Molecular Studies on Antigen Receptor Gene Recombination

本研究的主要目的是确定Endo-SR（与开关/体细胞凋亡有关的内切酶）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（Ig）开关重复序列的能力而发现的，该重复序列通常存在于Ig同种型开关重组断点处。 这种酶已被纯化到同质从牛脾核提取物和部分肽序列信息，从纯化的蛋白质已经获得。 用这些肽进行的序列同源性搜索显示与假设的人和C. elegans蛋白质鉴定的人和C. elegans Genome Projects. 最近有报道称，已鉴定的人类蛋白质编码一种核酸内切酶，该核酸内切酶与程序性细胞死亡期间诱导的核DNA降解有关。初步证据表明，C。elegans蛋白还编码一种与凋亡核DNA降解有关的核酸内切酶。最近分离的Endo-SR基因应大大有利于严格分析这种酶的体内调节和功能。 作为该分析的一部分，将通过靶向基因破坏产生Endo-SR突变体细胞系。 用这些Endo-SR缺陷细胞进行的测定应允许关于开关重组和/或凋亡是否需要这种核酸酶活性的明确结论。 这项研究应大大提高目前的分子和生化作用的酶，已牵连在淋巴细胞特异性DNA重组过程和一般的DNA降解过程诱导的细胞凋亡的理解。

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.