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Journal of Bacteriology, October 2003, p. 6158-6170, Vol. 185, No. 20
0021-9193/03/$08.00+0     DOI: 10.1128/JB.185.20.6158-6170.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Interfering with Different Steps of Protein Synthesis Explored by Transcriptional Profiling of Escherichia coli K-12{dagger}

Jeffrey Sabina,1 Nir Dover,2 Lori J. Templeton,2 Dana R. Smulski,2 Dieter Söll,1,3 and Robert A. LaRossa2*

Central Research and Development, DuPont Company, Wilmington, Delaware 19880-0173,2 Departments of Molecular Biophysics and Biochemistry,1 Chemistry, Yale University, New Haven, Connecticut 06520-81143

Received 22 May 2003/ Accepted 30 July 2003

Escherichia coli responses to four inhibitors that interfere with translation were monitored at the transcriptional level. A DNA microarray method provided a comprehensive view of changes in mRNA levels after exposure to these agents. Real-time reverse transcriptase PCRanalysis served to verify observations made with microarrays, and a chromosomal grpE::lux operon fusion was employed to specifically monitor the heat shock response. 4-Azaleucine, a competitive inhibitor of leucyl-tRNA synthetase, surprisingly triggered the heat shock response. Administration of mupirocin, an inhibitor of isoleucyl-tRNA synthetase activity, resulted in changes reminiscent of the stringent response. Treatment with kasugamycin and puromycin (targeting ribosomal subunit association as well as its peptidyl-transferase activity) caused accumulation of mRNAs from ribosomal protein operons. Abundant biosynthetic transcripts were often significantly diminished after treatment with any of these agents. Exposure of a relA strain to mupirocin resulted in accumulation of ribosomal protein operon transcripts. However, the relA strain's response to the other inhibitors was quite similar to that of the wild-type strain.

* Corresponding author. Mailing address: Central Research and Development, Biochemical Science and Engineering Experimental Station, P.O. Box 80173, Wilmington, DE 19880-0173. Phone: (302) 695-9264. Fax: (302) 695-9183. E-mail: Robert.A.LaRossa{at}usa.dupont.com.

{dagger} This work is dedicated to the memory of Professor Philip E. Hartman, Department of Biology, The Johns Hopkins University.

Journal of Bacteriology, October 2003, p. 6158-6170, Vol. 185, No. 20
0021-9193/03/$08.00+0     DOI: 10.1128/JB.185.20.6158-6170.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



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