Human POT1 is required for efficient telomere C-rich strand replication in the absence of WRN
Human POT1 is required for efficient telomere C-rich strand replication in the absence of WRN
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DOI：
10.1101/gad.544009
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发表时间：
2009-12-15
影响因子：
10.5
通讯作者：
Londono-Vallejo, Arturo
中科院分区：
文献类型：
--
作者：
Arnoult, Nausica;Saintome, Carole;Londono-Vallejo, Arturo
端粒复制的机制仍然不清楚。有人认为，通过滞后机制进行的富含G的端粒链复制以随机的方式需要WRN蛋白。在这里，我们表明，这一要求比以前认为的更系统。我们的数据是兼容的情况下，在没有WRN，DNA合成在复制叉是解耦的，从而使复制继续对C链，而单个G链积累。我们还表明，在细胞中，WRN和POT 1都是有限的，G-和C-丰富的端粒链缩短，这表明一个完整的复制块。在这种特定条件下，跨越两个POT 1-OB（寡核苷酸结合）折叠结构域的片段的表达能够恢复C（但不是G）链复制，表明POT 1与滞后链的结合允许在不存在WRN的情况下DNA合成解偶联。此外，体外实验表明，纯化的POT 1具有更高的亲和力端粒G-丰富的链比纯化的RPA。我们提出了一个模型，其中POT 1与RPA的端粒落后链的相对富集允许或不允许在复制叉的DNA合成的解偶联。我们的研究揭示了hPOT 1在端粒复制过程中的意想不到的作用。
Mechanisms of telomere replication remain poorly defined. It has been suggested that G-rich telomeric strand replication by lagging mechanisms requires, in a stochastic way, the WRN protein. Here we show that this requirement is more systematic than previously thought. Our data are compatible with a situation in which, in the absence of WRN, DNA synthesis at replication forks is uncoupled, thus allowing replication to continue on the C strand, while single G strands accumulate. We also show that in cells in which both WRN and POT1 are limiting, both G-and C-rich telomeric strands shorten, suggesting a complete replication block. Under this particular condition, expression of a fragment spanning the two POT1-OB (oligonucleotide-binding) fold domains is able to restore C (but not G) strand replication, suggesting that binding of POT1 to the lagging strand allows DNA synthesis uncoupling in the absence of WRN. Furthermore, in vitro experiments indicate that purified POT1 has a higher affinity for the telomeric G-rich strand than purified RPA. We propose a model in which the relative enrichments of POT1 versus RPA on the telomeric lagging strand allows or does not allow uncoupling of DNA synthesis at the replication fork. Our study reveals an unanticipated role for hPOT1 during telomere replication.