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Infection and Immunity, May 2003, p. 2787-2797, Vol. 71, No. 5
0019-9567/03/$08.00+0     DOI: 10.1128/IAI.71.5.2787-2797.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Modulation of Myosin Light-Chain Phosphorylation by p21-Activated Kinase 1 in Escherichia coli Invasion of Human Brain Microvascular Endothelial Cells

Rajyalakshmi S. Rudrabhatla,1 Sunil K. Sukumaran,1 Gary M. Bokoch,2 and Nemani V. Prasadarao1*

Division of Infectious Diseases, Childrens Hospital Los Angeles and Keck School of Medicine, University of Southern California, Los Angeles, California 90027,1 Department of Immunology and Cell Biology, The Scripps Research Institute, La Jolla, California 920372

Received 31 October 2002/ Returned for modification 21 January 2003/ Accepted 6 February 2003

Cytoskeletal dynamics, modulated by actin-myosin interactions, play an important role in Escherichia coli K1 invasion of human brain microvascular endothelial cells (HBMEC). Herein, we show that inhibitors of myosin function, butanedione monoxide and ML-7, significantly blocked the E. coli invasion of HBMEC. The invasive E. coli induces myosin light-chain (MLC) phosphorylation during the invasion process, which gets recruited to the site of actin condensation beneath the bacteria. We also show that invading E. coli downregulates the activity of p21-activated kinase 1 (PAK1), which is an upstream regulator of MLC kinase (MLCK). Overexpression of wild-type PAK1 and constitutively active PAK1 in HBMEC inhibits E. coli invasion significantly with a concomitant decrease in MLC phosphorylation. The inhibition of E. coli invasion by these PAK1 mutants is due to the absence of phospho-MLC at the actin condensation points. In contrast, the dominant-negative PAK1 shows no effect either on the invasion or on MLC phosphorylation or phospho-MLC recruitment to the actin focal points, suggesting that activated PAK1 inactivates MLCK. Taken together, these results suggest that E. coli invasion of HBMEC induces MLC phosphorylation by inhibiting the activity of PAK1 and the recruitment of phosphorylated MLC to the site of actin condensation beneath the bacteria for efficient internalization of E. coli into HBMEC.

* Corresponding author. Mailing address: Division of Infectious Diseases, MS #51, Childrens Hospital Los Angeles, 4650 Sunset Blvd., Los Angeles, CA 90027. Phone: (323) 669-5465. Fax: (323) 660-2661. E-mail: pnemani{at}chla.usc.edu.

Editor: J. N. Weiser

Infection and Immunity, May 2003, p. 2787-2797, Vol. 71, No. 5
0019-9567/03/$08.00+0     DOI: 10.1128/IAI.71.5.2787-2797.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Behling-Kelly, E., McClenahan, D., Kim, K. S., Czuprynski, C. J. (2007). Viable "Haemophilus somnus" Induces Myosin Light-Chain Kinase-Dependent Decrease in Brain Endothelial Cell Monolayer Resistance. Infect. Immun. 75: 4572-4581 [Abstract] [Full Text]  
  • Stockton, R. A., Schaefer, E., Schwartz, M. A. (2004). p21-activated Kinase Regulates Endothelial Permeability through Modulation of Contractility. J. Biol. Chem. 279: 46621-46630 [Abstract] [Full Text]  
  • Sukumaran, S. K., McNamara, G., Prasadarao, N. V. (2003). Escherichia coli K-1 Interaction with Human Brain Micro-vascular Endothelial Cells Triggers Phospholipase C-{gamma}1 Activation Downstream of Phosphatidylinositol 3-Kinase. J. Biol. Chem. 278: 45753-45762 [Abstract] [Full Text]  


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