Translation initiation factor IF1 is essential for cell viability in Escherichia coli

Abstract

Translation initiation factor IF1 is a highbly conserved element of the prokaryotic translational apparatus. It has been demonstrated earlier that the factor stimulates in vitro the initiation phase of protein synthesis. However, no mutation in its gene, infA, has been identified, and a role for IF1 in translation has not been demonstrated in vivo. To elucidate the function of IFi and determine if the protein is essential for cell growth, the chromosomal copy of infA was disrupted. Cell viability is maintained only when infA is expressed in trans from a plasmid, thereby demonstrating that WI1 is essential for cell growth in Escherichia coli. Cells depleted of IF1 exhibit few polsomes, suggesting that IFI functions in the initiation phase of protein synthesis.Initiation of protein synthesis in prokaryotes involves formation of a 30S preinitiation complex in which fMet-tRNA interacts with the initiation codon of an mRNA bound to the surface of the 30S ribosomal subunit. The efficiency and fidelity of formation of the 30S preinitiation complex is promoted by three initiation factors; IF1, IF2, and IF3. The properties and mechanism of action of the initiation factors have been reviewed recently (13). All three initiation factors bind to the 30S ribosomal subunit in the absence of other translational components. IF2 binds GTP and fMet-tRNAf, thereby ensuring that the tRNA bound to the 30S subunit is the initiator tRNA, fMet-tRNAf. 1F3 plays at least two roles: it acts as an antiassociation factor to maintain the ribosomal subunits in a dissociated state, and it promotes the selection of the initiator fMet-tRNA by monitoring the tRNA anticodon stem-loop region and the correctness of the anticodon-initiator codon interaction on the ribosome. The function of IF1 is less clear, since no specific role has yet been assigned to this protein.IF1 is the smallest of the initiation factors (Mr,8,100) and consists of 71 amino acid residues. The gene for IF1, infA, has been cloned, sequenced, and mapped to about 20 min on the Escherichia coli chromosome (25). The infA gene is transcribed by two promoters to yield two sizes of monocistronic mRNAs both ending at the same terminator (8). Recently, homologous genes have been identified in Bacillus subtilis and in the chloroplasts of several plants (4,23,28). The high degree of homology indicates that IF1 is a conserved protein and suggests that it plays an important role in the cell. However, our knowledge of the function of IF1 comes exclusively from a number of in vitro assays for translation. IF1 enhances the rates of 70S ribosome dissociation and subunit association but does not change the equilibrium position (10). It alone binds poorly to 30S ribosomal subunits, but when added with IF2 both factor-binding affinities are increased (32). IF1 stimulates IF2-dependent fMet-tRNAf binding to 30S or 70S ribosomes in the presence of mRNA. However, the best demonstration that IF1 truly functions in protein synthesis comes from the observation that ,-galactosidase synthesis in a DNA-l...

σ S -Dependent Gene Expression at the Onset of Stationary Phase in Escherichia coli : Function of σ S -Dependent Genes and Identification of Their Promoter Sequences

“…It is noteworthy that the newly identified rpoS-dependent genes do not appear to be essential genes as defined by Gerdes et al (26), except for infA (18), a finding consistent with the lack of major effects of the rpoS mutation on cell viability. To confirm the results of the whole-genome expression analysis, we performed RT-PCR experiments on a selection of differentially expressed genes belonging to each functional category (Fig.…”

Initiation of Protein Synthesis in Bacteria

“…Ejection of IF1 consequently opens the A-site for incoming aminoacyl-tRNAs. In vivo studies have shown that IF1-depleted cells have low growth rates and short polysomes (39). These data demonstrate that IF1 is essential for cell viability and suggest that one or more of its functions are crucial.…”

Determination of the Core of a Minimal Bacterial Gene Set

“…infA, infB, and infC are required for translation initiation and are also essential in E. coli (17,25) and S. aureus (22). The status of the infC gene in B. aphidicola BSg, initially annotated as a pseudogene due to the absence of a translation start codon (80), was recently changed (28) because, as in other enteric bacteria, it possesses an unusual AUU initiator codon.…”