Trypanosomes Have Six Mitochondrial DNA Helicases With One Controlling Kinetoplast Maxicircle Replication

Mol Cell. 2009 Aug 28;35(4):490-501. doi: 10.1016/j.molcel.2009.07.004. Epub 2009 Jul 30.

Abstract

Kinetoplast DNA (kDNA), the trypanosome mitochondrial DNA, contains thousands of minicircles and dozens of maxicircles interlocked in a giant network. Remarkably, Trypanosoma brucei's genome encodes 8 PIF1-like helicases, 6 of which are mitochondrial. We now show that TbPIF2 is essential for maxicircle replication. Maxicircle abundance is controlled by TbPIF2 level, as RNAi of this helicase caused maxicircle loss, and its overexpression caused a 3- to 6-fold increase in maxicircle abundance. This regulation of maxicircle level is mediated by the TbHslVU protease. Previous experiments demonstrated that RNAi knockdown of TbHslVU dramatically increased abundance of minicircles and maxicircles, presumably because a positive regulator of their synthesis escaped proteolysis and allowed synthesis to continue. Here, we found that TbPIF2 level increases following RNAi of the protease. Therefore, this helicase is a TbHslVU substrate and an example of a positive regulator, thus providing a molecular mechanism for controlling maxicircle replication.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • DNA Helicases / genetics
  • DNA Helicases / metabolism*
  • DNA Replication*
  • DNA, Kinetoplast / biosynthesis*
  • DNA, Mitochondrial / biosynthesis*
  • DNA, Protozoan / biosynthesis*
  • Gene Expression Regulation
  • Mutation
  • Peptide Hydrolases / metabolism
  • Protozoan Proteins / genetics
  • Protozoan Proteins / metabolism*
  • RNA Interference
  • Time Factors
  • Transfection
  • Trypanosoma brucei brucei / enzymology
  • Trypanosoma brucei brucei / genetics*
  • Trypanosoma brucei brucei / growth & development

Substances

  • DNA, Kinetoplast
  • DNA, Mitochondrial
  • DNA, Protozoan
  • Protozoan Proteins
  • Peptide Hydrolases
  • DNA Helicases