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Alternative mechanisms of ribosome stalling rescue in the gram‐negative bacterium Francisella tularensis
Author(s) -
Srinivas Pooja,
Goralski Tyler D. P.,
Keiler Kenneth C.,
Dunham Christine M.
Publication year - 2019
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2019.33.1_supplement.628.3
Subject(s) - ribosome , biology , translation (biology) , francisella tularensis , release factor , protein biosynthesis , stop codon , translational frameshift , start codon , microbiology and biotechnology , genetics , messenger rna , gene , rna , virulence
The number of new antibiotics being developed has plateaued, particularly compared to the sharp increase in antibiotic‐resistant bacteria plaguing humanity. Many current antibiotics target the bacterial ribosome, but have cross‐reactivity with eukaryotic ribosomes, causing host toxicity. One approach to prevent this is the identification and development of antibiotics that target bacterial‐specific pathways, such as ribosome stalling rescue. At any given moment, 5–10% of E. coli ribosomes are stalled. Stalled ribosome complexes arise when the ribosome halts during translation, which may arise due to the absence of an in‐frame stop codon, depletion of aminoacyl‐tRNAs, or proline‐rich motifs. Trans ‐translation, found in 99.9% of bacterial species, is the main rescue pathway for stalled complexes that arise due to no in‐frame stop codon. Most bacteria remain viable without trans‐ translation because they encode additional rescue systems, such as Alternative Rescue Factor A and B (ArfA and ArfB). The pathogenic bacterium F. tularensis appears to only encode trans‐ translation factors, but trans‐ translation deletion strains remain viable. How does F. tularensis survive without known ribosome rescue mechanisms present? Transposon mutagenesis data from the Keiler lab identifies a novel release factor, ArfT, which allows F. tularensis survival in the absence of trans‐ translation. In vitro translation assays find that ArfT facilitates peptidyl‐tRNA hydrolysis with both RF1 and RF2, in contrast to the model of ArfA‐RF2‐mediated rescue in other well‐studied systems. These data suggest that ArfT‐mediated ribosome rescue occurs via a novel mechanism. Our results indicate that F. tularensis 70S‐ArfT stalled complexes have an increase of RF1 binding, suggesting a novel mechanism of recognition by RF1. Future studies include structural and biochemical characterization of the molecular mechanism of ArfT and release factor binding to stalled F. tularensis 70S ribosome complexes. These studies will provide new insights on the fundamental mechanism and role of ribosome rescue in bacteria. Support or Funding Information T32‐GM008602 R01‐GM093278‐09S1 R01‐GM121650‐02 This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .