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Journal of Bacteriology, August 2007, p. 5779-5781, Vol. 189, No. 15
0021-9193/07/$08.00+0     doi:10.1128/JB.00453-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Thermoregulation of Escherichia coli hchA Transcript Stability

Aviram Rasouly, Yotam Shenhar, and Eliora Z. Ron^*

Department of Molecular Microbiology and Biotechnology, George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel, 69978

Received 27 March 2007/ Accepted 18 May 2007

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The conserved chaperone Hsp31 of Escherichia coli is transcribed at low temperatures by ^S and repressed by H-NS, whereas at high temperature, transcription is by ⁷⁰ independently of both ^S and H-NS. Here we present evidence for an additional, novel, temperature-dependent control of Hsp31 expression by increased transcript stability.

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Heat shock protein 31 (Hsp31), the product of the hchA (yedU) gene, is a conserved chaperone (7). Like all other chaperones, Hsp31 is a heat shock gene product, whose level is significantly up-regulated upon 7-min heat shock at 51°C (6). Strains with a mutation in the hchA gene show only a mild growth defect at high temperature, but they are severely defective in their ability to recover from exposure to lethal temperatures (4). Recent results indicated that the level of Hsp31 is induced upon temperature upshifts even in an rpoH mutant lacking ³² (5). It was shown that the hchA gene is transcriptionally regulated by the ^S transcriptional activator, which regulates the general stress response in Escherichia coli (2, 5, 8), and that the heat shock induction is presumably due to derepression by H-NS (5).

Here we show the existence of an additional, H-NS-independent thermal induction of hchA. This mechanism involves a significant increase in the stability of hchA transcripts upon a shift to higher temperatures. This is a novel method of thermoregulation to control the temperature-dependent expression of a chaperone.

Using quantitative real-time reverse transcription (RT)-PCR, the transcript levels of hchA were monitored at 30°C and 42°C in wild-type cultures and in mutants defective for RpoS and H-NS. The results were in agreement with previous findings showing a higher concentration of Hsp31 in H-NS deletion mutants and indicate that, at low temperature, hchA transcription is carried out by RpoS and repressed by H-NS (5). These results led to the assumption that H-NS represses hchA transcription at low temperature and that its release—at high temperature—derepresses the system. On the basis of this assumption, one would expect no induction of hchA in hns deletion mutants following temperature upshifts. However, the results presented in Fig. 1 clearly indicate that elevation from 30°C to 42°C results in a sevenfold induction of hchA even in cells depleted of H-NS. This unexpected temperature induction is independent of RpoS and is also independent of the heat shock sigma factor RpoH. These results indicated that hchA induction at 42°C cannot be explained by the release of H-NS alone.

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FIG. 1. Temperature-dependent induction of hchA in hns mutants. The levels of hchA transcripts were determined by RT-PCR in K-12 strain MG1655 and its mutant with a deletion of hns (1). Cultures were grown in Luria-Bertani medium with aeration to mid-log phase and shifted from 30°C to 42°C for 10 min. RNA isolation and quantitative real-time PCR were performed as previously described (3). Each experiment was based on at least four independent biological samples, each of which was run in triplicate. Bars show standard deviations.

Moreover, microarray analyses indicated that the heat shock induction of hchA is kinetically distinct from that of other heat shock genes. The global transcriptional response of E. coli MG1655 was determined at 5 and 15 min following a temperature shift from 30°C to 42°C. In these experiments, the transcript levels of known ³²-regulated heat shock genes were considerably lower after 15 min than at 5 min following the shift. These results reflect the previously reported transient nature of ³²-dependent thermoregulation. In contrast, the level of the hchA transcript was higher after 15 min of heat shock (Table 1). Moreover, we could show by quantitative RT-PCR that, at 42°C, hchA transcript levels were high for at least 60 min. At this time point, the level of transcript was still about 100-fold higher than before the temperature shift (Fig. 2). The high levels of Hsp31 and of hchA transcription that extended for long periods at 42°C were also noted previously (5). A possible explanation for these data are that the shift to higher temperatures involves a change in hchA transcript stability.

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TABLE 1. Relative transcript levels of heat shock genes encoding chaperones and proteases following a shift to 42°Ca

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FIG. 2. Levels of hchA transcripts following transfer to 42°C. The experiment was performed as described for Fig. 1, using the wild-type strain MG1655. Samples were taken at 0, 10, 30, and 60 min.

The stability of hchA transcripts was examined by quantitative RT-PCR determination of residual transcripts following the addition of the transcription inhibitor rifampin. The results summarized in Fig. 3 indicate that, indeed, hchA transcripts were considerably more stable at 42°C than at 30°C. The stabilization of transcripts at elevated temperatures is remarkable and not typical for other heat shock genes. The effects of temperature on the transcript levels of degP, also a non-³²-induced heat shock gene, are shown for comparison (Fig. 4).

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FIG. 3. Stability of hchA transcripts at elevated temperatures. Cultures of strain MG1655 were grown as described for Fig. 1. At an optical density at 600 nm of 0.4, half of the culture was transferred to 42°C and rifampin (500 µg/ml) was added to the two cultures after 5 min (t = 0). Samples were taken at intervals and treated as described for Fig. 1.

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FIG. 4. Stability of degP transcripts at elevated temperatures. The experiment was performed as described for Fig. 3.

In conclusion, the temperature-dependent induction of hchA transcription in E. coli is regulated by H-NS (5). However, this control mechanism offers only a partial explanation for the heat-induced hchA transcription, as there is a sevenfold induction in the level of hchA transcripts in hns deletion mutants, indicating the existence of an additional, H-NS-independent thermoregulation. One such additional control is the temperature-dependent increased stability of hchA transcripts, for which we present evidence in this communication. This thermoregulation of chaperone expression by transcript stabilization is unique and is an additional level of control of the expression of a chaperone gene whose transcription is controlled by the interplay of ^S and H-NS.

 
We thank Dvora Biran, Chen Katz, and Itzhak Mizrahi for helpful discussions.

This work was supported, in part, by the Morris and Manja Leigh Chair for Biophysics and Biotechnology.

 
* Corresponding author. Mailing address: Department of Molecular Microbiology and Biotechnology, George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, Israel, 69978. Phone: 972-3-6409379. Fax: 972-36414138. E-mail: eliora{at}post.tau.ac.il

Published ahead of print on 25 May 2007.

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1
1. Datsenko, K. A., and B. L. Wanner. 2000. One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc. Natl. Acad. Sci. USA 97:6640-6645.[Abstract/Free Full Text]
2
2. Lange, R., and R. Hengge-Aronis. 1991. Identification of a central regulator of stationary-phase gene expression in Escherichia coli. Mol. Microbiol. 5:49-59.[Medline]
3
3. Mizrahi, I., D. Biran, and E. Z. Ron. 2006. Requirement for the acetyl phosphate pathway in Escherichia coli ATP-dependent proteolysis. Mol. Microbiol. 62:201-211.[CrossRef][Medline]
4
4. Mujacic, M., M. W. Bader, and F. Baneyx. 2004. Escherichia coli Hsp31 functions as a holding chaperone that cooperates with the DnaK-DnaJ-GrpE system in the management of protein misfolding under severe stress conditions. Mol. Microbiol. 51:849-859.[CrossRef][Medline]
5
5. Mujacic, M., and F. Baneyx. 2006. Regulation of Escherichia coli hchA, a stress-inducible gene encoding molecular chaperone Hsp31. Mol. Microbiol. 60:1576-1589.[CrossRef][Medline]
6
6. Richmond, C. S., J. D. Glasner, R. Mau, H. Jin, and F. R. Blattner. 1999. Genome-wide expression profiling in Escherichia coli K-12. Nucleic Acids Res. 27:3821-3835.[Abstract/Free Full Text]
7
7. Sastry, M. S., K. Korotkov, Y. Brodsky, and F. Baneyx. 2002. Hsp31, the Escherichia coli yedU gene product, is a molecular chaperone whose activity is inhibited by ATP at high temperatures. J. Biol. Chem. 277:46026-46034.[Abstract/Free Full Text]
8
8. Weber, H., T. Polen, J. Heuveling, V. F. Wendisch, and R. Hengge. 2005. Genome-wide analysis of the general stress response network in Escherichia coli: ^S-dependent genes, promoters, and sigma factor selectivity. J. Bacteriol. 187:1591-1603.[Abstract/Free Full Text]

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Journal of Bacteriology, August 2007, p. 5779-5781, Vol. 189, No. 15
0021-9193/07/$08.00+0     doi:10.1128/JB.00453-07
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Rasouly, A. Articles by Ron, E. Z. Search for Related Content PubMed PubMed Citation Articles by Rasouly, A. Articles by Ron, E. Z.