Suppression of defective ribosome assembly in a rbfA deletion mutant by overexpression of Era, an essential GTPase in Escherichia coli.
Era is a small GTP-binding protein and essential for cell growth in Escherichia coli. It consists of two domains: N-terminal GTP-binding and C-terminal RNA-binding KH domains. It has been shown to bind to 16S rRNAs and 30S ribosomal subunits in vitro. Here, we report that a precursor of 16S rRNA accumulates in Era-depleted cells. The accumulation of the precursors is also seen in a cold-sensitive mutant, E200K, in which the mutation site is located in the C-terminal domain. The major precursor molecule accumulated seems to be 17S rRNA, containing extra sequences at both 5' and 3' ends of 16S rRNA. Moreover, the amounts of both 30S and 50S ribosomal subunits relative to the amount of 70S monosomes increase in Era-depleted and E200K mutant cells. The C-terminal KH domain has a high structural similarity to the RbfA protein, a cold shock protein that also specifically associates with 30S ribosomal subunits. RbfA is essential for cell growth at low temperature, and a precursor of 16S rRNA accumulates in an rbfA deletion strain. The 16S rRNA precursor seems to be identical in size to that accumulated in Era mutant cells. Surprisingly, the cold-sensitive cell growth of the rbfA deletion cells was partially suppressed by overproduction of the wild-type Era. The C-terminal domain alone was not able to suppress the cold-sensitive phenotype, whereas Era-dE, which has a 10-residue deletion in a putative effector region of the N-terminal domain, functioned as a more efficient suppressor than the wild-type Era. It was found that Era-dE suppressed defective 16S rRNA maturation, resuming a normal polysome profile to reduce highly accumulated free 30S and 50S subunits in the rbfA deletion cells. These results indicate that Era is involved in 16S rRNA maturation and ribosome assembly.
Inoue
Koichi
K
Department of Biochemistry, Robert Wood Johnson Medical School, 675 Hoes Lane, Piscataway, NJ 08854, USA.
Alsina
Janivette
J
Chen
Jingqiu
J
Inouye
Masayori
M
D015152:Blotting, Northern; D003080:Cold Temperature; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics*; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000096:biosynthesis* / Q000378:metabolism; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial; D009154:Mutation; D012329:RNA, Bacterial / Q000378:metabolism; D012336:RNA, Ribosomal, 16S / Q000378:metabolism; D016601:RNA-Binding Proteins; D012269:Ribosomal Proteins / Q000172:deficiency / Q000235:genetics*; D012270:Ribosomes / Q000378:metabolism*
D029968:Escherichia coli Proteins; D012329:RNA, Bacterial; D012336:RNA, Ribosomal, 16S; D016601:RNA-Binding Proteins; C092748:RbfA protein, E coli; D012269:Ribosomal Proteins; C068027:era protein, E coli; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins