Molecular Studies on Antigen Receptor Gene Recombination

本研究的主要目的是确定Endo-SR（参与开关/体细胞重组的内切酶）在b细胞特异性基因重组或其他DNA加工事件中的作用。Endo-SR最初是通过其优先切割免疫球蛋白（lg）开关重复序列的能力被发现的，这些序列通常在lg同型开关重组断点上发现。该酶已从牛脾核提取物中纯化到均匀性，并从纯化蛋白中获得部分肽序列信息。对这些肽进行的序列同源性搜索显示，它们与人类和秀丽隐杆线虫基因组计划中假设的人类和秀丽隐杆线虫蛋白质的预测氨基酸序列有显著的相似性。最近有报道称，鉴定出的人类蛋白编码一种与程序性细胞死亡过程中诱导的核DNA降解有关的内切酶。初步证据表明，秀丽隐杆线虫蛋白也编码一种参与凋亡核DNA降解的内切酶。最近对Endo-SR基因的分离将极大地促进对该酶的体内调控和功能的严格分析。作为该分析的一部分，将通过靶向基因破坏产生Endo-SR突变细胞系。对这些Endo-SR缺陷细胞进行的实验应该可以得出明确的结论，即这种核酸酶活性是否需要开关重组和/或凋亡。这项研究将极大地提高目前对一种酶的分子和生化作用的理解，这种酶在淋巴细胞特异性DNA重组过程和细胞凋亡诱导的一般DNA降解过程中都有牵连。

The major goal of this research is to determine the role of Endo-SR (Endonuclease implicated in Switch/Somatic Recombination) in B-cell specific gene recombination or other DNA processing events. Endo-SR was initially discovered through its ability to preferentially cleave immunoglobulin (lg) switch repeat sequences commonly found at lg isotype-switch recombination breakpoints. This enzyme has been purified to homogeneity from bovine spleen nuclear extracts and partial peptide sequence information from the purified protein has been obtained. Sequence homology searches performed with these peptides revealed significant similarities to the predicted amino acid sequences of hypothetical human and C. elegans proteins identified by the Human and C. elegans Genome Projects. The identified human protein has recently been reported to encode an endonuclease implicated in nuclear DNA degradation induced during programmed cell death. Preliminary evidence indicates that the C. elegans protein also encodes an endonuclease implicated in apoptotic nuclear DNA degradation. The recent isolation of the Endo-SR gene should greatly facilitate a rigorous analysis of the in vivo regulation and function of this enzyme. As part of this analysis, Endo-SR mutant cell lines will be generated by targeted gene disruption. Assays conducted with these Endo-SR deficient cells should allow definitively conclusions as to whether or not this nuclease activity is required for switch recombination and/or apoptosis. This research should greatly enhance current understanding of the molecular and biochemical roles of an enzyme that has been implicated in both a lymphocyte-specific DNA recombination process and a general DNA degradation process induced by apoptosis.