Dissociation of in vitro DNA deamination activity and physiological functions of AID mutants
Dissociation of in vitro DNA deamination activity and physiological functions of AID mutants
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DOI：
10.1073/pnas.0806641105
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发表时间：
2008-10-14
影响因子：
11.1
通讯作者：
Honjo, Tasuku
中科院分区：
文献类型：
--
作者：
Shivarov, Velizar;Shinkura, Reiko;Honjo, Tasuku
激活诱导型胞苷脱氨酶(AID)是DNA裂解所必需的，DNA裂解同时启动了Ig基因的体细胞超突变(SHM)和类切换重组(CSR)。已经提出了两种可供选择的AID切割DNA的机制：RNA编辑和DNA脱氨。为了支持后者，AID在无细胞系统中具有DNA脱氨活性，这被认为是代表其生理功能的。为了验证这一假设，我们产生了各种小鼠AID突变体，并比较了它们的DNA脱氨、CSR和SHM活性。在这里，我们比较了不同AID突变体的DNA脱氨基、CSR和SHM活性，发现它们的大部分CSR或SHM活性与他们的DNA脱氨基活性不成比例。具体地说，我们鉴定了一组突变体(H48A、L49A、R50A和N51A)，它们的DNA脱氨活性较低，但CSR活性相对完整。值得注意的是，一个AID突变体(N51a)保留了CSR功能，但失去了DNA脱氨活性。此外，与AID的N51同源的N57位的APOBEC1突变也取消了DNA脱氨活性，但保留了RNA编辑活性。这些结果表明，DNA脱氨活性并不能代表AID的生理功能。
Activation-induced cytidine deaminase (AID) is essential for the DNA cleavage that initiates both somatic hypermutation (SHM) and class switch recombination (CSR) of the Ig gene. Two alternative mechanisms of DNA cleavage by AID have been proposed: RNA editing and DNA deamination. In support of the latter, AID has DNA deamination activity in cell-free systems that is assumed to represent its physiological function. To test this hypothesis, we generated various mouse AID mutants and compared their DNA deamination, CSR, and SHM activities. Here, we compared DNA deamination, CSR, and SHM activities of various AID mutants and found that most of their CSR or SHM activities were disproportionate with their DNA deamination activities. Specifically, we identified a cluster of mutants (H48A, L49A, R50A, and N51A) with low DNA deamination activity but relatively intact CSR activity. Of note is an AID mutant (N51A) that retained CSR function but lost DNA deamination activity. In addition, an APOBEC1 mutation at N57, homologous to N51 of AID, also abolished DNA deamination activity but retained RNA editing activity. These results indicate that DNA deamination activity does not represent the physiological function of AID.