Postentry Events Are Responsible for Restriction of Productive Varicella-Zoster Virus Infection in Chinese Hamster Ovary Cells

Abstract

Varicella-zoster virus (VZV) is a human alphaherpesvirus that causes chicken pox (varicella) upon primary infection and shingles (zoster) upon reactivation of the virus from latently infected sensory neurons. In contrast to the broad host range of most alphaherpesviruses in cultured cells, productive VZV infection in vitro is restricted almost exclusively to cells derived from humans and other primates (64). Reported exceptions are guinea pig embryonic fibroblasts (62, 64), rabbit kidney cells (64,68), and cotton rat fibroblasts (54). For many viruses, restriction in cultured cells is determined by specific cellular receptors that must be engaged for virus binding and entry. After successfully entering a cell, viruses can encounter other blocks to productive infection (20,36). In studies using rats, cotton rats, or mice as in vivo models of VZV latency, viral DNA (1,6,27,[51][52][53][54]69), viral transcripts (6,27,51,54,69), and immediate early viral proteins (18,27,51) were detected in infected cells, primarily neurons, indicating that VZV is capable of entering rodent cells and initiating early events in the infectious cycle. Cultured rat neurons and the mouse neuroblastoma cell line neuro-2A also permitted VZV entry, but productive infection was blocked in both cases (4, 37). Thus, the restriction of productive VZV infection in cultured nonprimate cells may be governed primarily by postentry events.Much of our understanding of alphaherpesvirus entry comes from studies of herpes simplex virus type 1 (HSV-1) (60, 61). HSV-1 envelope glycoproteins gB and/or gC mediate binding of virus to cell surface glycosaminoglycans, predominantly heparan sulfate (59, 63). Following virus binding, gD engages one of several entry receptors (19,40,58,60). Fusion of viral and cellular membranes then ensues, either directly at the cell surface (15, 16) or following endocytosis (38,42,43) by a mechanism that requires the participation of gB, gD, a heterodimer of gH and gL, and a gD entry receptor (5,44,47,65). The emerging picture of entry from HSV-1 studies is one that involves multiple entry receptors and multiple entry pathways. This versatility likely contributes to the wide host range observed for HSV-1 in vitro and the success of this virus as a pathogen.Much less is known about the binding and entry of VZV, largely due to the difficulty in obtaining high titers of cell-free (CF) virus. An entry process similar to that described for HSV-1 has been proposed for VZV, commencing with binding of virions to heparan sulfate via gB (25, 70), followed by interaction of viral glycoproteins with an entry receptor (7, 70). The interaction of mannose-6-phosphate (M6P) groups found on at least four VZV glycoproteins (gB, gE, gH, and gI) with the cation-independent mannose-6-phosphate receptor (CI-MPR) is believed to facilitate virus entry (7,17,70). While the exact consortium of VZV proteins needed for virus-cell fusion has not been defined, results from cell-cell fusion assays (9,11,12,46), blocking assays with antiglycoprotein antibo...

Cholesterol Dependence of Varicella-Zoster Virion Entry into Target Cells

“…We found that preincubation of target cells with such agents (e.g., bafilomycin A at 10 nM) prior to the introduction of virus completely blocked infectivity. Similar findings have recently been reported by Finnen et al (14). Although this is compatible with a role for endosomal acidification in viral fusion, alternative explanations include a change in the trafficking of viral receptors following endosomal alkalinization.…”

“…Some evidence was recently provided that VZ virion entry into a nonpermissive cell type depends on a low-pH compartment, since it was partially inhibited by lysosomotropic agents (14). We sought to clarify whether VZ virions gain entry by fusing directly with the plasma membrane or instead undergo endocytosis prior to fusion with the membrane delimiting an intracellular compartment.…”

Low-pH-Dependent Changes in the Conformation and Oligomeric State of the Prefusion Form of Herpes Simplex Virus Glycoprotein B Are Separable from Fusion Activity

“…An emerging theme in herpesviral entry is that cellular endosomal pH is required, often in a cell-specific manner (23,30,40,41,46,50,63). gB conformation change is central to the molecular understanding of why herpesviruses require low pH to initiate infection.…”

Six1 Overexpression in Mammary Cells Induces Genomic Instability and Is Sufficient for Malignant Transformation

“…MCF12A human mammary epithelial cells were cultured, as previously described (26), in either 5% horse serum or 5% fetal bovine serum (FBS). NIH3T3 mouse fibroblasts were cultured as described (27). Stable clones in both of these cells lines were generated by transfection with a Six expression plasmid SIXFL (14) or pcDNA3.1(CAT) (Invitrogen), followed by selection in 400 Ag/mL G418.…”