Shen Hong Ming, Storb Ursula
Department of Molecular Genetics and Cell Biology, University of Chicago, 920 East 58th Street, Chicago, IL 60637, USA.
Proc Natl Acad Sci U S A. 2004 Aug 31;101(35):12997-3002. doi: 10.1073/pnas.0404974101. Epub 2004 Aug 24.
The activation-induced cytidine deaminase (AID) is required for somatic hypermutation (SHM) and class-switch recombination of Ig genes. It has been shown that in vitro, AID protein deaminates C in single-stranded DNA or the coding-strand DNA that is being transcribed but not in double-stranded DNA. However, in vivo, both DNA strands are mutated equally during SHM. We show that AID efficiently deaminates C on both DNA strands of a supercoiled plasmid, acting preferentially on SHM hotspot motifs. However, this DNA is not targeted by AID when it is relaxed after treatment with topoisomerase I, and thus, supercoiling plays a crucial role for AID targeting to this DNA. Most of the mutations are in negatively supercoiled regions, suggesting a mechanism of AID targeting in vivo. During transcription the DNA sequences upstream of the elongating RNA polymerase are negatively supercoiled, and this transient change in DNA topology may allow AID to access both DNA strands.
激活诱导的胞嘧啶脱氨酶（AID）是免疫球蛋白基因体细胞高频突变（SHM）和类别转换重组所必需的。研究表明，在体外，AID蛋白可使单链DNA或正在转录的编码链DNA中的C发生脱氨基作用，但不会使双链DNA中的C发生脱氨基作用。然而，在体内，SHM过程中两条DNA链发生的突变是均等的。我们发现，AID能有效地使超螺旋质粒的两条DNA链上的C发生脱氨基作用，且优先作用于SHM热点基序。然而，在用拓扑异构酶I处理后该DNA松弛时，AID不会靶向此DNA，因此，超螺旋对AID靶向该DNA起着关键作用。大多数突变发生在负超螺旋区域，这提示了AID在体内的靶向机制。在转录过程中，延伸的RNA聚合酶上游的DNA序列呈负超螺旋状态，而DNA拓扑结构的这种瞬时变化可能使AID能够接触到两条DNA链。
