Regulation of B Cell Gene Expression Patterns by Btk

描述（由申请方提供）：布鲁顿组分B酪氨酸激酶（Btk）是调节B细胞发育和功能的关键信号通路。Btk的突变导致人类的细胞免疫缺陷性X连锁α γ球蛋白血症（XLA）和小鼠的X连锁免疫缺陷性（xid）。Btk也是产生自身抗体和肥大细胞有效脱粒所必需的。深入了解Btk下游的信号通路可能会揭示B细胞免疫缺陷、自身免疫和过敏的新治疗靶点。本研究将检验以下假设：BCR交联导致基因表达调节受损，导致在Btk不存在的情况下观察到B细胞发育和功能缺陷。将比较来自野生型和Btklo小鼠的B细胞，其表达限制剂量的Btk。与Btk-/-或xid小鼠不同，Btklo小鼠具有正常数量的成熟B细胞。这些细胞由于Btk水平不足而保持功能受损，允许在成熟B细胞中鉴定Btk依赖性信号传导事件。我们已经开始使用cDNA微阵列分析来表征BCR诱导的基因表达中Btk依赖性的变化。我们的第一个目标是使用实时RT-PCR和Western印迹来确认这些结果，并确定Btk信号通路的哪些分支参与这些基因的调控。Btk调节基因在介导Btk启动信号的各种功能结果中的作用将评估如下。Btk调节的基因将在Btk-/-、Btk lo和野生型小鼠中表达为转基因，并且观察到它们恢复常规B细胞发育和功能中的Btk依赖性缺陷的能力。为了评估Btk调节基因在B-1细胞分化中的作用，将表达Btk调节基因的转基因与携带VH 12抗磷脂酰胆碱IG转基因的Btk-/-、Btklo和野生型小鼠杂交。该转基因驱动磷脂酰胆碱特异性B细胞的克隆扩增以及它们以Btk依赖性方式分化成B-1区室。这些研究将确定Btk介导的BCR信号传递调节常规B和B-1细胞的发育和功能的机制，并可能确定涉及B细胞失调的疾病的新治疗方法。

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.