In vivo involvement of mutated initiation factor IF1 in gene expression control at the translational level

Abstract

Edited by Lev KisselevAbstract The influence in vivo of mutated forms of translation initiation factor (IF1) on the expression of the lacZ or 3A 0 reporter genes, with different initiation and/or +2 codons, has been investigated. Reporter gene expression in these infA(IF1) mutants is similar to the wild-type strain. The results do not support the longstanding hypothesis that IF1 could perform discriminatory functions while blocking the aminoacyl-tRNA acceptor site (A-site) of the ribosome. One cold-sensitive IF1 mutant shows a general overexpression, in particular at low temperatures, of both reporter genes at the protein but not mRNA level.

Translation initiation region dependency of translation initiation in Escherichia coli by IF1 and kasugamycin

“…IF1 is essential for cell viability, although the reason for this is obscure. A set of mutants with mutations in infA , giving an altered IF1, has been isolated [38]. Some of these mutants are cold sensitive for growth, and have increased gene expression in an in vivo reporter system.…”

“…The influence of different sequences downstream of AUG on gene expression at the translational level has been well characterized [41]. Even though IF1 binds to the A‐site of the 30S subunit, different +2 codons do not cause significantly changed levels of protein expression in different IF1 mutant strains [38], indicating a lack of codon specificity. Using CVR40D, we used the 3A′/2A′ test system to analyze other plasmids with different sequences downstream of the initiation codon in 3A′ (downstream regions DR‐A, DR‐B, DR‐C, and DR‐D) (Fig.…”

“…However, it was found by Croitoru et al. [38] that cold‐sensitive chromosomal infA mutants with the mutations R40D and R69L had increased reporter gene expression in vivo as compared with the wild‐type strain. This depends on the TIR in the mRNA [38].…”

“…[38] that cold‐sensitive chromosomal infA mutants with the mutations R40D and R69L had increased reporter gene expression in vivo as compared with the wild‐type strain. This depends on the TIR in the mRNA [38]. The overexpression found in the mutant as compared with the wild‐type strain, using two different reporter gene systems, depends on the composition of the TIR.…”

Translation initiation factor IF1 of Bacillus stearothermophilus and Thermus thermophilus substitute for Escherichia coli IF1 in vivo and in vitro without a direct IF1–IF2 interaction

“…Translation initiation factor IF1 is a small (8.1 kDa) RNA‐binding protein consisting of five β‐strands and a loop connecting strands 3 and 4 containing a short and flexible α‐helix (Sette et al ., 1997). IF1 binds to the A‐site of the 30S ribosomal subunit, in the cleft between protein S12, helix 44 and the 530 loop of 16S rRNA (Moazed et al ., 1995; Carter et al ., 2001); the amino acid residues involved in/affected by the binding have been identified by site‐directed mutagenesis, NMR spectroscopy and crystallography (Gualerzi et al ., 1989; Sette et al ., 1997; Carter et al ., 2001; Croitoru et al ., 2005).…”