LL-37, a cationic antimicrobial peptide, has numerous immune-modulating effects. However, the identity of a receptor that mediates the responses in immune cells remains uncertain. We have recently demonstrated that LL-37 interacts with the αMI-domain of integrin α_Mβ₂ (Mac-1), a major receptor on the surface of myeloid cells, and induces a migratory response in Mac-1-expressing monocyte/macrophages as well as activation of Mac-1 on neutrophils. Here, we show that LL-37 and its C-terminal derivative supported strong adhesion of various Mac-1-expressing cells, including human embryonic kidney cells stably transfected with Mac-1, human U937 monocytic cells, and murine IC-21 macrophages. The cell adhesion to LL-37 was partially inhibited by specific Mac-1 antagonists, including monoclonal antibody against the α_M integrin subunit and neutrophil inhibitory factor, and completely blocked when anti-Mac-1 antibodies were combined with heparin, suggesting that cell surface heparan sulfate proteoglycans act cooperatively with integrin Mac-1. Coating both gram-negative and gram-positive bacteria with LL-37 significantly potentiated their phagocytosis by macrophages, and this process was blocked by a combination of anti-Mac-1 monoclonal antibody and heparin. Furthermore, phagocytosis by wild-type murine peritoneal macrophages of LL-37-coated latex beads, a model of foreign surfaces, was several fold higher than that of untreated beads. In contrast, LL-37 failed to augment phagocytosis of beads by Mac-1-deficient macrophages. These results identify LL-37 as a novel ligand for integrin Mac-1 and demonstrate that the interaction between Mac-1 on macrophages and bacteria-bound LL-37 promotes phagocytosis.

The broad recognition specificity exhibited by integrin α_Mβ₂ (Mac-1, CD11b/CD18) has allowed this adhesion receptor to play innumerable roles in leukocyte biology, yet we know little about how and why α_Mβ₂ binds its multiple ligands. Within α_Mβ₂, the α_MI-domain is responsible for integrin’s multiligand binding properties. To identify its recognition motif, we screened peptide libraries spanning sequences of many known protein ligands for α_MI-domain binding and also selected the α_M I-domain recognition sequences by phage display. Analyses of >1400 binding and nonbinding peptides derived from peptide libraries showed that a key feature of the α_MI-domain recognition motif is a small core consisting of basic amino acids flanked by hydrophobic residues. Furthermore, the peptides selected by phage display conformed to a similar pattern. Identification of the recognition motif allowed the construction of an algorithm that reliably predicts the α_MI-domain binding sites in the α_Mβ₂ ligands. The recognition specificity of the α_MI-domain resembles that of some chaperones, which allows it to bind segments exposed in unfolded proteins. The disclosure of the α_Mβ₂ binding preferences allowed the prediction that cationic host defense peptides, which are strikingly enriched in the α_MI-domain recognition motifs, represent a new class of α_Mβ₂ ligands. This prediction has been tested by examining the interaction of α_Mβ₂ with the human cathelicidin peptide LL-37. LL-37 induced a potent α_Mβ₂-dependent cell migratory response and caused activation of α_Mβ₂ on neutrophils. The newly revealed recognition specificity of α_Mβ₂ toward unfolded protein segments and cationic proteins and peptides suggests that α_Mβ₂ may serve as a previously proposed “alarmin” receptor with important roles in innate host defense.