DNA breaks in hypermutating immunoglobulin genes: evidence for a break-and-repair pathway of somatic hypermutation.
DNA breaks in hypermutating immunoglobulin genes: evidence for a break-and-repair pathway of somatic hypermutation.
复制标题
超突变免疫球蛋白基因中的 DNA 断裂：体细胞超突变断裂和修复途径的证据。
DOI：
10.1093/genetics/158.1.369
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发表时间：
2001
期刊：
影响因子：
3.3
通讯作者：
Maizels,N
中科院分区：
文献类型：
--
作者：
Kong,Q;Maizels,N
为了验证免疫球蛋白基因在体内的高突变采用一种途径，即DNA断裂被引入并随后修复以产生突变，我们使用了基于聚合酶链式反应的方法来检测和鉴定活跃高突变的原代小鼠生发中心B细胞的λ1基因的单链DNA断裂。我们发现，与非高突变B细胞相比，高突变B细胞的λ-1基因存在2-3倍的断裂，并且1.3%的生发中心B细胞含有与高突变相关的λ-1基因断裂。在顶部和底部的DNA链上都发现了断裂，并定位于λ1的活跃超突变区域，但双链断裂只占已发现的断裂的一部分。在高突变的B细胞中，几乎一半的断裂发生在热点，在这些热点上，发现了两个或更多独立的断裂。断裂点热点与特征序列基序相关：富嘧啶基序，RCTYT或CCYC；RGYW，与高突变热点相关的序列基序。在断裂点热点确定的序列基序应该为底物的设计提供信息，以表征参与超突变途径的活动。
To test the hypothesis that immunoglobulin gene hypermutationin vivoemploys a pathway in which DNA breaks are introduced and subsequently repaired to produce mutations, we have used a PCR-based assay to detect and identify single-strand DNA breaks in λ1 genes of actively hypermutating primary murine germinal center B cells. We find that there is a two- to threefold excess of breaks in λ1 genes of hypermutating B cells, relative to nonhypermutating B cells, and that 1.3% of germinal center B cells contain breaks in the λ1 gene that are associated with hypermutation. Breaks were found in both top and bottom DNA strands and were localized to the region of λ1 that actively hypermutates, but duplex breaks accounted for only a subset of breaks identified. Almost half of the breaks in hypermutating B cells occurred at hotspots, sites at which two or more independent breaks were identified. Breaksite hotspots were associated with characteristic sequence motifs: a pyrimidine-rich motif, eitherRCTYT or CCYC; andRGYW, a sequence motif associated with hypermutation hotspots. The sequence motifs identified at breaksite hotspots should inform the design of substrates for characterization of activities that participate in the hypermutation pathway.