Regulation of B cell Differentiation and Transformation by the Phosphoinositide 3-kinase (PI3K) Pathway

A large effort in our lab is to understand how components of the PI3K signaling pathway specifically contribute to B cell growth, proliferation and survival as well as functions unique to B cells, namely antigen receptor assembly, signaling and antibody production. Activation of the PI3K signaling network recruits cytosolic effectors to the plasma membrane via pleckstrin homology (PH) domain-binding to phosphatidylinositol 3,4,5-trisphosphate (PI(3,4,5)P3). In particular, PDK1 has been identified as a pivotal downstream effector of PI3K that activates Akt, p70S6kinase, PKC enzymes and other targets that control cell metabolism, quiescence, proliferation and survival. The Akt kinases have also been shown to negatively regulate the activity of the FOXO transcription factors. In doing so, expression of FOXO target genes is halted, leading to enhanced cell cycle progression and survival. In B cells, we posit that the PDK1-dependent pathway acts in synergy with the PI(3,4,5)P3-dependent Btk signaling pathway, which is crucial for NF-κB activation and Ca++ mobilization. We are currently defining the relative importance and distinct functions of the PDK1- and Btk-dependent pathways in B cells.

B cell Lymphomagenesis

In recent years, we have also focused on the regulation of PI(3,4,5)P3 in B cells through the opposing functions of PI3-kinase and the 3’-inositol phosphatase and tumor suppressor PTEN. This work has been extended to include a comparative functional analysis of PTEN versus the 5’-inositol phosphatase, SHIP. Both PTEN and SHIP hydrolyze PI(3,4,5)P3; however, we found that inactivation of Pten or Ship did not result in transformation, raising the postulate that PTEN and SHIP acted cooperatively to suppress transformation. Indeed, we found that mice lacking both of these phosphatases in the B lineage developed a lethal B lymphoma. These data are the first evidence that SHIP acts as a tumor suppressor in a coordinate fashion with PTEN to dampen PI3K signaling and suppress B cell neoplasia. We are utilizing this model to address the role of antigen and tonic/inducible BCR signaling in the generation and propagation of B lymphoma cells. As most B Non-Hodgkin Lymphoma (NHL) cells do not proliferate ex vivo, we are also investigating the environmental stimuli and tissue microenvironments that drive lymphoma progression.

Functional Distinctions of IgM vs. IgG-containing B cell Receptors

Molecular studies of memory B cell activation have been hampered by their relative scarcity and lack of effective technical approaches to unveil the distinctive activation pathways employed by memory versus naïve B cells. Most memory B cells express IgG-containing B cell receptors, while naïve B cells express IgM/IgD. Relative to IgM/IgD, IgG/IgE receptors have an extended cytoplasmic tail that contains conserved phosphotyrosine (pTyr) motifs and confers enhanced survival and proliferation. We are applying cutting-edge proteomic approaches, gene silencing methods and mouse models to elucidate differences in IgM/IgD- versus IgG-based signaling. As tyrosine kinase/phosphatase activity is known to be critical for BCR signaling, our analysis is focused on determining differences in induced tyrosine phosphorylation of downstream substrates by IgG versus IgM/IgD receptors. Determining the functional importance and signaling properties of newly identified downstream effectors should greatly aid our understanding of how IgG expression directs memory B cell propagation and differentiation.