Mild Acidic pH Inhibition of the Major Pathway of Herpes Simplex Virus Entry into HEp-2 Cells

Abstract

SUMMARYPenetration of the KOS strain of herpes simplex virus type 1 (HSV-1) and the MS and 333 strains of herpes simplex virus type 2 (HSV-2) into HEp-2 cells at pH 6.3 was at least 100-fold less efficient than at pH 7-4. Penetration of two low passage clinical isolates was completely blocked at pH 6.3. The syncytium-forming HSV-1 strains GC and MP were less sensitive than KOS to the mild acidic conditions. The inhibition was completely reversed upon neutralization of the medium. Penetration was assayed by plaque production following protection from acid inactivation upon virus entry. Penetration of HSV-1 KOS into Vero and HEL diploid fibroblast cells was similarly inhibited. HSV-1 KOS grown in 2-deoxy-D-glucose and monensin was also extensively inhibited at pH 6.3 but virus grown in 2-deoxy-D-glucose penetrated more slowly than normal virus at pH 7.4. Electron microscopy of HSV-1 KOS infection indicated that fusion and endocytosis occur at both pH 7.4 and 6.3 but that fusion predominates at pH 7-4 and endocytosis predominates at pH 6.3. These results indicate that fusion at the plasma membrane is the major route of productive entry for HSV, that Strains of HSV can differ in their pH dependence for penetration and this may determine whether virus infection can occur following endocytic uptake.

Roles for Endocytosis and Low pH in Herpes Simplex Virus Entry into HeLa and Chinese Hamster Ovary Cells

“…When viruses such as Semliki Forest virus are treated with buffers at pHs 4.8 to 6.5, their fusion glycoproteins are prematurely activated, so when the acid-treated virus is added to cells, it is incompetent for entry (3). As regards HSV, it has been known for some time that penetration of cell-bound virions is inhibited by exposure to sodium citrate buffered to the highly acidic pH of 3 (21,24,46).…”

Comparison of Heparin-sensitive Attachment of Pseudorabies Virus (PRV) and Herpes Simplex Virus Type 1 and Identification of Heparin-binding PRV Glycoproteins

“…Many studies on the entry of herpesviruses into the cell have been done (Epstein et al, 1964;Morgan et al, 1968;Dales & Silverberg, 1969;Smith & deHarven, 1974;DeLuca et al, 1981;Rosenthal et al, 1989), but little was known about attachment (Fuller & Spear, 1985;Johnson & Ligas, 1988) until it was shown recently by WuDunn & Spear (1989) that heparan sulphate proteoglycans present on the surface of many cell types (H66k et al, 1984) serve as an attachment receptor for herpes simplex virus (HSV) types 1 and 2. They used heparin, a glycosaminoglycan structurally related to heparan sulphate, to block specifically the attachment of HSV to the cell surface and to reduce the number of plaques formed by HSV on HEp-2 cells.…”

Bromodeoxyuridine-Labeled Viral Particles as a Tool for Visualization of the Immediate-Early Events of Human Cytomegalovirus Infection

“…This is in clear agreement with earlier EM studies showing capsids within the cytoplasm at approximately 20 min p.i. (13,17). We also visualized foci "rimming" the nucleus by 30 min p.i.…”

“…The advent of electron microscopy (EM) in the 1940s greatly aided the quest to capture the nature of the interaction of a virus with the host cell membrane and transport of the virion capsid (in the case of most DNA viruses) to the nuclear compartment. In the herpesvirus field, there have been several elegant studies utilizing EM to visualize the initial entry events of membrane fusion and the subsequent deposition of the capsid into the cytoplasm (13,17,22). Green fluorescent protein technology introduced the ability to specifically label viral capsid, tegument, and glycoproteins (and thus manufacture green virions), which could then be used to study the late life cycle events of assembly and export of herpesvirus virions in real time (6)(7)(8)16).…”