Glycoprotein D of herpes simplex virus encodes a domain which precludes penetration of cells expressing the glycoprotein by superinfecting herpes simplex virus

Abstract

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-l 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.

Herpes simplex virus type 1 glycoprotein K is not essential for infectious virus production in actively replicating cells but is required for efficient envelopment and translocation of infectious virions from the cytoplasm to the extracellular space

“…Infectious ⌬gK virions accumulated within the cytoplasm of Vero and HEp-2 cells, indicating that gK plays an important role in release of cytoplasmic virions to the extracellular spaces. A similar retention of infectious virions within cells was noted when Golgi-specified enzymatic functions were inhibited either by Golgi-specific inhibitors or in mutant cells deficient in specific Golgi-localized enzymes, indicating that the Golgi apparatus is important in virion egress (7,16,31). Previous studies indicated that gK contains high-mannose carbohydrate moieties which are characteristic of the RER, and indirect immunofluorescence with antipeptide antibodies failed to detect gK in the cytoplasm of infected cells (28,29).…”

“…An alternative model that involves de-envelopment at the outer nuclear membrane followed by envelopment at the rough ER (RER) and/or the Golgi complex has also been proposed (39,40,46,53,56). Golgi-dependent glycoprotein maturation is vital for cellular egress, because in cells defective in the processing of high-mannose precursor glycoproteins to mature glycoproteins, or when Golgi-dependent functions are inhibited by the ionophore monensin, infectious virions accumulate in the cytoplasm but are not transported into the extracellular space (7,16,31). HSV specifies at least three viral proteins which are involved in the transport of enveloped virions to extracellular spaces.…”

Viral determinants of the variable sensitivity of herpes simplex virus strains to gD-mediated interference

“…The mechanism of gD-mediated interference is not understood. It has been proposed that cell-associated gD interferes with a cell surface component required for viral entry or interacts with the virion itself, perhaps to disrupt interactions among the viral glycoproteins (3,4,8,11). Recently it was shown that a soluble recombinant form of gD becomes modified by the addition of mannose-6-phosphate (Man-6-P) to N-linked glycans and that this form can bind to Man-6-P receptors (2).…”

Characterization of a BHK(TK-) cell clone resistant to postattachment entry by herpes simplex virus types 1 and 2

“…Other cell lines resistant to virus entry have been used for the isolation of escape mutants that can grow more efficiently than wild-type virus on the resistant cells. Several groups have reported the properties of viruses carrying mutations in the gene encoding gD that grow more efficiently than the wild-type virus on cells that express gD or on a cell line that expresses the HSV-1 U s 11 gene product (2,7,8,34). If the mechanism of resistance to infection shown by 95-19 cells is essentially the same as that seen in these other resistant cell types, these mutant viruses should show a similarly enhanced ability to infect 95-19 cells.…”

“…Several groups have isolated lines that express gD from HSV-1 or HSV-2 and are resistant to HSV infection (6,23). These cells have been used for the selection of escape mutants of HSV-1 that can grow more efficiently than wild-type, and which map in the gD gene, suggesting that these cells fail to support an essential interaction between the cell surface and gD (2,(6)(7)(8).…”