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

Author:

Lee Shee Eun12,Shin Sung Heui3,Kim Soo Young12,Kim Young Ran12,Shin Dong Hyeon12,Chung Sun Sik12,Lee Zang Hee4,Lee Jee Yeon56,Jeong Kwang Choel56,Choi Sang Ho56,Rhee Joon Haeng12

Affiliation:

1. Department of Microbiology1 and

2. Institute of Medical Sciences,2

3. Chonnam National University Medical School, Department of Microbiology, Chosun University Medical School,3

4. Department of Microbiology and Immunology, Chosun University Dental School,4and

5. Department of Food Science and Technology5 and

6. Institute of Biotechnology,6 Chonnam National University, Kwangju, Republic of Korea

Abstract

ABSTRACT In an attempt to dissect the virulence regulatory mechanism in Vibrio vulnificus , we tried to identify the V. cholerae transmembrane virulence regulator toxRS ( toxRS Vc ) homologs in V. vulnificus . By comparing the sequences of toxRS of V. cholerae and V. parahaemolyticus ( toxRS Vp ), 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 Bgl II- Hin dIII fragment and a 1.2-kb Hin dIII fragment containing two complete open reading frames and one partial open reading frame attributable to toxR Vv , toxS Vv , and htpG Vv were cloned. ToxR Vv shared 55.0 and 63.0% sequence homology with ToxR Vc and ToxR Vp , respectively. ToxS Vv was 71.5 and 65.7% homologous to ToxS Vc and ToxS Vp , respectively. The amino acid sequences of ToxRS Vv showed transmembrane and activity domains similar to those observed in ToxRS Vc and ToxRS Vp . Western blot analysis proved the expression of ToxR Vv in V. vulnificus . ToxRS Vv enhanced, in an Escherichia coli background, the expression of the V. vulnificus hemolysin gene ( vvhA ) fivefold. ToxRS Vv also activated the ToxR Vc -regulated ctx promoter incorporated into an E. coli chromosome. A toxR Vv null mutation decreased hemolysin production. The defect in hemolysin production could be complemented by a plasmid harboring the wild-type gene. The toxR Vv mutation also showed a reversed outer membrane protein expression profile in comparison to the isogenic wild-type strain. These results demonstrate that ToxR Vv may regulate the virulence expression of V. vulnificus .

Publisher

American Society for Microbiology

Subject

Molecular Biology,Microbiology

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