STUDIES OF IMMUNOGLOBULIN GENE REARRANGEMENT

免疫球蛋白基因重排的研究

• 批准号: 2573012
• 负责人: M GELLERT
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/2573012
• 关键词: DNA binding protein; cell free system; gene rearrangement; immunoglobulin genes; nucleic acid sequence

The V(D)J recombination process that assembles functional immunoglobulin and T cell receptor genes in lymphoid cells is essential for generating the diversity of the immune response. The reaction is now known to occur in two stages. In the first stage, specific double-strand breaks are made at the target sites. The ends of the coding DNA sequence at these breaks are always joined back on themselves as DNA hairpins. Based on cellular experiments, this unusual type of cleavage was known to utilize the products of the RAG1 and RAG2 genes, but it was not clear how they functioned. The later stages of recombination are less specific; the final joined products are made with the use of many factors also involved in the repair of radiation damage. We have now reproduced the first cleavage reaction in a biochemically defined reaction requiring only the purified RAG-1 and RAG-2 proteins. A DNA fragment containing the recognition signal sequence (RSS) is cut to make a hairpin coding end and a blunt signal end, just as in vivo. Furthermore, in more demanding reaction conditions, cleavage requires a pair of RSSs of the different types that are needed for recombination in vivo. Thus the RAG proteins alone contain all the information for the specificity of V(D)J recombination. We have also found that the hairpins are made in a reaction very similar chemically to transpositional recombination. An evolutionary relation between V(D)J recombination and transposable elements had previously been suggested, and our results provide the strongest data so far in support of this idea.

V（D）J重组过程， 免疫球蛋白和T细胞受体基因在淋巴细胞中的表达， 对产生免疫反应的多样性至关重要。 的 现在已知反应分两个阶段进行。 在第一阶段， 在靶位点产生特异性双链断裂。端部 这些断裂处的DNA编码序列总是连接在一起， DNA发夹 根据细胞实验， 已知一种不寻常的裂解类型利用RAG 1的产物 和RAG 2基因，但尚不清楚它们如何发挥作用。 后期 重组的阶段不那么具体;最终的结合产物 是利用许多因素也参与修复 辐射损伤 我们现在已经重现了第一次裂解反应 在生化定义的反应中，仅需要纯化的RAG-1 和RAG-2蛋白。 含有识别信号的DNA片段 切割一个发夹序列（RSS）以产生一个发夹编码末端和一个钝信号 就像在体内一样。 此外，在要求更高的反应中， 条件下，切割需要一对不同类型的RSS 是体内重组所必需的。 因此，RAG蛋白 单独包含V（D）J特异性的所有信息 重组 我们还发现，发夹是在一个 化学反应与转座重组非常相似。一个 V（D）J重组与转座的进化关系 元素之前已经提出，我们的结果提供了 迄今为止最有力的数据支持了这一观点。