Human herpesvirus 6 (HHV-6) glycoproteins H and L (gH and gL, respectively) and the 80-kDa form of glycoprotein Q (gQ-80K) form a heterotrimeric complex that is found on the viral envelope and that is a viral ligand for human CD46. Besides gQ-80K, the gQ gene encodes an additional product whose mature molecular mass is 37 kDa (gQ-37K) and which is derived from a different transcript. Therefore, we designated gQ-80K as gQ1 and gQ-37K as gQ2. We show here that gQ2 also interacts with the gH-gL-gQ1 complex in HHV-6-infected cells and in virions. To examine how these components interact in HHV-6-infected cells, we performed pulse-chase studies. The results demonstrated that gQ2-34K, which is endo-␤-N-acetylglucosaminidase H sensitive and which is the precursor form of gQ2-37K, associates with gQ1-74K, which is the precursor form of gQ1-80K, within 30 min of the pulse period. After a 1-h chase, these precursor forms had associated with the gH-gL dimer. Interestingly, an anti-gH monoclonal antibody coimmunoprecipitated mainly gQ1-80K and gQ2-37K, with little gQ1-74K or gQ2-34K. These results indicate that although gQ2-34K and gQ1-74K interact in the endoplasmic reticulum, the gH-gL-gQ1-80K-gQ2-37K heterotetrameric complex arises in the postendoplasmic reticulum compartment. The mature complex is subsequently incorporated into viral particles.Human herpesvirus 6 (HHV-6) is a betaherpesvirus related to human herpesvirus 7 (HHV-7) and human cytomegalovirus (HCMV) and is a human pathogen of emerging clinical significance. HHV-6 was first isolated from the peripheral blood lymphocytes of patients with lymphoproliferative disorders and AIDS (34). HHV-6 isolates can be categorized as two variants, A (HHV-6A) and B (HHV-6B), on the basis of their in vitro growth properties, DNA restriction site polymorphisms, antigenicity, and host cell tropism (1,(3)(4)(5)45). HHV-6B is the causative agent of exanthem subitum (46).Herpesviruses encode a number of glycoproteins that are present in the envelope of the virion and that play an important role in viral infection, including attachment, penetration, cell-cell spread, and the envelopment and maturation of nascent viral particles. A number of studies have reported a role for glycoprotein H (gH) and glycoprotein L (gL) in the membrane fusion events involved in herpesvirus entry and cell-cell spread. Monoclonal antibodies (MAbs) raised to gH neutralize the activity of infectious virus (6,10,12,13,22,23,37). Similarly, antibodies against gL can inhibit virus infectivity and cell fusion (19,29,30).Previous studies showed that gH and gL form a heteromeric glycoprotein complex (11,17,18,22,33,47). Intermolecular disulfide bridges are required for complex formation in betaherpesviruses HCMV and HHV-6 (2, 18, 38) but not in alphaherpesviruses herpes simplex virus type 1, varicella-zoster virus, and bovine herpesvirus 1 (8,9,17,42). The Epstein-Barr virus (EBV) gH-gL complex includes a third glycoprotein, called gp42, which is encoded by the BZLF2 open reading frame (ORF) (21). The gCIII compl...

Human CD46 is a cellular receptor for human herpesvirus 6 (HHV-6). Virus entry into host cells requires a glycoprotein H (gH)-glycoprotein L (gL) complex. We show that the CD46 ectodomain blocked HHV-6 infection and bound a complex of gH-gL and the 80-kDa U100 gene product, designated glycoprotein Q, indicating that the complex is a viral ligand for CD46.

Human herpesvirus 6 (HHV-6) is a lymphotropic betaherpesvirus that productively infects T cells and monocytes. HHV-6 isolates can be differentiated into two groups, variants A and B (HHV-6A and HHV-6B). Here, we show a functional difference between HHV-6A and -6B in that HHV-6A induced syncytium formation of diverse human cells but HHV-6B did not. The syncytium formation induced by HHV-6A was observed 2 h after infection; moreover, it was found in the presence of cycloheximide, indicating that HHV-6A induced fusion from without (FFWO) in the target cells. Furthermore, the fusion event was dependent on the expression of the HHV-6 entry receptor, CD46, on the target cell membrane. In addition, we determined that short consensus repeat 2 (SCR2), -3, and -4 of the CD46 ectodomain were essential for the formation of the virus-induced syncytia. Monoclonal antibodies against glycoproteins B and H of HHV-6A inhibited the fusion event, indicating that the syncytium formation induced by HHV-6A required glycoproteins H and B. These findings suggest that FFWO, which HHV-6A induced in a variety of cell lines, may play an important role in the pathogenesis of HHV-6A, not only in lymphocytes but also in various tissues, because CD46 is expressed ubiquitously in human tissues.

. Here, we show that the HHV-6 U47 gene, which is a positional homolog of the human cytomegalovirus glycoprotein O (gO) gene, encodes a third component of the HHV-6 gH-gLcontaining envelope complex. A monoclonal antibody (MAb) against the amino terminus of HHV-6 gO reacted in immunoblots with protein species migrating at 120 to 130 kDa and 74 to 80 kDa in lysates of HHV-6-infected cells and with a 74-to 80-kDa protein species in purified virions. The 80-kDa form of gO was coimmunoprecipitated with an anti-gH MAb, but an anti-gQ MAb, which coimmunoprecipitated gH, did not coprecipitate gO. Furthermore, the gH-gL-gO complex did not bind to human CD46, indicating that the complex was not a ligand for CD46. These findings suggested that the viral envelope contains at least two kinds of tripartite complexes, gH-gL-gQ and gH-gL-gO, and that the gH-gL-gO complex may play a role different from that of gH-gL-gQ during viral infection. This is the first report of two kinds of gH-gL complexes on the viral envelope in a member of the herpesvirus family.Human herpesvirus 6 (HHV-6) was first isolated from the peripheral blood of patients with AIDS and lymphoproliferative disorders (8, 28). HHV-6 isolates can be classified into two variants, HHV-6A and HHV-6B. HHV-6B is the causative agent of exanthem subitum (40). The classification of the two variants is based on nucleotide sequence differences, as well as their immunological and biological characteristics (1-3, 6, 37).In the herpesvirus family, the envelope glycoproteins play critical roles in viral infection, including attachment, penetration, cell-to-cell spread, and the envelopment and maturation of nascent viral particles. In all of the human and animal herpesviruses studied to date, homologs of glycoprotein H (gH) and glycoprotein L (gL) have been found (9, 14-17, 20, 21, 26, 31-33, 38, 39, 41, 42). These two envelope glycoproteins, which associate to form a gH-gL complex, have been implicated as key participants in fusion events that are critical to herpesvirus infection. Studies of the HHV-6 gH (14, 27) and gL proteins have shown them to be representative gH and gL homologs (19,20).Santoro et al. showed that human CD46 is a cellular receptor for HHV-6 (30). Recently, we showed that HHV-6A, but not HHV-6B, mediates fusion from without in a variety of cells expressing human CD46 (23) and that anti-gH and anti-gB monoclonal antibodies (MAbs) inhibit the cell-cell fusion induced by HHV-6A. Furthermore, we found that the HHV-6A gH-gL complex interacts with one form of the U100 gene products, which we designated glycoprotein Q (gQ) (22), and we identified the gH-gL-gQ complex of HHV-6A as the viral ligand for human CD46 (25). Santoro et al. have also reported that HHV-6 gH associates with CD46 by a coimmunoprecipitation study (29).In the case of Epstein-Barr virus (EBV), a third viral glycoprotein, gp42, associates with the gH-gL complex (18,35,36). Recently, a third viral gene product of human cytomegalovirus (HCMV), glycoprotein O (gO), was identified as a member of the...

The characterization is reported of the human herpesvirus-6B (HHV-6B) rep/U94 gene, which is a homologue of the adeno-associated virus type 2 rep. In this study, a monoclonal antibody was produced against HHV-6B REP (anti-REP mAb). Immunofluorescence staining using the anti-REP mAb showed that REP was localized to the nucleus in HHV-6-infected MT4 cells. It was first detected at 24 h post-infection (p.i.) and accumulated to higher levels by 72 h p.i. REP may be expressed only at very low levels in HHV-6-infected cells : even when the late protein glycoprotein H was detected in nearly 90 % of HHV-6-infected cells, REP was detected in only a small percentage of them. Western blot analysis showed that the anti-REP mAb recognized a 56-kDa polypeptide in HHV-6B-infected MT4 cells. Furthermore, the REP protein was shown to bind single-stranded DNA.