Pritchard, S. M. | bioRxiv Science

bioRxiv2020

Herpes simplex virus entry by a non-conventional endocytic pathway

Herpes simplex virus 1 (HSV-1) causes significant morbidity and mortality in humans worldwide. HSV-1 enters epithelial cells via an endocytosis mechanism that is low pH-dependent. However, the precise intracellular pathway has not been identified, including the compartment where fusion occurs. In this study, we utilized a combination of molecular and pharmacological approaches to better characterize HSV entry by endocytosis. HSV-1 entry was unaltered in both cells treated with siRNA to Rab5 or Rab7 and cells expressing dominant-negative forms of these GTPases, suggesting entry is independent of the conventional endo-lysosomal network. The fungal metabolite brefeldin A (BFA) and the quinoline compound Golgicide A (GCA) inhibited HSV-1 entry via beta-galactosidase reporter assay and impaired incoming virus transport to the nuclear periphery, suggesting a role for trans Golgi network (TGN) functions and retrograde transport in HSV entry. Silencing of Rab9 or Rab11 GTPases, which are involved in the retrograde transport pathway, resulted in only a slight reduction in HSV infection. Together these results suggest that HSV enters host cells by an intracellular route independent of the lysosome-terminal endocytic pathway. IMPORTANCEHSV-1, the prototype alphaherpesvirus, is ubiquitous in the human population and causes lifelong infection that can be fatal in neonatal and immunocompromised individuals. HSV enters many cell types by endocytosis, including epithelial cells, the site of primary infection in the host. The intracellular itinerary for HSV entry remains unclear. We probed the potential involvement of several Rab GTPases in HSV-1 entry, and suggest that endocytic entry of HSV-1 is independent of the canonical lysosome-terminal pathway. A non-traditional endocytic route may be employed, such as one that intersects with the TGN. These results may lead to novel targets for intervention.

microbiology↗

bioRxiv2019

Herpes simplex virus glycoprotein C regulates low pH entry

Herpes simplex viruses (HSVs) cause significant morbidity and mortality in humans worldwide. Herpesviruses mediate entry by a multi-component, virus-encoded machinery. Herpesviruses enter cells by endosomal low pH and pH-neutral mechanisms in a cell-specific manner. HSV mediates cell entry via envelope glycoproteins gB, gD, and the heterodimer gH/gL regardless of pH or endocytosis requirements. HSV envelope proteins that function selectively in a given entry pathway have been elusive. Here we demonstrate that gC regulates cell entry and infection by a low pH pathway. Conformational changes in the core herpesviral fusogen gB are critical for membrane fusion. The presence of gC conferred a higher pH threshold to acid-induced antigenic changes in gB. Thus, gC may selectively facilitate low pH entry by regulating conformational changes in the fusion protein gB. We propose that gC modulates the HSV fusion machinery during entry into pathophysiologically relevant cells, such as human epidermal keratinocytes. ImportanceHerpesviruses are ubiquitous pathogens that cause lifelong latent infections and are characterized by multiple entry pathways. We propose that herpes simplex virus (HSV) gC plays a selective role in modulating HSV entry by a low pH pathway, such as into epithelial cells. gC facilitates conformational change of the main fusogen gB, a class III fusion protein. We propose a model whereby gC functions with gB, gD, and gH/gL to allow low pH entry. In the absence of gC, HSV entry occurs at a lower pH, coincident with trafficking to a lower pH compartment where gB changes occur at more acidic pHs. This study identifies a new function for gC and provides novel insight into the complex mechanism of HSV entry and fusion.

microbiology↗