Lee S T, Tarn C, Chang K P
Laboratory of Molecular Parasitology, Academia Sinica, Taipei, Taiwan.
Mol Biochem Parasitol. 1993 Apr;58(2):187-203. doi: 10.1016/0166-6851(93)90041-u.
To characterize the differences between kDNA minicircles of drug-resistant Leishmania mexicana amazonensis variants that show nuclear DNA amplification and minicircles of variants without nuclear DNA amplification, we sequenced minicircles from repeatedly cloned parasites. The dominant minicircles from arsenite- and tunicamycin-resistant parasites with DNA amplification were found to preexist as minor conserved divergent classes in parental wild-type cells. These classes shared very limited similarity with the predominant wild-type minicircle sequences or sequences from drug resistant parasites without amplification. These minor classes were preferentially selected to replicate in variants with DNA amplification and subsequently became the dominant sequences in these variants. Kinetic studies of the correlation between amplification and deamplification of the nuclear DNA and the switch in kDNA minicircle dominance indicated that factor(s) other than the amplified chromosomal DNA itself caused the minicircles to switch. Treating the kDNA networks isolated from cells at the switch transition period with single cutter endonucleases specific for either wild-type or variant-specific minicircles resulted in structural modifications consistent with both minicircle sequence classes being present simultaneously in the same network. This establishes the 'trans' nature of the switch.
为了表征显示核DNA扩增的抗药性亚马逊利什曼原虫变种的线粒体DNA小环与无核DNA扩增的变种的小环之间的差异，我们对来自反复克隆的寄生虫的小环进行了测序。发现来自具有DNA扩增的抗亚砷酸盐和抗衣霉素寄生虫的主要小环在亲本野生型细胞中以次要的保守分歧类预先存在。这些类别与主要的野生型小环序列或来自无扩增的抗药寄生虫的序列具有非常有限的相似性。这些次要类别在具有DNA扩增的变种中被优先选择进行复制，随后成为这些变种中的主要序列。对核DNA扩增与去扩增之间的相关性以及线粒体DNA小环优势转换的动力学研究表明，除了扩增的染色体DNA本身之外的其他因素导致了小环的转换。用对野生型或变种特异性小环特异的单切割内切核酸酶处理在转换过渡期从细胞中分离的线粒体DNA网络，导致结构修饰，这与同一网络中同时存在两种小环序列类别一致。这确立了转换的“反式”性质。
