MOLECULAR BASIS OF IMMUNOGLOBULIN GENE HYPERMUTATION

免疫前抗体谱系的进一步多样化是由 免疫球蛋白(Ig)基因体细胞突变的独特过程。这 流程紧密聚焦于重新安排的VDJ细分市场和即时 侧翼序列。突变率接近10-3/b.p/代 比其他估计的要大四个数量级 真核基因。这一过程可能是导致 自身免疫性疾病中的自身反应性抗体克隆 与B细胞肿瘤中某些癌基因的激活有关。这个 免疫球蛋白基因体细胞超突变的生物学基础尚不清楚。我们建议 通过克隆定位负责这一过程的Ig基因座区域 接近标记基因sup F突变的基因部分 由Ig启动和聚焦的片段可能会被引入到 这种侧翼靶基因类似于自然发生的VDJ 在染色体易位部位的连接和非Ig序列中。 这些变化将被一种敏感的微生物测试识别，当 利用该表达载体转化大肠杆菌指示菌株。一系列 的载体系统将被用来循环Ig片段通过各种 从细胞系到转基因动物的淋巴环境。 不同V区启动的突变率的比较可能允许 识别序列的检测对这一过程高度敏感。 对不同细胞系中发生的突变进行比较可能会发现 发育受调节的细胞成分的存在 影响突变过程。除了识别细胞 以及免疫球蛋白超突变的分子基础，这些实验将确定 无抗原条件下这一过程对抗体多样性的影响 选择。这些研究将导致对这一角色的更广泛的理解 免疫球蛋白突变在正常免疫反应中的潜在作用 自身免疫和B细胞恶性肿瘤，并将提供重要的见解 DNA诱变的更一般的方面。

Further diversification of the preimmune antibody repertoire is provided by a unique process of immunoglobulin (Ig) gene somatic mutation. This process is tightly focused to rearranged VDJ segments and immediate flanking sequences. The mutation rate approaches 10-3/b.p./generation which is four orders of magnitude greater than that estimated for other eukaryotic genes. This process may be responsible for the development of self reactive antibody clones in autoimmune disorders and has also been implicated in the activation of certain oncogenes in B cell tumors. The biologic bases of Ig gene somatic hypermutation is unknown. We propose to localize regions of the Ig locus responsible for this process by cloning portions of the gene in close proximity to a marker gene, sup F. Mutations initiated and focused by Ig fragments may be expected to be introduced into this flanking target gene similar to what occurs in naturally occurring VDJ joinings and in non Ig sequences at the site of chromosomal translocations. Such changes will be recognized by a sensitive microbiological assay when this plasmid is used to transform indicator strains of E. Coli. A series of vector systems will be used to cycle Ig segments through various lymphoid environments ranging from cell lines to transgenic animals. Comparison of mutation rates initiated by different V regions may allow detection of recognition sequences highly sensitive to this process. Comparison of mutations that occur in different cell lineages may identify the presence of developmentally regulated cellular components which influence the mutational process. In addition to identifying the cellular and molecular basis of Ig Hypermutation, these experiments will determine the impact of this process on antibody diversity in the absence of antigen selection. These studies will lead to a broader understanding of the role of Ig mutation in the normal immune response, its potential contribution to autoimmunity and B cell malignancy, and will provide important insights into more general aspects of DNA mutagenesis.