Stem-loop silencing reveals that a third mitochondrial DNA polymerase, POLID, is required for kinetoplast DNA replication in trypanosomes

Eukaryot Cell. 2008 Dec;7(12):2141-6. doi: 10.1128/EC.00199-08. Epub 2008 Oct 10.


Kinetoplast DNA (kDNA), the mitochondrial genome of trypanosomes, is a catenated network containing thousands of minicircles and tens of maxicircles. The topological complexity dictates some unusual features including a topoisomerase-mediated release-and-reattachment mechanism for minicircle replication and at least six mitochondrial DNA polymerases (Pols) for kDNA transactions. Previously, we identified four family A DNA Pols from Trypanosoma brucei with similarity to bacterial DNA Pol I and demonstrated that two (POLIB and POLIC) were essential for maintaining the kDNA network, while POLIA was not. Here, we used RNA interference to investigate the function of POLID in procyclic T. brucei. Stem-loop silencing of POLID resulted in growth arrest and the progressive loss of the kDNA network. Additional defects in kDNA replication included a rapid decline in minicircle and maxicircle abundance and a transient accumulation of minicircle replication intermediates before loss of the kDNA network. These results demonstrate that POLID is a third essential DNA Pol required for kDNA replication. While other eukaryotes utilize a single DNA Pol (Pol gamma) for replication of mitochondrial DNA, T. brucei requires at least three to maintain the complex kDNA network.

Publication types

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

MeSH terms

  • Animals
  • DNA Replication*
  • DNA, Kinetoplast / genetics*
  • DNA-Directed DNA Polymerase / genetics
  • DNA-Directed DNA Polymerase / metabolism*
  • Mitochondria / genetics
  • Mitochondria / metabolism
  • Mitochondrial Proteins / genetics
  • Mitochondrial Proteins / metabolism
  • Protozoan Proteins / genetics
  • Protozoan Proteins / metabolism*
  • RNA Interference*
  • RNA, Double-Stranded / genetics
  • Trypanosoma brucei brucei / enzymology*
  • Trypanosoma brucei brucei / genetics


  • DNA, Kinetoplast
  • Mitochondrial Proteins
  • Protozoan Proteins
  • RNA, Double-Stranded
  • DNA-Directed DNA Polymerase