| 其他摘要 | Several groups of protists (such as Microsporidia, Giardia, Trichomonas and, Entamoeba, and so on) have attracted long attention not only in medicine for their parasitical nosogenesis, but also in evolutionary biology for their unique evolutionary-status. It was once thought that they lacked mitochondria (amitochondriate), in addition to that some molecular phylogenetic analyses showed that they branched at the base of eukaryotes, thus not a few people considered that they were the most primitive eukaryotes which diverged before the endosymbiotic origin of mitochondria, and that they were the relicts of transformation between prokaryotes and eukaryotes. A hypothesis was formalized by calling them 'Archezoa' to denote the primitively amitochondriate condition. It was thought that they were very important for the study of the origin and evolution of eukaryotic cells. However, recently, a growing body of evidence challenged their evolutionary status. In the present paper, firstly, the genes encoding type II DNA topoisomerases were identified from Giardia lamblia, Trichomonas vaginalis and Entamoeba histolytica by PCR-amplifying, sequencing and database searching. The results of RT-PCR and sequence analysis showed that there exist no introns in all the genes. Sequence blast indicated that their sequences showed strong homology to known type II DNA topoisomerases, especially to eukaryotic DNA topoisomerase II. Using bioinformational methods, we have analyzed the characters of their encoded enzymes. Our analysis showed that G lamblia topoisomerase II was different from that of its human host, for example, six insertions in the ATPase domain and the central domain; a ~100aa longer central domain; a ~200aa shorter C-terminal domain containing rich charged residues. These detail features might be helpful to exploit drug selectivity for antigiardial therapy whose target enzyme is DNA topoisomerase II. Secondly, besides the above obtained sequences, the sequences of Encephalitozoon cuniculi and a series of eukaryotes of different evolutionary positions were obtained from GenBank, and then phylogenetic trees were reconstructed by various methods to study the evolutionary status of these amitochondrate protists. Simultaneously, the affection of "Long Branch Attraction (LBA)" to the phyologenetic trees was also investigated. Our results showed that owing to the unique characters of DNA topoisomerase II, LBA had little influence on the trees based on the enzyme. Our phylogenetic trees not only reflected the widely accepted phylogenetic relationships among the main eukaeyotic groups, but also showed that the evolutionary positions of those amitochondriate protists were different from those in previous reported trees: they were not the earliest branching eukaryotes, but emerged later than mitochondriate protozoa (e.g. Kinetoplastid and Mycetozoa); they belonged to different groups of eukaryotes with different evolutionary positions. In addition to the other recent evidence such as the discovery of mitochondria-like organelle, our results suggest that all these amitochondriate protists are not extremely primitive as was considered before, and they might be different groups of eukaryotes and diverged after the endosymbiotic origin of mitochondria, though some of them, such as GiardiUj are primitive eukaryotes and are of significance to the study of early evolution of eukaryotes. |
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