Autoselection of cytoplasmic yeast virus like elements encoding toxin/antitoxin systems involves a nuclear barrier for immunity gene expression

PLoS Genet. 2015 May 14;11(5):e1005005. doi: 10.1371/journal.pgen.1005005. eCollection 2015 May.

Abstract

Cytoplasmic virus like elements (VLEs) from Kluyveromyces lactis (Kl), Pichia acaciae (Pa) and Debaryomyces robertsiae (Dr) are extremely A/T-rich (>75%) and encode toxic anticodon nucleases (ACNases) along with specific immunity proteins. Here we show that nuclear, not cytoplasmic expression of either immunity gene (PaORF4, KlORF3 or DrORF5) results in transcript fragmentation and is insufficient to establish immunity to the cognate ACNase. Since rapid amplification of 3' ends (RACE) as well as linker ligation of immunity transcripts expressed in the nucleus revealed polyadenylation to occur along with fragmentation, ORF-internal poly(A) site cleavage due to the high A/T content is likely to prevent functional expression of the immunity genes. Consistently, lowering the A/T content of PaORF4 to 55% and KlORF3 to 46% by gene synthesis entirely prevented transcript cleavage and permitted functional nuclear expression leading to full immunity against the respective ACNase toxin. Consistent with a specific adaptation of the immunity proteins to the cognate ACNases, cross-immunity to non-cognate ACNases is neither conferred by PaOrf4 nor KlOrf3. Thus, the high A/T content of cytoplasmic VLEs minimizes the potential of functional nuclear recruitment of VLE encoded genes, in particular those involved in autoselection of the VLEs via a toxin/antitoxin principle.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Cloning, Molecular
  • Cytoplasm / metabolism*
  • Escherichia coli / genetics
  • Gene Expression Regulation, Fungal
  • Killer Factors, Yeast / genetics
  • Killer Factors, Yeast / metabolism*
  • Kluyveromyces / genetics
  • Kluyveromyces / metabolism*
  • Molecular Sequence Data
  • Pichia / genetics
  • Pichia / metabolism*
  • Plasmids
  • RNA, Fungal / genetics
  • Ribonucleases / genetics*
  • Ribonucleases / metabolism
  • Saccharomycetales / genetics
  • Saccharomycetales / metabolism*

Substances

  • Killer Factors, Yeast
  • RNA, Fungal
  • Ribonucleases
  • anticodon nuclease

Grants and funding

We acknowledge support by Deutsche Forschungsgemeinschaft and Open Access Publication Fund of University of Muenster. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.