STAT5 and IL-7 signaling are thought to control B-lymphopoiesis by regulating key transcription factor genes and activating VH gene segments at the Igh locus. Using conditional mutagenesis, we demonstrate that transgenic Bcl2 expression rescued the development of Stat5-deleted pro-B cells by compensating for the loss of Mcl-1. Ebf1 and Pax5 expression as well as VH gene recombination were normal in Bcl2-rescued pro-B cells lacking STAT5 or IL-7Rα. In agreement with this finding, STAT5-expressing pro-B cells contained little or no active chromatin at most VH genes. In contrast, Igk rearrangements were increased in STAT5-or IL-7Rα-deficient pro-B cells. Hence, STAT5 and IL-7 signaling control cell survival and the developmental ordering of immunoglobulin gene rearrangements by suppressing premature Igk recombination in pro-B cells.

The transcription factor Pax5 represses B lineage-inappropriate genes and activates B cell-specific genes in B lymphocytes. Here we have identified 170 Pax5-activated genes. Conditional mutagenesis demonstrated that the Pax5-regulated genes require continuous Pax5 activity for normal expression in pro-B and mature B cells. Expression of half of the Pax5-activated genes is either absent or substantially reduced upon Pax5 loss in plasma cells. Direct Pax5 target genes were identified based on their protein synthesis-independent activation by a Pax5-estrogen receptor fusion protein. Chromatin immunoprecipitation (ChIP) of Pax5 together with chromatin profiling by ChIP-on-chip analysis demonstrated that Pax5 directly activates the chromatin at promoters or putative enhancers of Pax5 target genes. The Pax5-activated genes code for key regulatory and structural proteins involved in B cell signaling, adhesion, migration, antigen presentation, and germinal-center B cell formation, thus revealing a complex regulatory network that is activated by Pax5 to control B cell development and function.

V(H)-DJ(H) recombination of the immunoglobulin heavy chain (Igh) locus is temporally and spatially controlled during early B cell development, and yet no regulatory elements other than the V(H) gene promoters have been identified throughout the entire V(H) gene cluster. Here, we discovered regulatory sequences that are interspersed in the distal V(H) gene region. These conserved repeat elements were characterized by the presence of Pax5 transcription factor-dependent active chromatin by binding of the regulators Pax5, E2A, CTCF, and Rad21, as well as by Pax5-dependent antisense transcription in pro-B cells. The Pax5-activated intergenic repeat (PAIR) elements were no longer bound by Pax5 in pre-B and B cells consistent with the loss of antisense transcription, whereas E2A and CTCF interacted with PAIR elements throughout early B cell development. The pro-B cell-specific and Pax5-dependent activity of the PAIR elements suggests that they are involved in the regulation of distal V(H)-DJ(H) recombination at the Igh locus.