Browse > Article
http://dx.doi.org/10.4014/jmb.0807.404

Role of Alkaline Serine Protease, Asp, in Vibrio alginolyticus Virulence and Regulation of Its Expression by LuxO-LuxR Regulatory System  

Rui, Haopeng (State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology)
Liu, Qin (State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology)
Wang, Qiyao (State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology)
Ma, Yue (State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology)
Liu, Huan (State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology)
Shi, Cunbin (Pearl River Fisheries Research Institute, Chinese Academy of Fishery Science)
Zhang, Yuanxing (State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology)
Publication Information
Journal of Microbiology and Biotechnology / v.19, no.5, 2009 , pp. 431-438 More about this Journal
Abstract
The alkaline serine protease asp, which was shown to be a virulence factor of Vibrio alginolyticus as a purified protein, was cloned from V. alginolyticus EPGS, a strain recently isolated from moribund Epinephelus coioides in an outbreak of vibriosis in a mariculture farm of Shenzhen. The asp null mutant was constructed by homologous recombination with suicide plasmid pNQ705-1. Compared with the wild-type strain, the asp null mutant exhibited a significant decrease of total extracellular protease activity, and caused a IS-fold decrease in virulence of V. alginolyticus. In our previous study, the luxO and $luxR_{val}$ genes from V. alginolyticus MVP01 were cloned and identified, and the luxO-$luxR_{val}$ regulatory couple was shown to regulate various genes expression, suggesting that it played a central role in the quorum sensing system of V. alginolyticus. In this study, the regulation of the asp gene was analyzed by using RT-PCR and quantitative real-time PCR methods; we proved that its transcription was greatly induced at the late stage of growth and was regulated by a luxO-$luxR_{val}$ regulatory system.
Keywords
Alkaline serine protease; asp; quorum sesing; Vibrio alginolyticus;
Citations & Related Records

Times Cited By Web Of Science : 15  (Related Records In Web of Science)
연도 인용수 순위
  • Reference
1 Balebona, M. C., M. J. Andreu, M. A. Bordas, I. Zorrilla, M. A. Morinigo, and J. J. Borrego. 1998. Pathogenicity of Vibrio alginolyticus for cultured gilt-head sea bream (Sparus aurata L.). Appl. Environ. Microbiol. 64: 4269-4275   PUBMED   ScienceOn
2 Croxatto, A., V. J. Chalker, J. Lauritz, J. Jass, A. Hardman, P. Williams, M. Camara, and D. L. Milton. 2002. VanT, a homologue of Vibrio harveyi LuxR, regulates serine, metalloprotease, pigment, and biofilm production in Vibrio anguillarum. J. Bacteriol. 184: 1617-1629   DOI   ScienceOn
3 Hare, P., T. Scott-Burden, and D. R. Woods. 1983. Characterization of extracellular alkaline proteases and collagenase induction in Vibrio alginolyticus. J. Gen. Microbiol. 129: 1141-1147   DOI   PUBMED   ScienceOn
4 Jobling, M. G. and R. K. Holmes. 1997. Characterization of hapR, a positive regulator of the Vibrio cholerae HA/protease gene hap, and its identification as a functional homologue of the Vibrio harveyi luxR gene. Mol. Microbiol. 26: 1023-1034   DOI   ScienceOn
5 Lee, K. K., S. R. Yu, and P. C. Liu. 1997. Alkaline serine protease is an exotoxin of Vibrio alginolyticus in kuruma prawn, Penaeus japonicus. Curr. Microbiol. 34: 110-117   DOI   ScienceOn
6 Liu, C. H., W. Cheng, J. P. Hsu, and J. C. Chen. 2004. Vibrio alginolyticus infection in the white shrimp Litopenaeus vannamei confirmed by polymerase chain reaction and 16S rDNA sequencing. Dis. Aquat. Organ. 61: 169-174   DOI   ScienceOn
7 Shao, C. P. and L. I. Hor. 2001. Regulation of metalloprotease gene expression in Vibrio vulnificus by a Vibrio harveyi LuxR homologue. J. Bacteriol. 183: 1369-1375   DOI   ScienceOn
8 Surette, M. G., M. B. Miller, and B. L. Bassler. 1999. Quorum sensing in Escherichia coli, Salmonella typhimurium, and Vibrio harveyi: A new family of genes responsible for autoinducer production. Proc. Natl. Acad. Sci. U.S.A. 96: 1639-1644   DOI   ScienceOn
9 Swift, S., M. J. Lynch, L. Fish, D. F. Kirke, J. M. Tomas, G. S. Stewart, and P. Williams. 1999. Quorum sensing-dependent regulation and blockade of exoprotease production in Aeromonas hydrophila. Infect. Immun. 67: 5192-5199   PUBMED
10 Ye, J., Y. Ma, Q. Liu, D. L. Zhao, Q. Y. Wang, and Y. X. Zhang. 2008. Regulation of Vibrio alginolyticus virulence by the LuxS quorum-sensing system. J. Fish Dis. 31: 161-169   DOI   ScienceOn
11 Bassler, B. L., M. Wright, R. E. Showalter, and M. R Silverman. 1993. Intercellular signalling in Vibrio harveyi: Sequence and function of genes regulating expression of luminescence. Mol. Microbiol. 9: 773-786   DOI   ScienceOn
12 Park, K. S., T. Ono, M. Rokuda, M. H. Jang, K. Okada, T. Iida, and T. Honda. 2004. Functional characterization of two type III secretion systems of Vibrio parahaemolyticus. Infect. Immun. 72: 6659-6665   DOI   ScienceOn
13 Croxatto, A., J. Pride, A. Hardman, P. Williams, M. Camara, and D. L. Milton. 2004. A distinctive dual-channel quorumsensing system operates in Vibrio anguillarum. Mol. Microbiol. 52: 1677-1689   DOI   ScienceOn
14 Deane, S. M., F. T. Robb, S. M. Robb, and D. R. Woods. 1989. Nucleotide sequence of the Vibrio alginolyticus calcium-dependent, detergent-resistant alkaline serine exoprotease A. Gene 76: 281-288   DOI   ScienceOn
15 Wang, Q., Q. Liu, Y. Ma, H. Rui, and Y. Zhang. 2007. LuxO controls extracellular protease, haemolytic activities and siderophore production in fish pathogen Vibrio alginolyticus. J. Appl. Microbiol. 103: 1525-1534   DOI   ScienceOn
16 Waters, C. M. and B. L. Bassler. 2005. Quorum sensing: Cellto- cell communication in bacteria. Annu. Rev. Cell Dev. Biol. 21: 319-346   DOI   ScienceOn
17 Zhu, J., M. B. Miller, R. E. Vance, M. Dziejman, B. L. Bassler, and J. J. Mekalanos. 2002. Quorum-sensing regulators control virulence gene expression in Vibrio cholerae. Proc. Natl. Acad. Sci. U.S.A. 99: 3129-3134   DOI   ScienceOn
18 Livak, K. J. and T. D. Schmittgen. 2001. Analysis of relative gene expression data using real-time quantitative PCR and the $2^{-\Delta\Delta C}T$ method. Methods 25: 402-408   DOI   ScienceOn
19 Aguirre-Guzman, G., H. Ruiz, and F. Ascencio. 2004. A review of extracellular virulence product of Vibrio species important in diseases of cultivated shrimp. Aquac. Res. 35: 1395-1404   DOI   ScienceOn
20 Dennis, J. J. and G. J. Zylstra. 1998. Plasposons: Modular selfcloning minitransposon derivatives for rapid genetic analysis of Gram-negative bacterial genomes. Appl. Environ. Microbiol. 64:2710-2715   PUBMED   ScienceOn
21 Takeuchi, H., Y. Shibano, K. Morihara, J. Fukushima, S. Inami, B. Keil, A. M. Gilles, S. Kawamoto, and K. Okuda. 1992. Structural gene and complete amino acid sequence of Vibrio alginolyticus collagenase. Biochem. J. 281: 703-708   DOI   ScienceOn
22 Denkin, S. M. and D. R. Nelson. 1999. Induction of protease activity in Vibrio anguillarum by gastrointestinal mucus. Appl. Environ. Microbiol. 65: 3555-3560   PUBMED   ScienceOn
23 Gomez-Leon, J., L. Villamil, M. L. Lemos, B. Novoa, and A. Figueras. 2005. Isolation of Vibrio alginolyticus and Vibrio splendidus from aquacultured carpet shell clam (Ruditapes decussatus) larvae associated with mass mortalities. Appl. Environ. Microbiol. 71: 98-104   DOI   ScienceOn
24 Liang, W., S. Wang, F. Yu, L. Zhang, G. Qi, Y. Liu, S. Gao, and B. Kan. 2003. Construction and evaluation of a safe, live, oral Vibrio cholerae vaccine candidate, IEM108. Infect. Immun. 71:5498-5504   DOI   ScienceOn
25 Miyamoto, C. M., P. V. Dunlap, E. G. Ruby, and E. A. Meighen. 2003. LuxO controls luxR expression in Vibrio harveyi: Evidence for a common regulatory mechanism in Vibrio. Mol. Microbiol. 48: 537-548   DOI   ScienceOn
26 Liu, Q., P. Wang, Y. Ma, and Y. Zhang. 2007. Characterization of the Vibrio alginolyticus fur gene and localization of essential amino acid sites in Fur by site-directed mutagenesis. J. Mol. Microbiol. Biotechnol. 13: 15-21   DOI   ScienceOn
27 Sultan, Z., S. Miyoshi, and S. Shinoda. 2006. Presence of LuxS/ AI-2 based quorum-sensing system in Vibrio mimicus: LuxO controls protease activity. Microbiol. Immunol. 50: 407-417   DOI   PUBMED
28 Sambrook, J., E. Fritsch, and T. Maniatis. 1989. Molecular Cloning: A Laboratory Manual, 2nd Ed, Cold Spring Harbor Laboratory, Press Cold Spring Harbor, New York, U.S.A
29 Long, S., M. A. Mothibeli, F. T. Robb, and D. R. Woods. 1981. Regulation of extracellular alkaline protease activity by histidine in a collagenolytic Vibrio alginolyticus strain. J. Gen. Microbiol. 127: 193-199   DOI   PUBMED   ScienceOn
30 Reed, L. J. and H. Muench 1938. A simple method of estimating fifty percent end points. Am. J. Hyg. 27: 493-497
31 Bassler, B. L. and R. Losick. 2006. Bacterially speaking. Cell 125: 237-246   DOI   ScienceOn
32 Rui, H., Q. Liu, Y. Ma, Q. Wang, and Y. Zhang. 2008. Roles of LuxR in regulating extracellular alkaline serine protease A, extracellular polysaccharide and mobility of Vibrio alginolyticus. FEMS Microbiol. Lett. 285: 155-162   DOI   ScienceOn
33 Cai, S. H., Z. H. Wu, J. C. Jian, and Y. S. Lu. 2007. Cloning and expression of the gene encoding an extracellular alkaline serine protease from Vibrio alginolyticus strain HY9901, the causative agent of vibriosis in Lutjanus erythopterus (Bloch). J. Fish Dis. 30: 493-500   DOI   ScienceOn
34 Storey, D. G., E. E. Ujack, H. R. Rabin, and I. Mitchell. 1998. Pseudomonas aeruginosa lasR transcription correlates with the transcription of lasA, lasB, and toxA in chronic lung infections associated with cystic fibrosis. Infect. Immun. 66: 2521-2528   PUBMED   ScienceOn