Abstract
Resistance-nodulation-division (RND) efflux pumps are associated with multidrug resistance in many gram-negative pathogens. The genome of Vibrio vulnificus encodes 11 putative RND pumps homologous to those of Vibrio cholerae and Escherichia coli. In this study, we analyzed three putative RND efflux pumps, showing homology to V. cholerae VexAB and VexCD and to E. coli AcrAB, for their functional roles in multidrug resistance of V. vulnificus. Deletion of the vexAB homolog resulted in increased susceptibility of V. vulnificus to bile acid, acriflavine, ethidium bromide, and erythromycin, whereas deletion of acrAB homologs rendered V. vulnificus more susceptible to acriflavine only. Deletion of vexCD had no effect on susceptibility of V. vulnificus to these chemicals. Upon exposure to these antibacterial chemicals, expression of tolCV1 and tolCV2, which are putative outer membrane factors of RND efflux pumps, was induced, whereas expression levels of vexAB, vexCD, and acrAB homologs were not significantly changed. Our results show that the V. vulnificus homologs of VexAB largely contributed to in vitro antimicrobial resistance with a broad substrate specificity that was partially redundant with the AcrAB pump homologs.
Article PDF
Similar content being viewed by others
Avoid common mistakes on your manuscript.
References
Bavro, V.N., Pietras, Z., Furnham, N., Pérez-Cano, L., Fernández-Recio, J., Pei, X.Y., Misra, R., and Luisi, B. 2008. Assembly and channel opening in a bacterial drug efflux machine. Mol. Cell. 30, 114–121.
Bina, J.E., Alm, R.A., Uria-Nickelsen, M., Thomas, S.R., Trust, T.J., and Hancock, R.E. 2000. Helicobacter pylori uptake and efflux: basis for intrinsic susceptibility to antibiotics in vitro. Antimicrob. Agents Chemother. 44, 248–254.
Bina, J.E., Provenzano, D., Wang, C., Bina, X.R., and Mekalanos, J.J. 2006. Characterization of the Vibrio cholerae vexAB and vexCD efflux systems. Arch. Microbiol. 186, 171–181.
Du, D., Wang, Z., James, N.R., Voss, J.E., Klimont, E., Ohene-Agyei, T., Venter, H., Chiu, W., and Luisi, B.F. 2014. Structure of the AcrAB-TolC multidrug efflux pump. Nature 509, 512–515.
Hwang, W., Lee, N.Y., Kim, J., Lee, M.A., Kim, K.S., Lee, K.H., and Park, S.J. 2011. Functional characterization of EpsC, a component of the Type II secretion system, in the pathogenicity of Vibrio vulnificus. Infect. Immun. 79, 4068–4080.
Kawano, H., Miyamoto, K., Yasunobe, M., Murata, M., Myojin, T., Tsuchiya, T., Tanabe, T., Funahashi, T., Sato, T., Azuma, T., et al. 2014. The RND protein is involved in the vulnibactin export system in Vibrio vulnificus M2799. Microb. Pathog. 75, 59–67.
Kim, H.M., Xu, Y., Lee, M., Piao, S., Sim, S.H., Ha, N.C., and Lee, K. 2010. Functional relationships between the AcrA hairpin tip region and the TolC aperture tip region for the formation of the bacterial tripartite efflux pump AcrAB-TolC. J. Bacteriol. 192, 4498–4503.
Kim, I.H., Wen, Y., Son, J.S., Lee, K.H., and Kim, K.S. 2013. The furiron complex modulates expression of the quorum-sensing master regulator, SmcR, to control expression of virulence factors in Vibrio vulnificus. Infect. Immun. 81, 2888–2898.
Kohler, T., Michea-Hamzehpour, M., Henze, U., Gotoh, N., Curty, L.K., and Pechère, J.C. 1997. Characterization of MexE-MexFOprN, a positively regulated multidrug efflux system of Pseudomonas aeruginosa. Mol. Microbiol. 23, 345–354.
Koronakis, V., Eswaran, J., and Hughes, C. 2004. Structure and function of TolC: the bacterial exit duct for proteins and drugs. Annu. Rev. Biochem. 73, 467–489.
Le Roux, F., Binesse, J., Saulnier, D., and Mazel, D. 2007. Construction of a Vibrio splendidus mutant lacking the metalloprotease gene vsm by use of a novel counterselectable suicide vector. Appl. Environ. Microbiol. 73, 777–784.
Lee, M., Jun, S.Y., Yoon, B.Y., Song, S., Lee, K., and Ha, N.C. 2012. Membrane fusion proteins of type I secretion system and tripartite efflux pumps share a binding motif for TolC in Gram-negative bacteria. PLoS One 7, e40460.
Lee, M., Kim, H.L., Song, S., Joo, M., Lee, S., Kim, D., Hahn, Y., Ha, N.C., and Lee, K. 2013. The α-barrel tip region of Escherichia coli TolC homologs of Vibrio vulnificus interacts with the MacA protein to form the functional macrolide-specific efflux pump MacABTolC. J. Microbiol. 51, 154–159.
Lee, S., Song, S., Lee, M., Hwang, S., Kim, J.S., Ha, N.C., and Lee, K. 2014. Interaction between the α-barrel tip of Vibrio vulnificus TolC homologs and AcrA implies the adapter bridging model. J. Microbiol. 52, 148–153.
Li, X.Z., Nikaido, H., and Poole, K. 1995. Role of mexA-mexB-oprM in antibiotic efflux in Pseudomonas aeruginosa. Antimicrob. Agents Chemother. 39, 1948–1953.
Lim, B., Ahn, S., Sim, M., and Lee, K. 2014. RNase III controls mltD mRNA degradation in Escherichia coli. Curr. Microbiol. 68, 518–523.
Misra, R. and Bavro, V.N. 2009. Assembly and transport mechanism of tripartite drug efflux systems. Biochim. Biophys. Acta. 1794, 817–825.
Nishino, K., Latifi, T., and Groisman, E.A. 2006. Virulence and drug resistance roles of multidrug efflux systems of Salmonella enterica serovar Typhimurium. Mol. Microbiol. 59, 126–141.
Park, J., Cho, Y., Chun, J., Seok, Y., Lee, J., Kim, K., Lee, K., Park, S., and Choi, S. 2011. Complete genome sequence of Vibrio vulnificus MO6-24/O. J. Bacteriol. 193, 2062–2063.
Poole, K., Tetro, K., Zhao, Q., Neshat, S., Heinrichs, D.E., and Bianco, N. 1996. Expression of the multidrug resistance operon mexAmexB-oprM in Pseudomonas aeruginosa: mexR encodes a regulator of operon expression. Antimicrob. Agents Chemother. 40, 2021–2028.
Rahman, M.M., Matsuo, T., Ogawa, W., Koterasawa, M., Kuroda, T., and Tsuchiya, T. 2007. Molecular cloning and characterization of all RND-type efflux transporters in Vibrio cholerae non-O1. Microbiol. Immunol. 51, 1061–1070.
Son, M., Moon, Y., Oh, M.J., Han, S.B., Park, K.H., Kim, J.G., and Ahn, J.H. 2012. Lipase and protease double-deletion mutant of Pseudomonas fluorescens suitable for extracellular protein production. Appl. Environ. Microbiol. 78, 8454–8462.
Song, S., Hwang, S., Lee, S., Ha, N.C., and Lee, K. 2014. Interaction mediated by the putative tip regions of MdsA and MdsC in the formation of a Salmonella-specific tripartite efflux pump. PLoS One 9, e100881.
Symmons, M.F., Bokma, E., Koronakis, E., Hughes, C., and Koronakis, V. 2009. The assembled structure of a complete tripartite bacterial multidrug efflux pump. Proc. Natl. Acad. Sci. USA 106, 7173–7178.
Tamura, N., Murakami, S., Oyama, Y., Ishiguro, M., and Yamaguchi, A. 2005. Direct interaction of multidrug efflux transporter AcrB and outer membrane channel TolC detected via site-directed disulfide cross-linking. Biochemistry 44, 11115–11121.
Val, M.E., Kennedy, S.P., El Karoui, M., Bonne, L., Chevalier, F., and Barre, F.X. 2008. FtsK-dependent dimer resolution on multiple chromosomes in the pathogen Vibrio cholerae. PLoS Genet. 4, e1000201.
Webber, M.A. and Piddock, L.J. 2003. The importance of efflux pumps in bacterial antibiotic resistance. J. Antimicrob. Chemother. 51, 9–11.
Wright, A.C., Morris, J.G. Jr., Maneval, D.R. Jr., Richardson, K., and Kaper, J.B. 1985. Cloning of the cytotoxin-hemolysin gene of Vibrio vulnificus. Infect. Immun. 50, 922–924.
Xu, Y., Lee, M., Moeller, A., Song, S., Yoon, B.Y., Kim, H.M., Jun, S.Y., Lee, K., and Ha, N.C. 2011a. Funnel-like hexameric assembly of the periplasmic adapter protein in the tripartite multidrug efflux pump in Gram-negative bacteria. J. Biol. Chem. 286, 17910–17920.
Xu, Y., Song, S., Moeller, A., Kim, N., Piao, S., Sim, S.H., Kang, M., Yu, W., Cho, H.S., Chang, I., et al. 2011b. Functional implications of an intermeshing cogwheel-like interaction between TolC and MacA in the action of macrolide-specific efflux pump MacAB-TolC. J. Biol. Chem. 286, 13541–13549.
Zgurskaya, H.I. and Nikaido, H. 1999. Bypassing the periplasm: reconstitution of the AcrAB multidrug efflux pump of Escherichia coli. Proc. Natl. Acad. Sci. USA 96, 7190–7195.
Author information
Authors and Affiliations
Corresponding authors
Additional information
These authors contributed equally to this work.
Rights and permissions
About this article
Cite this article
Lee, S., Yeom, JH., Seo, S. et al. Functional analysis of Vibrio vulnificus RND efflux pumps homologous to Vibrio cholerae VexAB and VexCD, and to Escherichia coli AcrAB. J Microbiol. 53, 256–261 (2015). https://doi.org/10.1007/s12275-015-5037-0
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s12275-015-5037-0