EBV attachment stimulates FHOS/FHOD1 redistribution and co-aggregation with CD21: formin interactions with the cytoplasmic domain of human CD21

Abstract

CD21 is a multifunctional receptor for Epstein-Barr virus (EBV), for C3dg and for CD23. Upon engagement of immune complexes CD21 modulates immunoreceptor signaling, linking innate and adaptive immune responses. The mechanisms enabling CD21 to independently relay information between the exterior and interior of the cell, however, remain unresolved. We show that formin homologue overexpressed in spleen (FHOS/FHOD1) binds the cytoplasmic domain of human CD21 through its C terminus. When expressed in cells, EGFP-FHOS localizes to the cytoplasm and accumulates with actin in membrane protrusions. Plasma membrane aggregation, redistribution and co-localization of both proteins are stimulated when EBV (ligand) binds CD21. Though widely expressed, FHOS RNA is most abundant in the littoral cell, a major constituent of the red pulp of human spleen believed to function in antigen filtration. Formins are molecular scaffolds that nucleate actin by a pathway distinct from Arp2/3 complex, linking signal transduction to actin reorganization and gene transcription. Thus, ligand stimulation of FHOS-CD21 interaction may transmit signals through promotion of cytoskeletal rearrangement. Moreover, formin recruitment to sites of actin assembly initiated by immunoreceptors could be a general mechanism whereby co-receptors such as CD21 modulate intracellular signaling.

Important but Differential Roles for Actin in Trafficking of Epstein-Barr Virus in B Cells and Epithelial Cells

“…Binding of virus cross-links CR2 and results in colocalization with the formin FHOS/FHOD1, which binds to the cytoplasmic domain of CR2 (15). Formins directly nucleate actin (17), and since actin dynamics seemed to be uniquely important to virus transport in epithelial cells, the possibility that CR2 might provide a link to cytoskeletal reorganization was very attractive.…”

Resting B cells as a transfer vehicle for Epstein–Barr virus infection of epithelial cells

“…We noted that virus-loaded B cells become transfer-competent at the same time that they acquire the ability to form conjugates with epithelial cells, suggesting that the two activities are related; indeed, both may be acquired as a consequence of EBV-induced cross-linking of the CD21͞CD19͞ CD81 receptor complex. Thus, CD21 cross-linking is known to have several downstream signaling effects in resting B cells (22)(23)(24) and, in a model system, leads to cytoskeletal reorganization (25). Certainly, EBV binding induced cocapping of the gp350͞receptor complex to one pole of the cell, and, after coculture with epithelium, that pole formed the point of intercellular contact.…”

Functional Divergence of Kaposi's Sarcoma-Associated Herpesvirus and Related Gamma-2 Herpesvirus Thymidine Kinases: Novel Cytoplasmic Phosphoproteins That Alter Cellular Morphology and Disrupt Adhesion

“…After being extensively washed (with PBS), cells were incubated with either goat anti-mouse phycoerythrin (Biosource International AMI4407) or goat anti-rabbit phycoerythrin (Biosource International ALI4407) conjugate at a 1:1,000 dilution in PBS for 30 min at RT. The details required for actin staining and cell mounting were described previously (18). Immunofluorescence images were taken under an 100ϫ oil objective on a Nikon E600 microscope with a SPOT2 digital camera with SPOTcam software, version 3.5.5.…”

“…Cell lysates were prepared from transfected adherent cells as previously described (18). Briefly, cells were collected by the addition of preboiled lysis buffer (50 mM Tris-1% sodium dodecyl sulfate [SDS]-0.5% ␤-mercaptoethanol), sonicated, and boiled for 5 min; protein concentration was quantified.…”