Gehring Alexandra M, Astling David P, Matsumi Rie, Burkhart Brett W, Kelman Zvi, Reeve John N, Jones Kenneth L, Santangelo Thomas J
Department of Biochemistry and Molecular Biology, Colorado State University, Fort Collins, CO, United States.
Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Aurora, CO, United States.
Front Microbiol. 2017 Oct 27;8:2084. doi: 10.3389/fmicb.2017.02084. eCollection 2017.
The initiation of DNA replication is typically tightly regulated by proteins that form initiation complexes at specific sequences known as replication origins. In Archaea and Eukaryotes, Cdc6, a near-universally conserved protein binds and facilitates the origin-dependent assembly of the replicative apparatus. TK1901 encodes Cdc6 in but, as we report here, TK1901 and the presumed origin of replication can be deleted from the genome of this hyperthermophilic Archaeon without any detectable effects on growth, genetic competence or the ability to support autonomous plasmid replication. All regions of the genome were equally represented in the sequences generated by whole genome sequencing of DNA isolated from strains with or without TK1901, inconsistent with DNA initiation occurring at one or few origins, and instead suggestive of replication initiating at many sites distributed throughout the genome. We were unable to generate strains lacking the recombination factors, RadA or RadB, consistent with cells, that are oligoploid (7-19 genomes per cell), employing a recombination-based mechanism of DNA replication. Deletion of the previously presumed origin region reduced the long-term viability of cultures supporting the possibility that retaining an origin-based mechanism of DNA initiation provides a survival mechanism for stationary phase cells with only one genome.
DNA复制的起始通常受到蛋白质的严格调控，这些蛋白质在特定序列（称为复制起点）处形成起始复合物。在古菌和真核生物中，Cdc6是一种几乎普遍保守的蛋白质，它结合并促进复制装置依赖于起点的组装。TK1901在[具体物种名称未给出]中编码Cdc6，但正如我们在此报告的那样，TK1901和假定的复制起点可以从古嗜热古菌的基因组中删除，而对生长、遗传感受态或支持自主质粒复制的能力没有任何可检测到的影响。从有或没有TK1901的菌株中分离的DNA进行全基因组测序所产生的序列中，基因组的所有区域都有同等的代表性，这与DNA起始发生在一个或几个起点不一致，而是提示复制在整个基因组中分布的许多位点起始。我们无法产生缺乏重组因子RadA或RadB（与[具体物种名称未给出]细胞一致，这些细胞是寡倍体（每个细胞有7 - 19个基因组），采用基于重组的DNA复制机制）的菌株。删除先前假定的起点区域降低了培养物的长期活力，这支持了保留基于起点的DNA起始机制为只有一个基因组的静止期细胞提供生存机制的可能性。
