Kras Is Critical for B Cell Lymphopoiesis

Small GTPases of the Ras subfamily regulate multiple signaling pathways and control numerous biological functions. Although the three major Ras members, Kras, Hras and Nras, are highly homologous, individual Ras gene can have distinct biological functions. Embryonic lethality of Kras-deficient mice precludes study of the biological role of Kras. Here, we generated and examined mice with hematopoietic- and B cell-specific deletion of Kras. In VavCreKras^fl/fl mice with hematopoietic deletion of Kras, the populations of bone marrow (BM) pre-, immature and mature B cells were reduced. The population of peripheral follicular (FO), marginal zone (MZ) and B1 mature B cells were also reduced in VavCreKras^fl/fl relative to control mice. In addition, BM chimeric mice with B cell-specific deficiency of Kras generated by transplantation of T- and B-null Rag1-deficient mice with a mixture of BM cells from VavCreKras^fl/fl mice and B cell-deficient μMT mice displayed a marked reduction of pre-, immature and mature B cells in the BM and mature B cells in the spleen. Thus, Kras deficiency intrinsically impairs early B cell development and late B cell maturation. Further, the effect of Kras deficiency on B-cell proliferation and survival was examined. The ³H-thymidine incorporation rate of Kras-deficient, relative to control, mature B cells in response to anti-IgM or anti-IgM plus IL-4 was markedly reduced. In addition, Kras-deficient B cells displayed a marked decrease of cell cycle entry and increase of cell apoptosis upon anti-IgM or anti-IgM plus IL-4 stimulation. Thus, Kras deficiency impairs BCR-induced B cell proliferation and survival. Lastly, the role of Kras in BCR signaling was studied. The level of total BCR-activated Ras (Ras-GTP) was largely reduced in Kras-deficient B cells. BCR-induced Ca²⁺ flux was comparable between Kras-deficient and control B cells. BCR-induced phosphorylation of Akt and IkBa was normal in mutant relative to control B cells. However, BCR-induced activation of ERK1/2 but not JNK or p38 was impaired in Kras-deficient relative to control B cells. Consistently, BCR-induced activation of Raf-1 and MEK1/2, the upstream activators of ERK1/2, was markedly reduced in mutant B cells. In addition, pre-BCR-induced ERK1/2 activation was impaired in Kras-deficient pre-B cells. Pre-BCR-induced activation of AP-1, the transcription factors downstream of ERK1/2, was decreased in mutant pre-B cells. Thus, Kras plays an important role in BCR- and pre-BCR-mediated activation of the ERK pathway in B cells. Taken together, these findings demonstrate that Kras is the Ras family member that critically regulates early B cell development and late B cell maturation through controlling the Raf-1/MEK/ERK pathway.