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Journal of Biochemistry Advance Access originally published online on January 16, 2009
Journal of Biochemistry 2009 145(4):461-466; doi:10.1093/jb/mvp004
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© The Authors 2009. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved

Colicin E5 Ribonuclease Domain Cleaves Saccharomyces cerevisiae tRNAs Leading to Impairment of the Cell Growth

Tetsuhiro Ogawa1, Makoto Hidaka1, Kenji Kohno2 and Haruhiko Masaki1,*

1Department of Biotechnology, The University of Tokyo, Yayoi, Bunkyo-ku, Tokyo 113-8657; and 2Graduate School of Biological Sciences, Nara Institute of Science and Technology (NAIST), Takayama, Ikoma, Nara 630-0192, Japan

*To whom correspondence should be addressed. Tel: +81 3 5841 3080, Fax: +81 3 5841 8016, E-mail: amasaki{at}mail.ecc.u-tokyo.ac.jp

Received November 24, 2008; Accepted December 19, 2008


  Abstract

Colicin E5 is a ribonuclease that specifically cleaves tRNATyr, tRNAHis, tRNAAsn and tRNAAsp of sensitive Escherichia coli cells by recognizing their anticodon sequences. Since all organisms possess universal anticodons of these tRNAs, colicin E5 was expected to potentially cleave eukaryotic tRNAs. Here, we expressed the active domain of colicin E5 (E5-CRD) in Saccharomyces cerevisiae and investigated its effects on growth. E5-CRD impaired growth of host cells by cleaving tRNATyr, tRNAHis, tRNAAsn and tRNAAsp in S. cerevisiae, which is the same repertoire as that in E. coli. This activity of E5-CRD was inhibited by the co-expression of its cognate inhibitor (ImmE5). Notably, the growth impairment by E5-CRD was reversible; cells restored the colony-forming activity after suppression of the E5-CRD expression. This seems different from the sharp killing effect of E5-CRD on E. coli. These results may provide insights into the role and behaviour of cytosolic tRNAs on cell growth and proliferation.

Key Words: transfer RNA, ribonuclease, Saccharomyces cerevisisiae, colicin, growth arrest

Abbreviations: BPB, bromophenol blue; BSA, bovine serum albumin; EDTA, ethylenediaminetetraacetic acid; PCR, polymerase chain reaction; Q, queuine; SD, synthetic dextrose medium; SG, synthetic galactose medium; TBE, Tris–borate–EDTA buffer; TGT, tRNA-guanine transglycosylase; XC, xylene cyanol; YPD, yeast peptone dextrose medium; {Psi}, pseudouridine; dGpdUp, deoxyguanylyl(3'-5')deoxyuridine 3'-monophosphate


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