Replication stalling at Friedreich's ataxia (GAA)n repeats in vivo
Replication stalling at Friedreich's ataxia (GAA)n repeats in vivo
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DOI：
10.1128/mcb.24.6.2286-2295.2004
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发表时间：
2004-03-01
影响因子：
5.3
通讯作者：
Mirkin, SM
中科院分区：
文献类型：
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作者：
Krasilnikova, MM;Mirkin, SM
将不同长度的friedrich 's ataxia (GAA)(n)重复序列克隆到酿酒酵母（Saccharymyces cerevisiae）质粒中，利用复制中间体双向电泳分析其对DNA复制的影响。我们发现突变前和疾病大小的重复序列在体内阻止了复制叉的进展，而正常大小的重复序列不影响复制。值得注意的是，观察到的酵母复制停滞的阈值重复长度（类似于40个重复）与人类重复扩增的阈值长度非常匹配。此外，复制停滞明显依赖于方向，只有当重复序列的同嘌呤链作为滞后链模板时才会出现。最后，发现（GAA）(n)（。）（TTC）(n)在扩张和收缩中都重复，重复的方向急剧增加，这是导致复制停滞的原因。这些数据首次直接证明了（GAA）(n)重复序列对体内DNA复制的影响。我们认为，体内重复引起的复制衰减是由于三重结构的形成。复制停滞与重复序列长度多态性之间的明显联系为重复序列扩展提供了一种新的模型。
Friedreich's ataxia (GAA)(n) repeats of various lengths were cloned into a Saccharymyces cerevisiae plasmid, and their effects on DNA replication were analyzed using two-dimensional electrophoresis of replication intermediates. We found that premutation- and disease-size repeats stalled the replication fork progression in vivo, while normal-size repeats did not affect replication. Remarkably, the observed threshold repeat length for replication stalling in yeast (similar to40 repeats) closely matched the threshold length for repeat expansion in humans. Further, replication stalling was strikingly orientation dependent, being pronounced only when the repeat's homopurine strand served as the lagging strand template. Finally, it appeared that length polymorphism of the (GAA)(n) (.) (TTC)(n) repeat in both expansions and contractions drastically increases in the repeat's orientation that is responsible for the replication stalling. These data represent the first direct proof of the effects of (GAA)(n) repeats on DNA replication in vivo. We believe that repeat-caused replication attenuation in vivo is due to triplex formation. The apparent link between the replication stalling and length polymorphism of the repeat points to a new model for the repeat expansion.