Regulation of B Cell Gene Expression Patterns by Btk

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DESCRIPTION( provided by the applicant): Bruton's component B tyrosine kinase (Btk) is a critical of signaling pathways that regulate B cell development and function. Mutations in Btk cause the cell immunodeficiency's X-linked a gammaglobulinemia (XLA) in humans and X-linked immunodeficiency (xid) in mice. Btk is also required for the production of autoantibodies and the efficient degranulation of mast cells. A thorough understanding of the signaling pathways downstream of Btk is likely to reveal new therapeutic targets for B cell immunodeficiencies,autoimmunity, and allergy. This study will test the hypothesis that impaired regulation of gene expression in response to BCR crosslinking results in the B cell developmental and functional defects observed in the absence of Btk. B cells from wild type and Btklo mice, which express a limiting dosage of Btk, will be compared. Unlike Btk-/- or xid mice, Btklo mice have normal numbers of mature B cells. These cells remain functionally impaired due to insufficient levels of Btk, allowing Btk-dependent signaling events to be identified in mature B cells. We have begun to characterize Btk-dependent changes in BCR-induced gene expression using cDNA microarray analysis. Our first aim is to confirm these results using real-time RT-PCR and Western blots and to define which branches of Btk signaling pathways are involved in the regulation of these genes. The role of Btk-regulated genes in mediating various functional outcomes of Btk-initiated signals will be assessed as follows. Btk-regulated genes will be expressed as transgenes in Btk-/-, Btk lo, and wild type mice and their ability to restore Btk-dependent defects in conventional B cell development and function observed. To assess the role of Btk-regulated genes in B-1 cell differentiation, transgenes expressing Btk regulated genes will be crossed to Btk-/-, Btklo, and wild type mice carrying a VH12 anti-phosphatidylcholine Ig transgene. This transgene drives the clonal expansion of phosphatidylcholine-specific B cells and their differentiation into the B-1 compartment in a Btk-dependent manner. These studies will define the mechanism by which Btk-mediated transmission of BCR signals regulates the development and function of both conventional B and B-1 cells and potentially identify novel therapeutic approaches for diseases involving B cell disregulation.