Dissection of the Human Pre-B Cell Receptor (pre-BCR) Signalling Cascade.

Pre-BCR acts as a critical checkpoint guards in pre-B cell selection and expansion. There is also an increasing body of evidence suggesting a role for this receptor in leukemogenesis. However, the molecular events that follow its stimulation remain largely unknown. Most of the functions that have been so far attributed to this receptor are speculative or deduced from genetic studies conducted in mice, using either knock out or transgenic mice, which makes difficult to figure out whether these functions are directly or indirectly triggered via pre-BCR stimulation. Moreover, one should also be cautious in extrapolating data from mice to humans, as many discrepancies in B cell development have been reported between both species. We extensively analyzed herein the pre-BCR signalling machinery using the human 697 and Nalm6 pre-B cell lines and F(ab′)2 anti-human heavy chain μ to stimulate pre-BCR. We showed that stimulation of this receptor results in a rapid phosphorylation of Lyn, Blk, Syk, BLNK, LAB as well as adaptors that were so far considered as T-cell specific, together with Vav, Btk, PLCγ2 and Akt, which in turns activates FKHRL and GSK3. It also induces activation of Ras and the 3 major downstream arms of the Ras/MAPKinase pathway as well as number of transcription factors that will be further discussed. Its stimulation is followed by cell cycle progression and proliferation that are associated with p27^kip1 degradation and Rb phosphorylation and with c-Myc upregulation. Moreover, pre-BCR aggregation induces phosphorylation of Bcl2 on residue serine 70 suggesting that this receptor mediates anti-apoptotic signal. In addition, CD19 coligation was found to potentiate pre-BCR signalling capabilities. c-Cbl and SHIP1 are rapidly phosphorylated upon pre-BCR aggregation, probably acting as negative regulators of this receptor signalling. We bring also evidence that Syk plays a pivotal role in pre-BCR signalling cascade and functions. Altogether these findings demonstrate that pre-BCR is directly implied in the control of proliferation and survival signals in pre-B cells and describes the multiple molecular events involved in these processes. We also show that pre-BCR stimulation induces downmodulation of rag1/2 and zap70 gene transcription but not of λ5, vpre-b, CD79a, E2A and EBF. Besides its major interest in understanding the regulation of the first steps of B cell development, analysis of pre-BCR associated-signalling pathways in leukemic B cell progenitors would certainly be helpful to better characterize the ongogenic events and identify new therapeutic targets in acute lymphoblastic leukemia.