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A monoclonal antibody, designated as MAb 6E2, specific for human herpesvirus 6 variant B (HHV-6B) was derived from the spleen of a mouse immunized with lysates of HHV-6B(Z29) cord blood mononuclear cells. MAb 6E2 reacts by immunofluorescence with all the HHV-6B strains tested (Z29, CV, Hashimoto and SF) and fails to react with variant A prototypes, GS and U1102. The immunofluorescence staining was punctate and localized to the cytoplasm. The protein reacting with MAb 6E2 was identified as protein 48 000 in apparent Mr value by immunoaffinity chromatography of lysates of HHV-6B-infected mononuclear cells. © 1995.

Cell lines that constitutively express herpes simplex virus 1 (HSV-1) glycoprotein H (gH-1 ) failed to synthesize the mature form of gH and accumulated a precursor-like form of the glycoprotein, which was retained intracellularly, most likely in RER. Fine-structure analysis of the oligosaccharides present in recombinant gH revealed oligosaccharides processed by RER enzymes; sialylated complex-type and biantennary oligosaccharides, which are assembled in the trans-Golgi, were absent. A small fraction had the characteristics of oligosaccharides processed by the early mannosidases of the Golgi. These findings suggest that a defect in the transport out of RER to the Golgi may account for the intracellular retention of the immature form of gH in cells that express the glycoprotein constitutively. Upon superinfection of cells expressing gH-1 with HSV-2, recombinant gH-1 underwent maturation, indicating that a viral function is required to attain full processing of gH. The known HSV glycoproteins do not appear to carry out this function, since in cells infected with deletion mutants in gD, gG, gE, and gE-gl, with a spontaneous gC- mutant, or with a temperature-sensitive mutant in gB, maturation of gH occurred independently of the presence or of the maturation of the single glycoproteins tested. The present findings together with previous observations on HSV, human CMV, and the EBV homologue of gH suggest that inability of gH to undergo full processing in the absence of viral protein (s) is a property of gH.

Hyperimmune rabbit and mouse sera raised to human herpesvirus 7 (HHV-7)-infected cells and an immune human serum identified 20 [35S]methionine-[35S]cysteine-labelled proteins specific for HHV-7-infected cord blood mononuclear cells, ranging in apparent M(r) from 136K to 30K. The major proteins had apparent M(r) values of 121K, 100K, 87K, 85K, 60K, 51K, 46K, 42K, 40K and 36K. The human serum also identified seven [3H]glucosamine-labelled glycoproteins, with apparent M(r) values of 100K, 89K, 82K, 67K, 63K, 53K and 41K. Four monoclonal antibodies (MAbs) specific for HHV7-infected cells were derived. Two reacted with a family of five antigenically related polypeptides (87K, 85K, 70K, 61K and 57K in apparent M(r)), designated as the p85 complex. Two reacted with 121K and 51K M(r) proteins designated as p121 and p51, respectively. Human sera react with high frequency with the p85 complex and to a lesser extent with p121; hence these two proteins appear to be immunodominant for both humans and laboratory animals. The hyperimmune mouse serum and some of the MAbs showed some cross-reactivity with HHV-6A(U1102)- and 6B(Z29)-infected cells. The implications of cross-reactivity with respect to the human immune response to HHV-6 and -7 infections and prevalence analyses are discussed.

Pulse-chase experiments in conjunction with quantitative immunoprecipitation have been used to study the time-course of conversion from precursor to mature form of herpes simplex virus 1 glycoproteins C, D and B (gC, gD, and gB). The experimental systems employed were two infected cell lines and cells that constitutively express gD or gB. The relative rates of conversion among the glycoproteins did not vary in the systems used; the rate of maturation of gC was about two-fold higher than that of gD which, in turn, was about one and a half-fold higher than that of gB. Treatment with phosphonoacetate which inhibits viral DNA synthesis and hence virion morphogenesis induced a striking increase in the time course of conversion of immature gC, gD, and gB to fully glycosylated forms when measured late in the infection. The model of HSV glycoproteins maturation as integral components of the virion envelope is discussed. © 1988.

Earlier studies have shown that herpes simplex viruses adsorb to but do not penetrate permissive baby hamster kidney clonal cell lines designated the BJ series and constitutively expressing the herpes simplex virus 1 (HSV-1) glycoprotein D (gD). To investigate the mechanism of the restriction, the following steps were done. First, wild-type HSV-1 strain F [HSV-1(F)] virus was passaged blindly serially on clonal line BJ-1 and mutant viruses [HSV-1(F)U] capable of penetration were selected. The DNA fragment capable of transferring the capacity to infect BJ cells by marker transfer contains the gD gene. The mutant gD, designated gDU, differed from wild-type gD only in the substitution of Leu-25 by proline. gDU reacted with monoclonal antibodies which neutralize virus and whose epitopes encompass known functional domains involved in virus entry into cells. It did not react with the monoclonal antibody AP7 previously shown to react with an epitope which includes Leu-25. Second, cell lines expressing gDU constitutively were constructed and cloned. Unlike the clonal cell lines constitutively expressing gD (e.g., the BJ cell line), those expressing gDU were infectable by both HSV-1(F) and HSV-1(F)U. Lastly, exposure of BJ cells to monoclonal antibody AP7 rendered the cells capable of being infected with HSV-1(F). The results indicate that (i) gD expresses a specific function, determined by sequences at or around Leu-25, which blocks entry of virus into cells synthesizing gD, (ii) the gD which blocks penetration by superinfecting virus is located in the plasma membrane, (iii) the target of the restriction to penetration is the identical domain of the gD molecule contained in the envelope of the superinfecting virus, and (iv) the molecular basis of the restriction does not involve competition for a host protein involved in entry, as was previously thought.