The brown algae Pl.LSU/2 group II intron-encoded protein has functional reverse transcriptase and maturase activities

PLoS One. 2013;8(3):e58263. doi: 10.1371/journal.pone.0058263. Epub 2013 Mar 11.

Abstract

Group II introns are self-splicing mobile elements found in prokaryotes and eukaryotic organelles. These introns propagate by homing into precise genomic locations, following assembly of a ribonucleoprotein complex containing the intron-encoded protein (IEP) and the spliced intron RNA. Engineered group II introns are now commonly used tools for targeted genomic modifications in prokaryotes but not in eukaryotes. We speculate that the catalytic activation of currently known group II introns is limited in eukaryotic cells. The brown algae Pylaiella littoralis Pl.LSU/2 group II intron is uniquely capable of in vitro ribozyme activity at physiological level of magnesium but this intron remains poorly characterized. We purified and characterized recombinant Pl.LSU/2 IEP. Unlike most IEPs, Pl.LSU/2 IEP displayed a reverse transcriptase activity without intronic RNA. The Pl.LSU/2 intron could be engineered to splice accurately in Saccharomyces cerevisiae and splicing efficiency was increased by the maturase activity of the IEP. However, spliced transcripts were not expressed. Furthermore, intron splicing was not detected in human cells. While further tool development is needed, these data provide the first functional characterization of the PI.LSU/2 IEP and the first evidence that the Pl.LSU/2 group II intron splicing occurs in vivo in eukaryotes in an IEP-dependent manner.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Line
  • Endoribonucleases / metabolism
  • Gene Expression
  • Gene Order
  • Humans
  • Introns*
  • Nucleic Acid Conformation
  • Nucleotidyltransferases / metabolism
  • Phaeophyta / genetics*
  • Phaeophyta / metabolism*
  • Proteins / genetics*
  • Proteins / metabolism*
  • RNA / chemistry
  • RNA / genetics
  • RNA / metabolism
  • RNA Splicing
  • RNA-Directed DNA Polymerase / metabolism
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / metabolism
  • Vault Ribonucleoprotein Particles / metabolism

Substances

  • Proteins
  • Vault Ribonucleoprotein Particles
  • RNA
  • Nucleotidyltransferases
  • mRNA maturase
  • RNA-Directed DNA Polymerase
  • Endoribonucleases

Grant support

This work was supported by the Association Française contre les myopathies (AFM, http://www.afm-telethon.fr/); the Institut national de la santé et de la recherche médicale (INSERM http://www.inserm.fr); the University of Evry Val d’Essonne (www.univ-evry.fr); and by the 7th European Commission Framework Program (FP7) “PERSIST” (agreement 222878). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.