STUDIES OF IMMUNOGLOBULIN GENE REARRANGEMENT

在淋巴发育过程中，功能性免疫球蛋白和T细胞受体基因通过称为V（D）J重组的过程从基因片段组装，这对于产生免疫应答的多样性至关重要。在该反应的第一阶段，通过RAG 1/RAG 2蛋白复合物在靶位点处产生特异性双链断裂。随后的步骤需要重新连接使用许多蛋白质因子，这些因子也用于修复X射线损伤。我们继续研究该途径的两半。1）我们之前将RAG蛋白进行的转座重组描述为副反应，并提出这种异常活性可能是白血病和淋巴瘤中染色体易位的来源。我们现在已经使用生物化学测定来显示RAG蛋白能够引起这样的易位，尽管这还没有在体内得到证实。2）已知Xrcc 4蛋白结合DNA连接酶IV并刺激其在V（D）J重组或辐射损伤修复中的活性。Xrcc 4蛋白的晶体结构，现在已经与W. Yang和M. Junop类似于参与DNA代谢的许多其他方面的SMC蛋白质家族。DNA连接酶IV的结合位点位于Xrcc的长螺旋茎4.3） 已知Mre 11/Rad 50/Nbs 1复合物参与许多类型的遗传重组。我们现在已经证明，Mre 11核酸酶可以产生通常在V（D）J重组连接中发现的短同源性。Mre 11作为二聚体桥接两个DNA末端，使它们能够感知彼此的序列，并且当同源性被揭示时，核酸酶暂停以允许连接。4）组蛋白变体H2 AX的磷酸化是DNA损伤后的早期事件。我们现在已经证明（与W。Bonner和E. Rogakou of NCI）认为，辐射诱导的磷酸化H2 AX的核灶是涉及多种修复因子的灶的前体。这一结果将染色质水平的初始信号传导与随后的DNA修复联系起来。和Gellert，M.（2000年）的第10/2000号决议。Cell 101，625- 633. T.，和Gellert，M.（2000年）的第10/2000号决议。Proc. Natl. Acad.Sci. U S A 97，6409- 6414. Paull，T.T.等人（2000）Curr. 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.