BRIGHT FUNCTION IN IMMUNODEFICIENCY DISEASE

X连锁免疫缺陷(XID)小鼠和X连锁人类 无丙种球蛋白血症(XLA)，表现为血清免疫球蛋白水平较低 而不是普通人。许多患者缺乏可检测到的B淋巴细胞。 布鲁顿酪氨酸激酶(BTK)在XID和XLA中都发生了突变，但 这种激酶的突变导致B细胞缺陷的机制是 未知。Bright，(免疫球蛋白重链B细胞调节因子 转录)，是一个70 kDa的DNA结合蛋白，主要在B 淋巴细胞。它与小鼠免疫球蛋白内的几个区域结合 重链基因，并一直与增加在 免疫球蛋白RNA水平。明亮的表达和DNA结合活性可以 在正常成人脾屏幕上被多个刺激物诱导。近期 研究表明，来自XID小鼠的受刺激的淋巴细胞不能产生 明亮的DNA结合活性；即使不存在明亮的蛋白质。 其他实验表明，Bright和BTK在正常情况下直接相互作用 老鼠。因此，光明的活动可能需要一个功能正常的BTK。 拟议的研究将解决这样的假设，即缺陷的BTK导致 可能至少部分解释了低谷的不活跃的光明形式 在XID中观察到血清免疫球蛋白水平，进而在XLA中观察到。这个 具体目标是。1)研究BTK对Bright DNA的重要性- 通过共表达、免疫沉淀和鉴定获得结合活性 翻译后修饰，2)以确定功能是否正确 在XLA患者的B细胞中存在，使用迁移率变化分析，3)到 通过产生Xid表型确定Bright与Xid表型的关系 Bright的显性阴性形式，然后在转基因小鼠中表达， 4)确定Bright在免疫球蛋白基因座中的功能 体外转录和拓扑异构酶分析，以及5)鉴定 受迁移率的光明相互作用潜在调节的其他基因 移位试验和抗体促进了克隆。这些研究将提供 对Bright在XID和人类中的潜在作用的重要新见解 免疫缺陷疾病，XLA，并将有助于我们理解 免疫球蛋白基因调控。

X-linked immunodeficient (xid) mice and humans with X-linked agammaglobulinemia (XLA), exhibit lower levels of serum immunoglobulin than normal individuals. Many patients lack detectable B lymphocytes. Bruton's tyrosine kinase (BTK) is mutated in both xid and XLA, but the mechanism by which mutations in this kinase cause B cell defects is unknown. Bright, (B cell regulator of immunoglobulin heavy chain transcription), is a 70 kDa DNA-binding protein expressed primarily in B lymphocytes. It binds to several regions within the murine immunoglobulin heavy chain locus, and has been associated with increases in immunoglobulin RNA levels. Bright expression and DNA-binding activity can be induced in normal adult spleen screens by a number of stimuli. Recent studies showed that stimulated lymphocytes from xid mice did not produce Bright DNA-binding activity; even though Bright protein was not present. Other experiments suggest that Bright and BTK interact directly in normal mice. Thus, Bright activity may require a functional BTK. The proposed studies will address the hypothesis that defective BTK leads to inactive forms of Bright that might at least partially explain the low serum immunoglobulin levels observed in xid, and by extension, XLA. The specific aims are. 1) to investigate the importance of BTK for Bright DNA- binding activity by co-expression, immunoprecipitation, and identification of post-translational modifications, 2) to determine if functional Bright is present in B cells from XLA patients using mobility shift assays, 3) to determine the relationship of Bright to the xid phenotype by producing dominant negative forms of Bright and expressing then in transgenic mice, 4) to determine how Bright functions in the immunoglobulin locus using in vitro transcription and topoisomerase assays, and 5) to identify additional genes potentially regulated by Bright interactions by mobility shift assay and antibody facilitated cloning. These studies will provide important new insights into Bright's potential role in xid and the human immunodeficiency disease, XLA, and will contribute to our understanding of immunoglobulin gene regulation.