34 | Chapter 2 homodimer, two surrogate light chains (SLC) and the Igα and Igβ units which possess the signaling capacity 155. At this stage, the rapid clonal expansion comes to a halt, mainly because of the cell-cycle arrest, which is required for the Rag1/2 expression and Ig gene recombination. The pre-BCR signal promotes the expansion of large pre-B cells, which remain dependent on IL7 signaling. The IL7R-induced STAT5 signaling also inhibits the Igk recombination, presumably through direct binding to Igk intron enhancer156,157. Therefore, in order to initiate Igk or Igl recombination, IL7 signaling must attenuate so that the large pre-B cells can escape their proliferative state. The crosstalk between the IL7R signaling and the pre-BCR signaling ensures successful progression of B-cell development and the maintenance of genome integrity by ensuring that proliferation and gene recombination are mutually exclusive. Importantly, pre-BCR signaling has the ability to counteract the IL7-dependent proliferation of pre-B cells through B-cell linker (BLNK)-mediated inhibition of the PI3K–AKT pathway158–160. The rapid clonal expansion is limited by the pre-BCR-driven RAS–ERK signaling pathway that operates to diminish proliferation. This occurs through the activation of the developmentally restricted transcription factor Aiolos (an Ikaros family member encoded by Ikzf3161). Aiolos then suppresses the transcription of Myc and Cyclin D3. In addition, in response to pre-BCR signaling, BLNK also mediates upregulation of interferon regulatory factor 4 (IRF4), which on the one hand further enhances the expression of Ikaros and Aiolos, aiding to limit the cell proliferation, and on the other hand, promotes light chain rearrangement by attenuating the IL7 signaling and promoting the binding of E2A to Rag promoter160,162. BCR signaling BCR serves as a regulator controlling the initial stages of development within the bone marrow. Even in the absence of an antigen, in resting B lymphocytes, there is an observable low-level phosphorylation of signaling intermediates, so-called “tonic signaling”, driven by the surface expression of BCR. The tonic BCR signaling is crucial for cell survival and its deletion results in loss of mature B cells163. These survival signals are mediated primarily through PI3K: while constitutive activity of nuclear factor kappa B (NF-kB) or MAPK or overexpression of BCL2 failed to prevent the loss of mature B cells in B cells with conditionally deleted BCR (BCRnegcells), the conditional PI3K signaling was sufficient to sustain the survival of mature B cells in BCRnegcells164. Following the discovery of AKT being the effector of PI3K165, it has been demonstrated that Akt (also known as protein kinase B or PKB) is recruited to the membrane and activated downstream of BCR166. In immature B cells with competent BCR the tonic BCR-signaling is low, which was shown to turn off RAG expression through the downstream effectors of PI3K: AKT, but also phospholipase C (PLC)gamma2 (PLCg) and Bruton’s tyrosine kinase (BTK)140,167,168 and in this way thought to protect the stability of the genome in developing B cells against excessive RAG1/2 ac-
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