Lymphocyte Function-Associated Antigen 1 Is a Receptor forPasteurella haemolyticaLeukotoxin in Bovine Leukocytes

Abstract

Pasteurella (Mannheimia) haemolytica leukotoxin (Lkt) causes cell type-and species-specific effects in ruminant leukocytes. Recent studies indicate that P. haemolytica Lkt binds to bovine CD18, the common subunit of all ␤2 integrins. We designed experiments with the following objectives: to identify which member of the ␤2 integrins is a receptor for Lkt; to determine whether Lkt binding to the receptor is target cell (bovine leukocytes) specific; to define the relationships between Lkt binding to the receptor, calcium elevation, and cytolysis; and to determine whether a correlation exists between Lkt receptor expression and the magnitude of target cell cytolysis. We compared Lkt-induced cytolysis in neutrophils from control calves and from calves with bovine leukocyte adhesion deficiency (BLAD), because neutrophils from BLAD-homozygous calves exhibit reduced ␤2 integrin expression. The results demonstrate for the first time that Lkt binds to bovine CD11a and CD18 (lymphocyte function-associated antigen 1 [LFA-1]). The binding was abolished by anti-CD11a or anti-CD18 monoclonal antibody (MAb). Lkt-induced calcium elevation in bovine alveolar macrophages (BAMs) was inhibited by anti-CD11a or anti-CD18 MAb (65 to 94% and 37 to 98%, respectively, at 5 and 50 Lkt units per ml; P < 0.05). Lkt-induced cytolysis in neutrophils and BAMs was also inhibited by anti-CD11a or anti-CD18 MAb in a concentration-dependent manner. Lkt bound to porcine LFA-1 but did not induce calcium elevation or cytolysis. In neutrophils from BLAD calves, Lkt-induced cytolysis was decreased by 44% compared to that of neutrophils from control calves (P < 0.05). These results indicate that LFA-1 is a Lkt receptor, Lkt binding to LFA-1 is not target cell specific, Lkt binding to bovine LFA-1 correlates with calcium elevation and cytolysis, and bovine LFA-1 expression correlates with the magnitude of Lkt-induced target cell cytolysis.Leukotoxin (Lkt) and lipopolysaccharide produced by Pasteurella (Mannheimia) haemolytica serotype 1 are considered to be the primary virulence factors contributing to lung injury in bovine pneumonic pasteurellosis (BPP) (33,36,38,40), a disease of substantial economic importance to the beef and dairy cattle industries in North America (7,28,39). Lkt is a member of a family of gram-negative bacterial exotoxins termed RTX (for repeats in toxin) cytolysins (3). Although most RTX cytolysins interact with a variety of cell types from many different species (6), cytolysins produced by Actinobacillus actinomycetemcomitans, Actinobacillus pleuropneumoniae (ApxIIIA), and P. haemolytica are known to have cell type-and species-specific effects. The leukotoxin (LtxA) of A. actinomycetemcomitans, a human pathogen, interacts only with cells of the lymphocytic and monomyelocytic lineages of humans and some nonhuman primates (23); the Lkt of P. haemolytica, a ruminant pathogen, interacts only with ruminant leukocytes causing activation and cytolysis (4,15,25,33,40). A study by Lally et al. (23) has determined that two RTX cytolysins, ...

Phylogenetic relationship of equine Actinobacillus species and distribution of RTX toxin genes among clusters

“…Differences in the toxic activities are most probably due to specific binding of the toxin to receptors on the target cells and/or differences in the signaling cascade after toxin binding. This was shown for other RTX toxins like the leukotoxin Lkt of Mannheimia haemolytica which after binding to the LFA-1 receptor is toxic for bovine leukocytes but has no effect on porcine or equine lymphocytes [10,11,19].…”

How Mannheimia haemolytica defeats host defence through a kiss of death mechanism

“…Indeed, inactivation of M. haemolytica leukotoxin by a gene knockout hardly causes any further pulmonary lesions although the wild-type and mutant strains were equally capable of colonising the upper respiratory tracts of the calves [165]. Moreover, necrosis of neutrophils can be reproduced in vitro with purified LKT [5,34,40,46,88,89,161,162,176]. Biologically, it is worth noting that even if LKT is able to bind leukocytes from various animal species, it is only cytotoxic for ruminant leukocytes, suggesting that the interaction specificity between LKT and ruminant leukocytes could be responsible for the ruminant-specificity of M. haemolytica [92,149,161].…”

“…These data suggest that the binding of LKT on ruminant LFA-1 is liable for the virulence specificity of M. haemolytica against ruminants. The precise identification of the subunit that binds LKT appears controversed [88], even if CD18 seems probable [5,46,109]. Fortunately, the recent cloning, sequencing and characterisation of the Bos taurus CD11a [56] will give the first opportunity to express homologous and heterologous LFA-1 in vitro to definitely answer the question.…”

Mannheimia haemolytica

“…34,35 This species specificity stems from the selective interaction of LKT with the b 2 integrin LFA-1 (lymphocyte function-associated antigen 1; CD11a/CD18) on target host cells. 35,40,48,59,71,77,78 Integrins are expressed on most cell types and are involved in cell-cell interactions and cell-extracellular matrix interactions, whereas subtype b 2 integrins are expressed exclusively on leukocytes, including T lymphocytes, macrophages, neutrophils, monocytes, and dendritic cells. 6 Despite a consensus that LKT-induced specificity is due to b 2 integrin binding, a complete agreement is lacking on the specific b 2 integrin and subunits of b 2 integrin required for receptor-ligand interaction.…”