Physiological analysis of the role of truB in Escherichia coli: a role for tRNA modification in extreme temperature resistance

• Published in: Microbiology (Society for General Microbiology) Vol. 148; no. 11; pp. 3511 - 3520
• Main Authors: Kinghorn, Seonag M, O'Byrne, Conor P, Booth, Ian R, Stansfield, Ian
• Format: Journal Article
• Language: English
• Published: Reading Soc General Microbiol 01.11.2002; Society for General Microbiology
• Subjects: Action of physical and chemical agents on bacteria; Bacterial Outer Membrane Proteins - metabolism; Bacteriology; Biological and medical sciences; Escherichia coli; Escherichia coli - metabolism; Escherichia coli - physiology; Fundamental and applied biological sciences. Psychology; Gene Deletion; Hot Temperature; Intramolecular Lyases - genetics; Intramolecular Lyases - physiology; Intramolecular Transferases; Microbiology; OmpA protein; OmpX protein; Phenotype; pseudouridine-55 ( psi 55) synthase; RNA, Transfer - metabolism; Saccharomyces cerevisiae Proteins; sigma E Factor; Sigma Factor - metabolism; Transcription Factors - metabolism; trmA gene; tRNA Methyltransferases - genetics; tRNA Methyltransferases - physiology; truB gene; Bacteria; tRNA; Thermal resistance; Enzyme; Escherichia coli; Enterobacteriaceae
• ISSN: 1350-0872; 1465-2080
• DOI: 10.1099/00221287-148-11-3511

Department of Molecular & Cell Biology, University of Aberdeen, Institute of Medical Sciences, Foresterhill, Aberdeen AB25 2ZD, UK 1 Author for correspondence: Ian Stansfield. Tel: +44 1224 273106. Fax: +44 1224 273144. e-mail: i.stansfield{at}abdn.ac.uk The truB gene of Escherichia coli encodes the pseudouridine-55 ( 55) synthase and is responsible for modifying all tRNA molecules in the cell at the U55 position. A truB null mutant grew normally on all growth media tested, but exhibited a competitive disadvantage in extended co-culture with its wild-type progenitor. The mutant phenotype could be complemented by both the cloned truB gene and by a D48C, catalytically inactive allele of truB . The truB mutant also exhibited a defect in survival of rapid transfer from 37 to 50 °C. This mutant phenotype could be complemented by the cloned truB gene but not by a D48C, catalytically inactive allele of truB . The temperature sensitivity of truB mutants could be enhanced by combination with a mutation in the trmA gene, encoding an m 5 U-methyltransferase, modifying the universal U54 tRNA nucleoside, but not by mutations in trmH , encoding the enzyme catalysing the formation of Gm18. The truB mutant proteome contained altered levels of intermediates involved in biogenesis of the outer-membrane proteins OmpA and OmpX. The truB mutation also reduced the basal expression from two E promoters, degP and rpoH P3. Three novel aspects to the phenotype of truB mutants were identified. Importantly the data support the hypothesis that TruB-effected 55 modification of tRNA is not essential, but contributes to thermal stress tolerance in E. coli , possibly by optimizing the stability of the tRNA population at high temperatures. Keywords: pseudouridine-55 synthase, heat stress, trmA , sigma E Abbreviations: Cm, chloramphenicol; Kan, kanamycin; OMP, outer-membrane protein; Tet, tetracycline; , pseudouridine