Incongruent end structures of leading and lagging telomeres dictate the nature of end replication problem

The end replication problem refers to the incomplete replication of parental DNA at telomeres, a process whose molecular depiction is hampered by the complex nature of telomere ends. Here we recapitulate this process using a synthetic de novo telomere in yeast and delineate distinct molecular fates of telomere ends in vivo. We show that the lagging strand telomere carries a ~10 nt 3’ overhang, while the leading strand telomere has a Yku-protected blunt end, which is prevalent on native telomeres. Moreover, we find that RNase H2 is mainly responsible for the removal of the last RNA primer. In the absence of RNase H2 activity, RNA primer retention on the lagging strand telomere attenuates telomere erosion and delays senescence in telomerase-null cells. These findings highlight incongruent end structures on telomeres and clarify that the primary culprit behind end replication problem is the incompletely replicated lagging strand telomere.末端复制问题（end replication problem）指亲本DNA在端粒（telomeres）处无法完成复制的现象，该过程的分子机制阐释因端粒末端复杂的结构特性而受阻。本研究借助酵母中的从头合成人工端粒（synthetic de novo telomere）重现了这一过程，并阐明了体内端粒末端的独特分子结局。研究发现，滞后链端粒（lagging strand telomere）带有一段约10 nt的3'端突出（3’ overhang），而前导链端粒（leading strand telomere）则存在受Yku蛋白保护的平末端，该结构在天然端粒中广泛存在。此外，本研究证实核糖核酸酶H2（RNase H2）主要负责移除最后一段RNA引物（RNA primer）。在缺失核糖核酸酶H2活性的情况下，滞后链端粒上存留的RNA引物会减弱端粒侵蚀（telomere erosion）进程，并延缓端粒酶缺陷细胞（telomerase-null cells）的细胞衰老。本研究结果揭示了端粒末端结构的异质性，并明确了末端复制问题的主要根源正是复制不完全的滞后链端粒。