Nonmuscle myosin heavy chain IIA mediates Epstein–Barr virus infection of nasopharyngeal epithelial cells

Abstract

EBV causes B lymphomas and undifferentiated nasopharyngeal carcinoma (NPC). Although the mechanisms by which EBV infects B lymphocytes have been extensively studied, investigation of the mechanisms by which EBV infects nasopharyngeal epithelial cells (NPECs) has only recently been enabled by the successful growth of B lymphoma Mo-MLV insertion region 1 homolog (BMI1)-immortalized NPECs in vitro and the discovery that neuropilin 1 expression positively affects EBV glycoprotein B (gB)-mediated infection and tyrosine kinase activations in enhancing EBV infection of BMI1-immortalized NPECs. We have now found that even though EBV infected NPECs grown as a monolayer at extremely low efficiency (<3%), close to 30% of NPECs grown as sphere-like cells (SLCs) were infected by EBV. We also identified nonmuscle myosin heavy chain IIA (NMHC-IIA) as another NPEC protein important for efficient EBV infection. EBV gH/gL specifically interacted with NMHC-IIA both in vitro and in vivo. NMHC-IIA densely aggregated on the surface of NPEC SLCs and colocalized with EBV. EBV infection of NPEC SLCs was significantly reduced by NMHC-IIA siRNA knock-down. NMHC-IIA antisera also efficiently blocked EBV infection. These data indicate that NMHC-IIA is an important factor for EBV NPEC infection. Epstein-Barr virusBV is a nearly ubiquitous human γ-herpesvirus that causes B-cell lymphomas and nasopharyngeal carcinoma (NPC), indicative of tropism for both cell types (1-3). Until recently, the molecular mechanisms of EBV infection of B lymphocytes were better understood than the mechanisms of epithelial cell infection (4). EBV attachment to the B-cell membrane is mediated by interactions between EBV glycoprotein 350 (gp350) and complement receptor type 2 (CR2 or CD21) (5) or CD35 (6). EBV gp42 binding to HLA class II triggers EBV fusion with B cells in the presence of EBV glycoprotein B (gB) and gH/gL (7,8). For epithelial cells, gH/gL and gB are important for EBV infection (4, 9, 10). Epithelial cells lack HLA class II expression; thus, gp42 cannot trigger EBV and cell fusion. Instead, gp42 inversely suppresses the infection (11), and an antibody against gp350 can enhance infections of CD21/CD35-negative epithelial cells (12). The gH/gL heterodimer is required for virus entry (4) and may be involved in binding (13), as well as fusion of EBV (14-17). However, the crystal structure of EBV gH/gL does not show any known fusion domain (18). It is now thought that gH/gL regulates the fusion function of gB (19). Binding of gH/gL to a subset of αv integrins (e.g., αvβ 5 , αvβ 6 , or αvβ 8 ) provides the initial trigger for gB-mediated fusion (16,20,21). However, E1D1(gH/gL) antibody or CL59(gH) antibody, with a different epitope, can impair epithelial cell infection (20,22). Thus, multiple gH/gL domains are critical to EBV infection, and gH/gL may interact with proteins in addition to integrins. Direct interaction of EBV gB amino acids 23-431 with neuropilin 1 (NRP1) and its associated tyrosine kinases is critical for EBV infection of nasopharyngeal ep...

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