Abstract
Trypanosomes have an unusual mitochondrial genome, called kinetoplast DNA, that is a giant network containing thousands of interlocked minicircles. During kinetoplast DNA synthesis, minicircles are released from the network for replication as theta-structures, and then the free minicircle progeny reattach to the network. We report that a mitochondrial protein, which we term p38, functions in kinetoplast DNA replication. RNA interference (RNAi) of p38 resulted in loss of kinetoplast DNA and accumulation of a novel free minicircle species named fraction S. Fraction S minicircles are so underwound that on isolation they become highly negatively supertwisted and develop a region of Z-DNA. p38 binds to minicircle sequences within the replication origin. We conclude that cells with RNAi-induced loss of p38 cannot initiate minicircle replication, although they can extensively unwind free minicircles.
Publication types
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Research Support, N.I.H., Extramural
MeSH terms
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Animals
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Base Sequence
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DNA Replication / physiology*
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DNA, Kinetoplast / biosynthesis*
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DNA, Kinetoplast / genetics
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DNA, Kinetoplast / ultrastructure
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DNA-Binding Proteins / antagonists & inhibitors
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / isolation & purification
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DNA-Binding Proteins / metabolism*
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Genes, Protozoan
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Microscopy, Electron
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Mitochondrial Proteins / antagonists & inhibitors
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Mitochondrial Proteins / genetics
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Mitochondrial Proteins / isolation & purification
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Mitochondrial Proteins / metabolism*
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Models, Biological
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Proteomics
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Protozoan Proteins / antagonists & inhibitors
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Protozoan Proteins / genetics
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Protozoan Proteins / isolation & purification
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Protozoan Proteins / metabolism*
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RNA Interference
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Trypanosoma brucei brucei / genetics
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Trypanosoma brucei brucei / metabolism*
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Trypanosoma brucei brucei / ultrastructure
Substances
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DNA, Kinetoplast
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DNA-Binding Proteins
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Mitochondrial Proteins
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Protozoan Proteins