Affiliation:
1. Department of Pathobiological Sciences, University of Wisconsin-Madison, Madison, Wisconsin
Abstract
ABSTRACT
Mannheimia haemolytica
is the principal bacterial pathogen of the bovine respiratory disease complex. Its most important virulence factor is a leukotoxin (LKT), which is a member of the RTX family of exotoxins produced by many gram-negative bacteria. Previous studies demonstrated that LKT binds to the β
2
-integrin LFA-1 (CD11a/CD18) on bovine leukocytes, resulting in cell death. In this study, we demonstrated that depletion of lipid rafts significantly decreases LKT-induced bovine lymphoblastoid cell (BL-3) death. After binding to BL-3 cells, some of the LKT relocated to lipid rafts in an LFA-1-independent manner. We hypothesized that after binding to LFA-1 on BL-3 cells, LKT moves to lipid rafts and clathrin-coated pits via a dynamic process that results in LKT internalization and cytotoxicity. Knocking down dynamin-2 by small interfering RNA reduced both LKT internalization and cytotoxicity. Similarly, expression of dominant negative Eps15 protein expression, which is required for clathrin coat formation, reduced LKT internalization and LKT-mediated cytotoxicity to BL-3 cells. Finally, we demonstrated that inhibiting actin polymerization reduced both LKT internalization and LKT-mediated cytotoxicity. These results suggest that both lipid rafts and clathrin-mediated mechanisms are important for LKT internalization and cytotoxicity in BL-3 cells and illustrate the complex nature of LKT internalization by the cytoskeletal network.
Publisher
American Society for Microbiology
Subject
Infectious Diseases,Immunology,Microbiology,Parasitology
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