBF1 promoter by OBF-1 constitutes a positive feedback loop. The OBF-1 gene contains two active start codons giving rise to nuclear and myristoylated cytoplasmic isoforms. The second part of this thesis describes the investigation of the physiological role of each isoform. BAC transgenic mice expressing only one isoform were generated to decipher this issue. The nuclear isoform was found to be the main player among the isoforms, as the corresponding transgenic mice did not have any OBF-1 associated phenotype and most of the OBF-1 regulated genes were under its control. IL-7 dependent cultures of PreBI cells from mice deficient for OBF-1 were reported to proliferate faster. In fact the cytoplasmic OBF-1 isoform might have a role under these culture conditions, as the nuclear OBF-1 isoform could only partially rescue this hyperproliferation.