Sadofsky-RAG1 N-terminal domain binding interactions

描述（由申请人提供）：脊椎动物的适应性免疫系统依赖于称为V（D）J重组的DNA重组系统，以产生免疫球蛋白和T细胞受体的库.这一重要步骤在误用时也会导致染色体易位。这种重组过程的大部分调节发生在DNA被切割的初始阶段。蛋白质RAG 1和RAG 2形成位点特异性核酸酶，其切割适当的DNA靶标，并且据信协调重组反应。RAG 1蛋白具有一个大的N-末端结构域，包含一个RING基序，其作为DNA结合蛋白和核酸酶的已知酶功能是可消耗的。我已经表明，这个域可以作为一个泛素连接酶，并提出它起着调节作用。这一提议将使实验室能够研究RAG 1 N-末端结构域与通过双杂交筛选鉴定的几种蛋白质之间的相互作用。将检查几个属性，包括修改状态，诱导的变化，在亚细胞定位，蛋白质周转，以及是否与这些蛋白质的相互作用影响的重组目标的选择。我们的目标是确定和表征涉及RAG 1 N-末端结构域的蛋白质相互作用。预计这些将调节免疫系统发育的关键步骤，并可能导致与该过程相关的基因组不稳定性。

DESCRIPTION (provided by applicant): The adaptive immune system of vertebrates depends upon a DNA recombination system, called V(D)J recombination, to generate the repertoire of immunoglobulin and T- cell receptors. This essential step also leads to chromosomal translocation when misapplied. Much of the regulation of this recombination process occurs at the initial stage when the DNA is cut. The proteins RAG1 and RAG2 form the site-specific nuclease that cuts the appropriate DNA targets and, it is believed, coordinate the recombination reaction. The RAG1 protein possesses a large N-terminal domain, containing a RING motif, which is expendable with respect to the known enzymatic functions as a DNA binding protein and a nuclease. I have shown that this domain can act as an ubiquitin ligase and propose that it plays a regulatory role. This proposal will allow the laboratory to study the interaction between the RAG1 N-terminal domain and several proteins that have been identified through a two-hybrid screen. Several properties will be examined including modification status, induced changes in subcellular localization, protein turn-over and whether the interaction with these proteins influences the choice of recombination targets. Our goal is to identify and characterize protein interactions involving the RAG1 N- terminal domain. These are expected to regulate a critical step in the development of the immune system, and may contribute to genomic instability associated with that process.