The two eIF4A helicases in Trypanosoma brucei are functionally distinct

Nucleic Acids Res. 2006 May 10;34(9):2495-507. doi: 10.1093/nar/gkl290. Print 2006.

Abstract

Protozoan parasites belonging to the family Trypanosomatidae are characterized by an unusual pathway for the production of mRNAs via polycistronic transcription and trans-splicing of a 5' capped mini-exon which is linked to the 3' cleavage and polyadenylation of the upstream transcript. However, little is known of the mechanism of protein synthesis in these organisms, despite their importance as agents of a number of human diseases. Here we have investigated the role of two Trypanosoma brucei homologues of the translation initiation factor eIF4A (in the light of subsequent experiments these were named as TbEIF4AI and TbEIF4AIII). eIF4A, a DEAD-box RNA helicase, is a subunit of the translation initiation complex eIF4F which binds to the cap structure of eukaryotic mRNA and recruits the small ribosomal subunit. TbEIF4AI is a very abundant predominantly cytoplasmic protein (over 1 x 10(5) molecules/cell) and depletion to approximately 10% of normal levels through RNA interference dramatically reduces protein synthesis one cell cycle following double-stranded RNA induction and stops cell proliferation. In contrast, TbEIF4AIII is a nuclear, moderately expressed protein (approximately 1-2 x 10(4) molecules/cell), and its depletion stops cellular proliferation after approximately four cell cycles. Ectopic expression of a dominant negative mutant of TbEIF4AI, but not of TbEIF4AIII, induced a slow growth phenotype in transfected cells. Overall, our results suggest that only TbEIF4AI is involved in protein synthesis while the properties and sequence of TbEIF4AIII indicate that it may be the orthologue of eIF4AIII, a component of the exon junction complex in mammalian cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Amino Acid Substitution
  • Animals
  • Eukaryotic Initiation Factor-4A / analysis
  • Eukaryotic Initiation Factor-4A / genetics
  • Eukaryotic Initiation Factor-4A / physiology*
  • Isoenzymes / chemistry
  • Isoenzymes / genetics
  • Models, Molecular
  • Molecular Sequence Data
  • Mutation
  • Protozoan Proteins / analysis
  • Protozoan Proteins / genetics
  • Protozoan Proteins / physiology*
  • RNA Interference
  • RNA, Messenger / metabolism
  • Sequence Homology, Amino Acid
  • Trypanosoma brucei brucei / enzymology*
  • Trypanosoma brucei brucei / genetics
  • Trypanosoma brucei brucei / growth & development

Substances

  • Isoenzymes
  • Protozoan Proteins
  • RNA, Messenger
  • Eukaryotic Initiation Factor-4A