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Infection and Immunity, September 2005, p. 5458-5467, Vol. 73, No. 9
0019-9567/05/$08.00+0     doi:10.1128/IAI.73.9.5458-5467.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Examination of the Coordinate Effects of Pseudomonas aeruginosa ExoS on Rac1

Claudia L. Rocha,2,{dagger} Elizabeth A. Rucks,1,{dagger} Deanne M. Vincent,1,{dagger} and Joan C. Olson1*

Department of Microbiology, Immunology and Cell Biology, West Virginia University Health Sciences Center, Morgantown, West Virginia 26505,1 Department of Pathology and Laboratory Medicine, Medical University of South Carolina, Charleston, South Carolina 294252

Received 22 December 2004/ Returned for modification 1 February 2005/ Accepted 9 May 2005

Exoenzyme S (ExoS) is a bifunctional toxin directly translocated into eukaryotic cells by the Pseudomonas aeruginosa type III secretory (TTS) process. The amino-terminal GTPase-activating (GAP) activity and the carboxy-terminal ADP-ribosyltransferase (ADPRT) activity of ExoS have been found to target but exert opposite effects on the same low-molecular-weight G protein, Rac1. ExoS ADP-ribosylation of Rac1 is cell line dependent. In HT-29 human epithelial cells, where Rac1 is ADP-ribosylated by TTS-ExoS, Rac1 was activated and relocalized to the membrane fraction. Arg66 and Arg68 within the GTPase-binding region of Rac1 were identified as preferred sites of ExoS ADP-ribosylation. The modification of these residues by ExoS would be predicted to interfere with Rac1 inactivation and explain the increase in active Rac1 caused by ExoS ADPRT activity. Using ExoS-GAP and ADPRT mutants to examine the coordinate effects of the two domains on Rac1 function, limited effects of ExoS-GAP on Rac1 inactivation were evident in HT-29 cells. In J774A.1 macrophages, where Rac1 was not ADP-ribosylated, ExoS caused a decrease in the levels of active Rac1, and this decrease was linked to ExoS-GAP. Using immunofluorescence staining of Rac1 to understand the cellular basis for the targeting of ExoS ADPRT activity to Rac1, an inverse relationship was observed between Rac1 plasma membrane localization and Rac1 ADP-ribosylation. The results obtained from these studies have allowed the development of a model to explain the differential targeting and coordinate effects of ExoS GAP and ADPRT activity on Rac1 within the host cell.

* Corresponding author. Mailing address: Department of Microbiology, Immunology and Cell Biology, Robert C. Byrd Health Sciences Center, PO Box 9177, West Virginia University, Morgantown, WV 26506-9177. Phone: (304) 293-5843. Fax: (304) 293-7823. E-mail: jolson{at}hsc.wvu.edu.

Editor: D. L. Burns

{dagger} C.L.R., E.A.R., and D.M.V. contributed equally to this study.

Infection and Immunity, September 2005, p. 5458-5467, Vol. 73, No. 9
0019-9567/05/$08.00+0     doi:10.1128/IAI.73.9.5458-5467.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Stirling, F. R., Evans, T. J. (2006). Effects of the type III secreted pseudomonal toxin ExoS in the yeast Saccharomyces cerevisiae.. Microbiology 152: 2273-2285 [Abstract] [Full Text]  


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