Tsai Chia-Lun, Drejer Anna H, Schatz David G
Department of Molecular Biophysics and Biochemistry, Section of Immunobiology, Yale University School of Medicine, New Haven, Connecticut 06510, USA.
Genes Dev. 2002 Aug 1;16(15):1934-49. doi: 10.1101/gad.984502.
In addition to creating the DNA double strand breaks that initiate V(D)J recombination, the RAG proteins are thought to play a critical role in the joining phase of the reaction. One such role, suggested by in vitro studies, might be to ensure the structural integrity of postcleavage complexes, but the significance of such a function in vivo is unknown. We have identified RAG1 mutants that are proficient in DNA cleavage but defective in their ability to interact with coding ends after cleavage and in the capture of target DNA for transposition. As a result, these mutants exhibit severe defects in hybrid joint formation, hairpin coding end opening, and transposition in vitro, and in V(D)J recombination in vivo. Our results suggest that the RAG proteins have an architectural function in facilitating proper and efficient V(D)J joining, and a protective function in preventing degradation of broken ends prior to joining.
除了产生引发V(D)J重组的DNA双链断裂外，RAG蛋白被认为在该反应的连接阶段发挥关键作用。体外研究表明，其中一个作用可能是确保切割后复合物的结构完整性，但这种功能在体内的意义尚不清楚。我们已经鉴定出RAG1突变体，它们在DNA切割方面表现正常，但在切割后与编码末端相互作用以及捕获用于转座的靶DNA的能力方面存在缺陷。因此，这些突变体在体外杂交接头形成、发夹编码末端打开和转座以及体内V(D)J重组方面表现出严重缺陷。我们的结果表明，RAG蛋白在促进正确和高效的V(D)J连接方面具有结构功能，并且在连接之前防止断裂末端降解方面具有保护功能。
