Decoding and functional characterization of the tonic B cell Antigen receptor signaling Network in human B cells

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Beyond the development, activation, and differentiation of B cells the B cell antigen receptor (BCR) controls their survival. The BCR-mediated survival signals are referred to as tonic BCR signals because they occur in the absence of antigen binding. Despite substantial evidence for an importance of the phosphatidyl-3-kinase signaling pathway the molecular details of the tonic BCR signal transduction remain poorly understood. A functional characterization of the underlying signaling processes is, however, important to improve our understanding of survival signals in B cells and their dysregulation in B cell neoplasia. Hence, the overall aim of this project is the elucidation of tonic BCR signal transduction in primary human B cells to identify and functionally characterize key processes with a particular focus on the functional interplay of cytoskeleton-related processes and BCR signaling events that we will study in a comparative manner between normal human B cells and lymphoma cells. We will address this aim with three subprojects that will be carried out in close collaboration between our laboratories at the Universities of Göttingen and Frankfurt.Based on our previous studies of tonic BCR signaling in BL cells, our first subproject will aim at investigating tonic BCR signaling in normal human B cells by phosphoproteomics. By using specific small molecule inhibitors, we will address the role of the BCR-proximal effectors as well as the cytoskeleton in the context of tonic BCR signaling. We will systematically compare these results with the phosphoproteome data that we have obtained for tonic BCR signaling in lymphoma cells.In the second subproject, we will, based on our preliminary data, mechanistically characterize tonic BCR signaling processes on a molecular level. We will generate B cell lines that are deficient for expression of certain signal effectors. Because of the pivotal role of the actin cytoskeleton in controlling BCR signals, we will mainly focus on candidate proteins that are putative regulators of the cytoskeleton. The cell lines will allow us to analyze the role of certain cytoskeletal regulators in tonic BCR signal transduction and will be used to perform comparative analyses in primary human B cells.In the third subproject we will study spacial aspects of tonic BCR signaling. Here, we will focus on the processes that stabilize preformed signaling complexes to control tonic BCR signaling with a particular focus on the role of the actin-based cytoskeleton.