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）：初步研究已确定 可以在细胞上表达的一组不寻常的人μ重链 在没有替代光或常规光的情况下，作为预BCR的一部分的表面 店这些异常的mu重链，首先在一项研究中被鉴定， 前BCR信号传导缺陷的患者似乎能够激活细胞 并诱导轻链基因重排，但不促进扩增 或B细胞前体的长期存活。轻链独立链 具有预期的分子量并具有完整的VH和CH 1结构域。 这些μ重链的转录物可以在VH 3和VH 4中找到。 家庭它们占重排的VH 3的大约10%， 在正常的人前B细胞中存在μ重链基因;然而，在正常的人前B细胞中没有发现它们。 正常前B细胞或成熟B细胞。拟议的实验将 对这些异常重链进行生化表征， 他们的回应。具体目标是：（1）确定特定的 允许细胞表面表达的V（D）J重排内的序列，或 在没有轻链的情况下分泌。异常穆海的束缚 还将检查ER伴侣BiP的链; 2）使用两种培养物 系统，RAG 2缺陷型前B细胞的逆转录病毒感染和四环素 BCR在Jurkat T细胞中的可诱导重建，以评估其作用 细胞表面异常μ重链的表达 活化标志物，信号转导磷酸化的改变 分子和转录本数量的变化， 原B细胞或前B细胞分化阶段;和3）产生转基因的 表达正常或异常mu重链并表征B细胞的小鼠 在这些老鼠身上的发展。转基因小鼠将与 对于所涉及的信号转导蛋白或分子是无效的或转基因的 细胞凋亡更好地了解这些不寻常的生物化学性质 mu重链和机制，他们被删除应阐明 正常前B细胞向前B细胞转化的要求。拟议 这些研究亦会提供有关 正常和异常抗体库。