STUDIES OF IMMUNOGLOBULIN GENE REARRANGEMENT

在淋巴发育过程中，功能性免疫球蛋白和T细胞受体基因通过一种称为V(D)J重组的过程从基因片段组装而来，这是产生多样化免疫反应所必需的。在这个反应的第一阶段，RAG1/RAG2蛋白复合体在目标位置进行特定的双链断裂。重新连接所需的后续步骤使用了许多蛋白质因子，这些蛋白质因子也用于修复X射线损伤。我们继续研究这两条途径的一半1)我们之前描述了RAG蛋白进行的转座重组是一种副反应，并认为这种异常活动可能是白血病和淋巴瘤染色体易位的一个来源。我们现在已经使用生化分析来证明RAG蛋白能够引起这种易位，尽管这还没有在体内得到证实。2)已知Xrcc4蛋白与DNA连接酶IV结合，并刺激其在V(D)J重组或辐射损伤修复中的活性。Xrcc4蛋白的晶体结构现在已经与杨伟强和M·朱诺普合作确定，类似于参与DNA代谢许多其他方面的SMC蛋白家族。DNA连接酶IV的结合部位位于Xrcc4.3的长螺旋茎中)Mre11/Rad50/Nbs1复合体参与了许多类型的遗传重组。我们现在已经证明，Mre11核酸酶可以产生在V(D)J重组连接中常见的短同源。Mre11作为一种二聚体连接两个DNA末端，使它们能够检测彼此的序列，当同源性被发现时，核酸酶暂停以允许连接。4)组蛋白变异体H2AX的磷酸化是DNA损伤后的早期事件。我们现在已经证明(与NCI的W.Bonner和E.Rogakou合作)，辐射诱导的磷酸化H_2AX核焦点是涉及多种修复因子的焦点的前体。这一结果将染色质水平上的初始信号与DNA的后来修复联系在一起。细胞101,625-633.Paull，T.和Gellert，M.(2000)。程序娜塔莉。理工学院。U S A 97,6409-6414保尔，T.T.等.(2000)Curr.比奥尔。10,886-895。

During lymphoid development, functional immunoglobulin and T cell receptor genes are assembled from gene segments by a process called V(D)J recombination, which is essential for generating the diversity of the immune response. In the first stage of this reaction, specific double-strand breaks are made at the target sites by the RAG1/RAG2 protein complex. The later steps necessary for rejoining employ many protein factors that are also used for repair of X-ray damage. We continue to investigate both halves of the pathway.1) We previously described a transpositional recombination carried out by the RAG proteins as a side reaction, and suggested this aberrant activity as a possible source of chromosomal translocations in leukemias and lymphomas. We have now used a biochemical assay to show that the RAG proteins are capable of causing such translocations, although this has not yet been demonstrated in vivo.2) The Xrcc4 protein is known to bind to DNA ligase IV and to stimulate its activity in V(D)J recombination or the repair of radiation damage. The crystal structure of the Xrcc4 protein, which has now been determined in collaboration with W. Yang and M. Junop, resembles the SMC family of proteins involved in many other aspects of DNA metabolism. The binding site for DNA ligase IV lies in the long helical stem of Xrcc4.3) The Mre11/Rad50/Nbs1 complex is known to be involved in many types of genetic recombination We have now shown that the Mre11 nuclease can produce the short homologies often found in V(D)J recombination junctions. Mre11 acts as a dimer to bridge two DNA ends, allows them to sense each other's sequence, and the nuclease pauses to allow ligation when a homology is revealed.4) Phosphorylation of the histone variant H2AX is an early event after DNA damage. We have now shown (in collaboration with W. Bonner and E. Rogakou of NCI) that radiation-induced nuclear foci of phosphorylatd H2AX are precursors of foci involving multiple repair factors. This result links initial signaling at the level of chromatin to later repair of DNA.Melek, M., and Gellert, M. (2000). Cell 101, 625-633.Paull, T. T., and Gellert, M. (2000). Proc. Natl. Acad.Sci. U S A 97, 6409-6414.Paull, T.T. et al. (2000) Curr. Biol. 10, 886-895.