RNA interference of a trypanosome topoisomerase II causes progressive loss of mitochondrial DNA.
We studied the function of a Trypanosoma brucei topoisomerase II using RNA interference (RNAi). Expression of a topoisomerase II double-stranded RNA as a stem-loop caused specific degradation of mRNA followed by loss of protein. After 6 days of RNAi, the parasites' growth rate declined and the cells subsequently died. The most striking phenotype upon induction of RNAi was the loss of kinetoplast DNA (kDNA), the cell's catenated mitochondrial DNA network. The loss of kDNA was preceded by gradual shrinkage of the network and accumulation of gapped free minicircle replication intermediates. These facts, together with the localization of the enzyme in two antipodal sites flanking the kDNA, show that a function of this topoisomerase II is to attach free minicircles to the network periphery following their replication.
Wang
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Department of Biological Chemistry, Johns Hopkins Medical School, 725 N. Wolfe Street, Baltimore, MD 21205, USA.
Englund
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PT
D000818:Animals; D004261:DNA Replication; D004250:DNA Topoisomerases, Type II / Q000235:genetics / Q000378:metabolism*; D018105:DNA, Kinetoplast / Q000235:genetics* / Q000378:metabolism / Q000648:ultrastructure; D004272:DNA, Mitochondrial / Q000235:genetics* / Q000378:metabolism / Q000648:ultrastructure; D016054:DNA, Protozoan / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D007700:Kinetics; D010641:Phenotype; D012330:RNA, Double-Stranded / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016053:RNA, Protozoan / Q000235:genetics*; D013997:Time Factors; D014346:Trypanosoma brucei brucei / Q000201:enzymology* / Q000235:genetics* / Q000254:growth & development
D018105:DNA, Kinetoplast; D004272:DNA, Mitochondrial; D016054:DNA, Protozoan; D012330:RNA, Double-Stranded; D012333:RNA, Messenger; D016053:RNA, Protozoan; D004250:DNA Topoisomerases, Type II