Characterization of the Human Cytomegalovirus gH/gL/UL128-131 Complex That Mediates Entry into Epithelial and Endothelial Cells

Abstract

The entry of human cytomegalovirus (HCMV) into biologically relevant epithelial and endothelial cells involves endocytosis followed by low-pH-dependent fusion. This entry pathway is facilitated by the HCMV UL128, UL130, and UL131 proteins, which form one or more complexes with the virion envelope glycoprotein gH/gL. gH/gL/UL128-131 complexes appear to be distinct from the gH/gL/gO complex, which likely facilitates entry into fibroblasts. In order to better understand the assembly and protein-protein interactions of gH/gL/ UL128-131 complexes, we generated HCMV mutants lacking UL128-131 proteins and nonreplicating adenovirus vectors expressing gH, gL, UL128, UL130, and UL131. Our results demonstrate that UL128, UL130, and UL131 can each independently assemble onto gH/gL scaffolds. However, the binding of individual UL128-131 proteins onto gH/gL can significantly affect the binding of other proteins; for example, UL128 increased the binding of both UL130 and UL131 to gH/gL. Direct interactions between gH/UL130, UL130/UL131, gL/UL128, and UL128/UL130 were also observed. The export of gH/gL complexes from the endoplasmic reticulum (ER) to the Golgi apparatus and cell surface was dramatically increased when all of UL128, UL130, and UL131 were coexpressed with gH/gL (with or without gO expression). Incorporation of gH/gL complexes into the virion envelope requires transport beyond the ER. Thus, we concluded that UL128, UL130, and UL131 must all bind simultaneously onto gH/gL for the production of complexes that can function in entry into epithelial and endothelial cells.

The pentameric complex of human Cytomegalovirus: cell tropism, virus dissemination, immune response and vaccine development

“…[6]) and then the pentameric complex in 2008 (Ryckman et al [10]) were found to be indispensable for virus and viral product transfer from endothelial cells to leukocytes (from Revello and Gerna [148]). …”

“…A significant contribution to the development of a HCMV vaccine was the identification of the pentameric complex (PC), which consists of HCMV gH, gL, pUL128, pUL130 and pUL131 [6][7][8][9][10].…”

Differential kinetics of human cytomegalovirus load and antibody responses in primary infection of the immunocompetent and immunocompromised host

“…In addition, the neutralizing antibody response was determined in parallel by inoculating virus-serum mixtures onto ARPE-19 epithelial cells and HELF cells. The double neutralization assay was based on the recent discovery that HCMV requires the pentamer to infect epithelial/endothelial/dendritic cells Ryckman et al, 2008), while infection of HELF cells requires the presence of gH/gL/gO on the HCMV envelope (Zhou et al, 2013). It has been demonstrated and repeatedly confirmed that neutralizing titres determined in epithelial cells are far higher than those obtained in HELF cells (Cui et al, 2008;Gerna et al, 2008).…”

“…The IgG antibody responses to the glycoprotein complexes representing the pentamer (gH/gL/ pUL128L), the dimer gH/gL (Hahn et al, 2004;Ryckman et al, 2008) and gB were investigated. In addition, the neutralizing antibody response was determined in parallel by inoculating virus-serum mixtures onto ARPE-19 epithelial cells and HELF cells.…”

“…Subsequently, it was shown that the UL128, UL130 and UL131 locus (thus referred to as UL128L) gene products formed a pentameric complex with gH/gL (gH/gL/pUL128/pUL130/pUL131), which was required for infection of both endothelial and epithelial cells (Wang & Shenk, 2005;Ryckman et al, 2008;. Multiple antigenic sites of the pentamer have been shown to elicit very potent neutralizing mAbs with high frequency, unlike gB which induces a predominant number of binding but non-neutralizing mAbs (Macagno et al, 2010;Pötzsch et al, 2011;Fouts et al, 2012).…”