Shen Hong Ming
Department of Molecular Genetics and Cell Biology, University of Chicago, Chicago, IL 60637, USA.
Mol Immunol. 2007 Feb;44(5):974-83. doi: 10.1016/j.molimm.2006.03.015. Epub 2006 May 11.
Activation-induced cytidine deaminase (AID) is likely responsible for DNA cytidine deamination, although it may also act as an RNA deaminase. It functions on single-stranded DNA, the non-template strand in double-stranded DNA during transcription, or both strands in supercoiled DNA. To ask whether AID is able to deaminate cytidine at DNA breaks, plasmids, containing a SnaBI site (TAC downward arrowGTA) that forms blunt ends after digestion with SnaBI, were generated. If AID deaminates cytidine at the upstream blunt end, the ATG start codon in either of two drug resistance genes will be regenerated after ligation and replication in UDG-null E. coli cells. This study shows that AID targets cytidine at the break. The extent of deamination activity beyond the break is correlated with the base composition in the break region. If the break region is A, T-rich, C > T transitions are extensive. However, when the break region is not A, T-rich, mutations are mainly restricted to the break, similar to findings in vivo. The results indicate that AID has activity on double strand breaks (DSBs). Based on previous and current findings, a somatic hypermutation (SHM) model is proposed, in which collision between the transcription apparatus and the replication fork generates DSBs. After AID acts on break ends, the error-prone DNA repair machinery fixes and creates mutations.
激活诱导的胞苷脱氨酶（AID）可能负责DNA胞苷脱氨，尽管它也可能作为RNA脱氨酶起作用。它作用于单链DNA、转录过程中双链DNA的非模板链或超螺旋DNA的两条链。为了探究AID是否能够在DNA断裂处使胞苷脱氨，构建了含有SnaBI位点（TAC向下箭头GTA）的质粒，该位点在用SnaBI消化后形成平端。如果AID在上游平端使胞苷脱氨，那么在无尿嘧啶DNA糖基化酶（UDG）的大肠杆菌细胞中进行连接和复制后，两个耐药基因中的任何一个中的ATG起始密码子将被重新生成。这项研究表明AID靶向断裂处的胞苷。断裂处之外的脱氨活性程度与断裂区域的碱基组成相关。如果断裂区域富含A、T，C>T转换就会广泛发生。然而，当断裂区域不富含A、T时，突变主要局限于断裂处，这与体内的发现相似。结果表明AID对双链断裂（DSB）有活性。基于之前和当前的发现，提出了一种体细胞超突变（SHM）模型，其中转录装置与复制叉之间的碰撞产生DSB。在AID作用于断裂末端后，易出错的DNA修复机制进行修复并产生突变。
