Kinetoplast maxicircle DNA replication in Crithidia fasciculata and Trypanosoma brucei

Laura Rocco Carpenter, Paul T. Englund

Research output: Contribution to journalArticlepeer-review

Abstract

Kinetoplast DNA, the mitochondrial DNA of trypanosomatids, is composed of several thousand minicircles and a few dozen maxicircles, all of which are topologically interlocked in a giant network. We have studied the replication of maxicircle DNA, using electron microscopy to analyze replication intermediates from both Crithidia fasciculata and Trypanosoma brucei. Replication intermediates were stabilized against branch migration by introducing DNA interstrand cross-links in vivo with 4,5',8- trimethylpsoralen and UV radiation. Electron microscopy of individual maxicircles resulting from a topoisomerase II decatenation of kinetoplast DNA networks revealed intact maxicircle θ structures. Analysis of maxicircle DNA linearized by restriction enzyme cleavage revealed branched replication intermediates derived from θ structures. Measurements of the linearized branched molecules in both parasites indicate that replication initiates in the variable region (a noncoding segment characterized by repetitive sequences) and proceeds unidirectionally, clockwise on the standard map.

Original languageEnglish (US)
Pages (from-to)6794-6803
Number of pages10
JournalMolecular and cellular biology
Volume15
Issue number12
DOIs
StatePublished - Dec 1995

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

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