Contributions of herpes simplex virus type 1 envelope proteins to entry by endocytosis

Tri Komala Sari

Herpes simplex virus (HSV) can transit at least three distinct cellular routes during viral entry. These pathways include: endocytosis (either pH-dependent or - independent) and direct penetration at the cell surface (pH-independent) (Fig. 1). In contrast to other human herpesviruses, such as EBV and HCMV, HSV envelope proteins that are specifically required for one pathway but not another have not been identified. HSV envelope proteins glycoprotein B (gB), gD, gH, and gL are essential for entry regardless of entry pathway. HSVs deleted for gE, gG, gI, gJ, gK, gM, UL20, UL45 or Us9 replicate to varying extents in Vero cells, which support non-endocytic entry. We used individual gene deletion mutants of HSV type 1 (HSV-1) in an analysis of the roles of gE, gG, gI, gJ, gK, gM, UL20, UL45 and Us9 in the endocytic entry of HSV-1. To measure entry and infectivity of the HSV deletion mutants on cells that support pHindependent endocytic entry (B78-nectin-1), we used a beta-galactosidase reporter assay and a plaque assay, respectively. The results indicated that gE, gG, gI, gJ, gK, gM, UL20, UL45 and Us9 are dispensable for pH-independent endocytic entry. Purified HSV virions are irreversibly inactivated for entry by in vitro treatment with low pH, a hallmark of viruses that enter via pH-activated membrane fusion mechanisms. The target(s) of inactivation in the HSV envelope is not known. Entry of each of the mutant viruses was inactivated by pHs of 5.0 or 6.0 to an extent similar to wild type HSV. Each mutant virus tested retained the ability to enter CHO-nectin-1 cells by pH-dependent endocytosis. Also, entry of each of the mutant HSVs into CHO-nectin-1 cells was blocked by the pHaltering agent ammonium chloride. Together the results suggest that gE, gG, gI, gJ, gM, UL45 and Us9 are non-essential for entry by either form of endocytosis. Interestingly, we found that entry of gK- and UL20-null mutant HSVs was defective in CHO-nectin-1 cells under the conditions tested, suggesting gK and UL20 may potentially have roles in pHdependent entry at low multiplicity of infection.

Contributions of herpes simplex virus type 1 envelope proteins to entry by endocytosis

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