Supplementary data for: Encyclopaedia of family A DNA polymerases localized in organelles: Evolutionary contribution of bacteria including the proto-mitochondrion

DNA polymerases (DNAPs) synthesize DNA from deoxyribonucleotides in a semi-conservative manner and serve as the core of DNA replication and repair machinery. In eukaryotic cells, there are two genome-containing organelles, mitochondria and plastids, that were derived from an α-proteobacterium and a cyanobacterium, respectively. Except for rare cases of genome-lacking mitochondria and plastids, both organelles must be served by nucleus-encoded DNAPs that localize and work in them to maintain their genomes. The evolution of organellar DNAPs has yet to be fully understood because of two unsettled issues. First, the diversity of organellar DNAPs has not been elucidated in the full spectrum of eukaryotes. Second, it is unclear when the DNAPs that were used originally in the endosymbiotic bacteria giving rise to mitochondria and plastids were discarded, as the organellar DNAPs known to date show no phylogenetic affinity to those of the extant α-proteobacteria or cyanobacteria. In this study, we identified from diverse eukaryotes 134 family A DNAP sequences, which were classified into 10 novel types, and explored their evolutionary origins. The subcellular localizations of selected DNAPs were further examined experimentally. The results presented here suggest that the diversity of organellar DNAPs has been shaped by multiple transfers of the PolI gene from phylogenetically broad bacteria, and their occurrence in eukaryotes was additionally impacted by secondary plastid endosymbioses. Finally, we propose that the last eukaryotic common ancestor may have possessed two mitochondrial DNAPs, POP and a candidate of the direct descendant of the proto-mitochondrial DNAP, rdxPolA, identified in this study.

DNA聚合酶（DNA polymerases, DNAPs）以半保留方式由脱氧核糖核苷酸合成DNA，是DNA复制与修复机制的核心组分。在真核细胞中，存在两类携带基因组的细胞器——线粒体（mitochondria）与质体（plastids），它们分别起源于α-变形菌（α-proteobacterium）和蓝细菌（cyanobacterium）。除极少数缺乏基因组的线粒体和质体案例外，这两类细胞器必须依赖由细胞核编码、并定位至其中发挥功能的DNAP，以维持自身基因组的稳定。细胞器DNAP的演化机制尚未完全阐明，存在两个悬而未决的问题：其一，细胞器DNAP的多样性尚未在全谱系真核生物中得到系统解析；其二，目前已知的细胞器DNAP与现存α-变形菌或蓝细菌的同源酶均无系统发育亲缘关系，因此尚不清楚起源于内共生细菌的原始线粒体与质体DNAP究竟在何时被取代。本研究从多样的真核生物类群中鉴定出134条A家族DNA聚合酶（family A DNAP）序列，将其划分为10个新型类别，并探究了它们的演化起源。研究进一步通过实验手段验证了部分候选DNAP的亚细胞定位。本研究结果表明，细胞器DNAP的多样性是由多起跨宽泛系统发育范围的细菌PolI基因水平转移所塑造的，而真核生物中这类酶的出现还受到次生质体内共生事件的额外影响。最后，本研究提出，最后真核共同祖先可能拥有两种线粒体DNAP：POP，以及本研究中鉴定的原线粒体DNAP（proto-mitochondrial DNAP）的直接同源候选rdxPolA。