古菌拥有与真核生物类似、而与细菌截然不同的DNA复制机器。古菌与真核生物的引发酶同属于古菌-真核生物引发酶（archaeo-eukaryotic primase, AEP）超家族，而细菌引发酶则属于DnaG超家族。此前，古菌AEP被认为是仅含催化亚基PriS和非催化亚基PriL的异二聚体PriSL。本实验室近期发现，部分古菌编码一种新的引发酶非催化亚基PriX或其同源蛋白；此外，还发现极端嗜热古菌—硫化叶菌PriSL具有依赖模板的跨模板合成能力，因而有可能参与DNA双链断裂的非同源末端连接修复。本实验室根据已有结果推测，最后的共同祖先拥有AEP和DnaG双引发酶系统。本项目拟以硫化叶菌为模式，阐明PriX及其同源蛋白的结构与生化性质多样性，解析古菌AEP在DNA复制和DNA双链断裂修复过程中的作用及其机制，揭示古菌AEP的功能演化规律。本项目的实施有助于破解AEP多种生理功能的演化奥秘。

Archaea possess a DNA replication apparatus similar to that of Eukarya but distinctly different from that of Bacteria. DNA primases from Archaea and Eukarya belong to the archaeo-eukaryotic primase (AEP) superfamily, while bacterial primases are members of the DnaG superfamily. Until recently, an archaeal AEP was considered to be a heterodimer (PriSL) of a catalytic subunit (PriS) and a noncatalytic subunit (PriL). Our laboratory has recently shown that a number of archaea encode PriX, a novel noncatalytic primase subunit, or its homologue. We have also found that PriSL from the hyperthermophilic archaeon Sulfolobus solfataricus is capable of template-dependent polymerization across discontinuous templates. This observation has led to the suggestion that the enzyme participates in the nonhomologous end-joining (NHEJ) repair of DNA double-strand breaks (DSBs). We have further speculated that the last universal common ancestor (LUCA) possessed both an AEP and a DnaG. This project will assess, using Sulfolobus as a model, the structural and biochemical diversity of PriX and its homologues, elucidate the roles of archaeal AEPs in DNA replication and DSB repair, and shed light on the functional evolution of this protein superfamily. The implementation of the project will help solve the puzzle of the evolution of multiple functions of AEPs.