A Novel Serine-rich Motif in the Intercellular Adhesion Molecule 3 Is Critical for Its Ezrin/Radixin/Moesin-directed Subcellular Targeting

Abstract

The relationship between adhesion receptors and the cytoskeleton is crucial for leukocyte migration and cell-cell interactions (1). In migrating leukocytes, ICAMs 1 are redistributed to the cellular uropod, a membrane protrusion at the rear of the cells (2). ICAMs act as both adhesion and signaling receptors and have partially overlapping functions (3). The ␤ 2 integrin LFA-1 specifically binds to the intercellular adhesion molecules ICAM-1, -2, and -3 (4 -6). ICAM-3 is highly expressed in naive T cells and interacts preferentially with two additional ligands, the integrin ␣ D ␤ 2 (7) and DC-SIGN, a novel C-type lectin expressed in dendritic cells that binds with high affinity to ICAM-3 (8). ICAM-3-DC-SIGN interaction seems to play a key role during the initiation of the immune response. ICAMs are co-localized with members of the ezrin, moesin, and radixin family of proteins in several cell types (9, 10). The latter proteins are thought to connect constituents of the plasma membrane with the cytoskeleton and are commonly associated with protrusive formations such as microvilli, microspikes, and filopodia (11,12). In leukocytes, ERM proteins are concentrated at the tips of microvilli (13,14), facilitating macrophage phagocytosis (15) and CD95-mediated apoptosis of T cells (16). Interactions between ICAM-1, ICAM-2, and moesin/ezrin/radixin have been studied previously in vitro (10, 17). The functional relevance of such interactions has been illustrated for natural killer cellmediated killing of target cells (18) and for microvillar organization during cortical morphogenesis (19).We have investigated here the dynamics of the interaction of ERM proteins with ICAM-3 and the critical residues in its cytoplasmic region, accounting both for binding to moesin and ezrin and for targeting of this adhesion receptor to the cellular uropod. EXPERIMENTAL PROCEDURESAntibodies, Cells, and Reagents-The anti-ICAM-3 HP2/19, TP1/24 and TP1/25; anti-CD4 HP2/6; anti-ICAM-1 RR1.1; anti-LFA-1␣ TS1/11; anti-CD45 D3/9; and anti-moesin/radixin 38/87 mAb have been described (2, 9, 20). The blocking anti-ICAM-3 140.11 mAb was generously provided by Dr. R. Vilella (Hospital Clinic, Barcelona, Spain). The anti-LFA-1␣ NKI-L16 mAb was generously provided by Dr. C. G. Figdor (University Hospital, Nijmegen, The Netherlands). The anti-ICAM-3 polyclonal antiserum (pAb) was generously provided by Dr. D. Simmons (Imperial Cancer Research Fund Laboratories, Oxford, United Kingdom). The anti-moesin 95/2 and the anti-ezrin 90/3 pAbs were raised in rabbits by immunization with recombinant human moesin and purified human ezrin, respectively, and the former was purified by affinity chromatography (13). The affinity-purified polyclonal antibody 454 raised against a unique peptide from mouse moesin was generously provided by Dr.

CD93 is required for maintenance of antibody secretion and persistence of plasma cells in the bone marrow niche

“…Moesin is known to be important in linking transmembrane proteins to the cytoskeleton. This contributes to a redistribution of the actin cytoskeleton that has been shown to be essential for phagocytosis, migration, and adhesion (33,34). As this molecule seems to be implicated in survival in BM niches, it makes sense that GC and memory B cells do not express this marker.…”

Activation of SRC Tyrosine Kinases in Response to ICAM-1 Ligation in Pulmonary Microvascular Endothelial Cells

“…25). Functioning as a linkage between membrane proteins and the submembrane F-actin cytoskeleton, ezrin plays important roles in organizing membrane structures such as microvilli and in targeting membrane proteins such ICAM-3 during lymphocyte migration (25,43). Although not well understood, the activity of ERM proteins appears to be regulated by phosphorylation at a conserved threonine residue at the C terminus and/or binding to phosphatidylinositol 4,5-bisphosphate.…”

SLIC‐1/sorting nexin 20: A novel sorting nexin that directs subcellular distribution of PSGL‐1

“…The crystal structure of p40phox bound to PtdIns(3)P has recently been solved and Arg58 in p40phox was shown to form the most extensive interaction with the bound phospholipid [45]. A single amino acid mutation to Gln at this position was reported to effect loss of phospholipid binding without affecting the overall fold of the protein.…”