Platelet Endothelial Cell Adhesion Molecule-1 (PECAM-1) Homophilic Adhesion Is Mediated by Immunoglobulin-like Domains 1 and 2 and Depends on the Cytoplasmic Domain and the Level of Surface Expression

Abstract

PECAM-1/CD31 is vascular cell adhesion and signaling molecule of the Ig superfamily that plays a role in neutrophil recruitment at inflammatory sites and may be involved the release of leukocytes from the bone marrow and in cardiovascular development. The interactions of PECAM-1 with its ligands are complex in that it is able to bind both with itself (homophilic adhesion) or with non-PECAM-1 ligands (heterophilic adhesion). Although the factors that regulate ligand binding are not fully understood, these interactions are regulated in part by its large cytoplasmic domain, a region of 118 amino acids encoded by 8 exons of its gene (exons 9 -16). The purpose of this work was to better define the mechanisms of PECAM-1-dependent homophilic adhesion by analyzing the binding interactions of L-cells expressing full-length and selectively mutated forms of human, murine, and human/murine chimeric PECAM-1 molecules in an established aggregation assay. These studies demonstrate that 1) the minimal length of the cytoplasmic domain required for cellular aggregation is represented within the sequences encoded by exons 9 and 10, 2) removal or addition of the sequences encoded by exon 14 from the cytoplasmic domain can determine whether the mechanism of aggregation is a heterophilic calciumdependent process or a homophilic calcium-independent process, 3) high levels of surface expression of PE-CAM-1 on the cell surface change the mechanism of aggregation from heterophilic to homophilic, and 4) PE-CAM-1-dependent homophilic binding appears to involve the direct interaction of only the first two extracellular Ig-like domains. These data suggest that PECAM-1-ligand interactions can be regulated through multiple pathways including alterations of the cytoplasmic domain and the level of surface expression.

Influence of PECAM-1 ligand interactions on PECAM-1-dependent cell motility and filopodia extension

“…1993, 1994; Sun et al. 1996a). In these so called “mixed aggregation studies”, homophilic interactions result in aggregates that are enriched with cells expressing the molecule of interest, whereas heterophilic binding (to other molecules on surface of the cell) are responsible for aggregates that are composed of both cell types.…”

“…PECAM‐1 within the membrane of the filopodia is not freely diffusible to the rest of the cell surface and thus its surface density in the filopodia is likely to be high, conditions that might favor these cis‐homophilic interactions (Sun et al. 1996a). Clustering of PECAM‐1 subsequently enables high‐affinity heterophilic binding to proteoglycans in the matrix that may be reminiscent of the affinity modulation observed with clustered integrins (Maheshwari et al.…”

“…1993, 1994; Sun et al. 1996a). Specifically for the 5 cell aggregates, aggregates were defined as homophilic if they contained 4 or 5 PECAM‐1‐expressing REN cells; heterophilic if 2 or 3 PECAM‐1‐expressing cells were present; or nonspecific if the aggregate was composed of one or no PECAM‐1‐expressing cell.…”

“…For homophilic interactions, molecules on adjacent cells overlap each other in an antiparallel manner in which Ig‐like domain 1 of one molecule binds to Ig‐like domain 2 of the counter molecule (Sun et al. 1996a, 1996b). In terms of factors that regulate the specificity of ligand interaction, a high surface density, antibody‐mediated engagement of the membrane proximal Ig‐like domain 6 and the absence of sequences from the cytoplasmic domain coded by exon 14 of the PECAM‐1 gene promote homophilic over GAG‐dependent heterophilic adhesion (Sun et al.…”

“…In terms of factors that regulate the specificity of ligand interaction, a high surface density, antibody‐mediated engagement of the membrane proximal Ig‐like domain 6 and the absence of sequences from the cytoplasmic domain coded by exon 14 of the PECAM‐1 gene promote homophilic over GAG‐dependent heterophilic adhesion (Sun et al. 1996a, 1996b). In addition, homophilic adhesion is dependent on the glycosylation state of the PECAM‐1 molecule (Lee et al.…”