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Journal of Bacteriology, June 2000, p. 3405-3415, Vol. 182, No. 12
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Vibrio vulnificus Has the Transmembrane Transcription Activator ToxRS Stimulating the Expression of the Hemolysin Gene vvhA

Shee Eun Lee,1,2 Sung Heui Shin,3 Soo Young Kim,1,2 Young Ran Kim,1,2 Dong Hyeon Shin,1,2 Sun Sik Chung,1,2 Zang Hee Lee,4 Jee Yeon Lee,5,6 Kwang Choel Jeong,5,6 Sang Ho Choi,5,6 and Joon Haeng Rhee1,2,*

Department of Microbiology1 and Institute of Medical Sciences,2 Chonnam National University Medical School, Department of Microbiology, Chosun University Medical School,3 Department of Microbiology and Immunology, Chosun University Dental School,4 and Department of Food Science and Technology5 and Institute of Biotechnology,6 Chonnam National University, Kwangju, Republic of Korea

Received 29 October 1999/Accepted 22 March 2000

In an attempt to dissect the virulence regulatory mechanism in Vibrio vulnificus, we tried to identify the V. cholerae transmembrane virulence regulator toxRS (toxRSVc) homologs in V. vulnificus. By comparing the sequences of toxRS of V. cholerae and V. parahaemolyticus (toxRSVp), we designed a degenerate primer set targeting well-conserved sequences. Using the PCR product as an authentic probe for Southern blot hybridization, a 1.6-kb BglII-HindIII fragment and a 1.2-kb HindIII fragment containing two complete open reading frames and one partial open reading frame attributable to toxRVv, toxSVv, and htpGVv were cloned. ToxRVv shared 55.0 and 63.0% sequence homology with ToxRVc and ToxRVp, respectively. ToxSVv was 71.5 and 65.7% homologous to ToxSVc and ToxSVp, respectively. The amino acid sequences of ToxRSVv showed transmembrane and activity domains similar to those observed in ToxRSVc and ToxRSVp. Western blot analysis proved the expression of ToxRVv in V. vulnificus. ToxRSVv enhanced, in an Escherichia coli background, the expression of the V. vulnificus hemolysin gene (vvhA) fivefold. ToxRSVv also activated the ToxRVc-regulated ctx promoter incorporated into an E. coli chromosome. A toxRVv null mutation decreased hemolysin production. The defect in hemolysin production could be complemented by a plasmid harboring the wild-type gene. The toxRVv mutation also showed a reversed outer membrane protein expression profile in comparison to the isogenic wild-type strain. These results demonstrate that ToxRVv may regulate the virulence expression of V. vulnificus.

* Corresponding author. Mailing address: Department of Microbiology, Chonnam National University Medical School, 5-1 Hak-Dong, Dong-Ku, Kwangju 501-190, Republic of Korea. Phone: 82-62-220-4136. Fax: 82-62-228-7294. E-mail: jhrhee{at}chonnam.chonnam.ac.kr.

Journal of Bacteriology, June 2000, p. 3405-3415, Vol. 182, No. 12
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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