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J. Biol. Chem., Vol. 280, Issue 6, 4864-4872, February 11, 2005

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Pseudomonas Invasion of Type I Pneumocytes Is Dependent on the Expression and Phosphorylation of Caveolin-2^*

David W. Zaas, Mathew J. Duncan, Guojie Li¶, Jo Rae Wright||, and Soman N. Abraham¶**

From the Division of Pulmonary, Allergy, and Critical Care Medicine, Department of Medicine, the Department of Molecular Genetics and Microbiology, the ^||Department of Cell Biology, the ^**Department of Immunology, and the ^¶Department of Pathology, Duke University Medical Center, Durham, North Carolina 27710

Pseudomonas aeruginosa is a major cause of pneumonia in patients with cystic fibrosis and other immuncompromising conditions. Here we showed that P. aeruginosa invades type I pneumocytes via a lipid raft-mediated mechanism. P. aeruginosa invasion of rat primary type I-like pneumocytes as well as a murine lung epithelial cell line 12 (MLE-12) is inhibited by drugs that remove membrane cholesterol and disrupt lipid rafts. Confocal microscopy demonstrated co-localization of intracellular P. aeruginosa with lipid raft components including caveolin-1 and -2. We generated caveolin-1 and -2 knockdowns in MLE-12 cells by using RNA interference techniques. Decreased expression of caveolin-2 significantly impaired the ability of P. aeruginosa to invade MLE-12 cells. In addition, the lipid raft-dependent tyrosine phosphorylation of caveolin-2 appeared to be a critical regulator of P. aeruginosa invasion.

Received for publication, October 14, 2004 , and in revised form, November 12, 2004.

^* This work was supported by National Institute of Health Grants 1F32-DK065412, 5R01 AI150021-03, 1 R21 AI056101-01A1, and 5 R01 HL68072 and by a Senior Investigator award from the Sandler Family Foundation on Asthma Research. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

To whom correspondence should be addressed: Duke University Medical Center, Box 3020, Duke University Medical Center, Durham, NC 27710. Tel.: 919684-6942; Fax: 919-684-2021; E-mail: soman.abraham{at}duke.edu.

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