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Infection and Immunity, March 1999, p. 1025-1033, Vol. 67, No. 3
0019-9567/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Role of DnaK in In Vitro and In Vivo Expression of Virulence Factors of Vibrio cholerae

Sabyasachi Chakrabarti,dagger Nilanjan Sengupta, and Rukhsana Chowdhury*

Biophysics Division, Indian Institute of Chemical Biology, Calcutta 700 032, India

Received 2 September 1998/Returned for modification 9 November 1998/Accepted 8 December 1998

The dnaK gene of Vibrio cholerae was cloned, sequenced, and used to construct a dnaK insertion mutant which was then used to examine the role of DnaK in expression of the major virulence factors of this important human pathogen. The central regulator of several virulence genes of V. cholerae is ToxR, a transmembrane DNA binding protein. The V. cholerae dnaK mutant grown in standard laboratory medium exhibited phenotypes characteristic of cells deficient in ToxR activity. Using Northern blot analysis and toxR transcriptional fusions, we demonstrated a reduction in expression of the toxR gene in the dnaK mutant strain together with a concomitant increase in expression of a htpG-like heat shock gene that is located immediately upstream and is divergently transcribed from toxR. This may be due to increased heat shock induction in the dnaK mutant. In vivo, however, although expression from heat shock promoters in the dnaK mutant was similar to that observed in vitro, expression of both toxR and htpG was comparable to that by the parental strain. In both strains, in vivo expression of toxR was significantly higher than that observed in vitro, but no reciprocal decrease in htpG expression was observed. These results suggest that the modulation of toxR expression in vivo may be different from that observed in vitro.

* Corresponding author. Mailing address: Biophysics Division, Indian Institute of Chemical Biology, 4 Raja S.C. Mullick Road, Calcutta 700 032, India. Phone: 91 33 473 0350. Fax: 91 33 473 0350/5197. E-mail: iichbio{at}giascl01.vsnl.net.in.

dagger Present address: Department of Cell Biology, Yale University School of Medicine, New Haven, Conn.

Infection and Immunity, March 1999, p. 1025-1033, Vol. 67, No. 3
0019-9567/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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