NEGATIVE SELECTION AT THE PRO-B TO PRE-B CELL TRANSITION

PRO-B 到 PRE-B 细胞转变时的负选择

• 批准号: 6511408
• 负责人: MARY ELLEN CONLEY
• 金额: $ 30万
• 依托单位: ST. JUDE CHILDREN'S RESEARCH HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-01 至 2004-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6511408
• 关键词: B cell receptor; B lymphocyte; Retroviridae; autoimmunity; cell differentiation; gene expression; gene rearrangement; genetically modified animals; immunoglobulin genes; laboratory mouse; molecular chaperones; phosphorylation; protein binding; protein biosynthesis; receptor expression; tissue /cell culture

DESCRIPTION (provided by applicant): Preliminary studies have identified an unusual group of human mu heavy chains that can be expressed on the cell surface as part of a pre-BCR in the absence of surrogate or conventional light chains. These aberrant mu heavy chains, which were first identified in a patient with a defect in pre-BCR signaling, appear to be able to activate cells and induce rearrangement of light chain genes but they do not promote expansion or long-term survival of B-cell precursors. The light chain independent chains are of the expected molecular weight and have intact VH and CH1 domains. Transcripts for these muheavy chains can be found in both the VH3 and VH4 families. They constitute approximately 10 percent of the rearranged VH3 muheavy chain genes in normal human pro-B-cells; however, they are not found in normal pre-B-cells or mature B-cells. The proposed experiments will biochemically characterize these aberrant heavy chains and examine the responses elicited by them. The specific aims are 1) determine the particular sequences within the V(D)J rearrangement that allow cell surface expression or secretion in the absence of light chains. The binding of the aberrant muheavy chains to the ER chaperone BiP will also be examined; 2) use two culture systems, retroviral infection of RAG2 deficient pro-B-cells and a tetracycline inducible reconstitution of the BCR in Jurkat T-cells, to evaluate the effects of expression of the aberrant muheavy chain on cell surface expression of activation markers, alterations in phosphorylation of signal transduction molecules and shifts in the amount of transcripts that characterize the pro-B-cell or pre-B-cell stage of differentiation; and 3) produce transgenic mice expressing the normal or aberrant mu heavy chain and characterize B-cell development in these mice. The transgenic mice will be crossed with mice that are null or transgenic for signal transduction proteins or molecules involved in apoptosis. A better understanding of the biochemical nature of these unusual mu heavy chains and the mechanisms by which they are removed should elucidate requirements for the normal pro-B-cell to pre-B-cell transition. The proposed studies will also provide valuable information about the development of the normal and abnormal antibody repertoire.

描述（由申请人提供）：初步研究已经确定 可以在细胞上表达的不寻常的人类MU重链 在没有替代或常规光的情况下，表面是前BCR的一部分 链。这些异常的MU重链，首先在 前BCR信号缺陷的患者似乎能够激活细胞 并诱导轻链基因的重排，但不会促进扩展 或B细胞前体的长期生存。轻链独立连锁 具有预期的分子量，具有完整的VH和CH1结构域。 这些Muheavy链的成绩单都可以在VH3和VH4中找到 家庭。它们约占重新排列VH3的10％ 正常人类pro-b细胞中的muheavy链基因；但是，它们没有发现 正常的前B细胞或成熟的B细胞。提出的实验将 生化是这些异常的重链的特征，并检查 他们引起的回应。具体目的是1）确定特定的 V（d）J重排内的序列，允许细胞表面表达或 在没有光链的情况下分泌。异常的穆希维的结合 还将检查ER伴侣BIP的链条； 2）使用两种文化 系统，RAG2缺乏B-B细胞的逆转录病毒感染和四环素 BCR在Jurkat T细胞中的诱导重构，以评估效果 在细胞表面表达上的异常木链的表达 激活标记，信号转导的磷酸化改变 分子和转录量的转移量 pro-B细胞或B细胞分化阶段； 3）产生转基因 表达正常或异常MU重链并表征B细胞的小鼠 这些小鼠的发育。转基因小鼠将与小鼠交叉 涉及信号转导蛋白或分子的无效或转基因 在凋亡中。更好地理解这些不寻常的生化本质 MU重链及其去除的机制应阐明 对正常的B-Cell前B细胞过渡的要求。提议 研究还将提供有关开发的宝贵信息 正常和异常抗体库。