Human cytomegalovirus glycoprotein B variants affect viral entry, cell fusion, and genome stability

Abstract

Human cytomegalovirus (HCMV), like many other DNA viruses, can cause genome instability and activate a DNA damage response (DDR). Activation of ataxia-telangiectasia mutated (ATM), a kinase activated by DNA breaks, is a hallmark of the HCMV-induced DDR. Here we investigated the activation of caspase-2, an initiator caspase activated in response to DNA damage and supernumerary centrosomes. Of 7 HCMV strains tested, only strain AD169 activated caspase-2 in infected fibroblasts. Treatment with an ATM inhibitor or inactivation of PIDD or RAIDD inhibited caspase-2 activation, indicating that caspase-2 was activated by the PIDDosome. A set of chimeric HCMV strains was used to identify the genetic basis of this phenotype. Surprisingly, we found a single nucleotide polymorphism within the AD169 UL55 ORF, resulting in a D275Y amino acid exchange within glycoprotein B (gB), to be responsible for caspase-2 activation. As gB is an envelope glycoprotein required for fusion with host cell membranes, we tested whether gB(275Y) altered viral entry into fibroblasts. While entry of AD169 expressing gB(275D) proceeded slowly and could be blocked by a macropinocytosis inhibitor, entry of wild-type AD169 expressing gB(275Y) proceeded more rapidly, presumably by envelope fusion with the plasma membrane. Moreover, gB(275Y) caused the formation of syncytia with numerous centrosomes, suggesting that cell fusion triggered caspase-2 activation. These results suggest that gB variants with increased fusogenicity accelerate viral entry, cause cell fusion, and thereby compromise genome stability. They further suggest the ATM-PIDDosome-caspase-2 signaling axis alerts the cell of potentially dangerous cell fusion.

Development of a TaqMan Based Real‐Time Fluorescent Quantitative PCR Assay for Detection of Porcine Cytomegalovirus in Semen

“…e gB gene is highly conserved within the Herpesviridae family, and is widely used as a candidate gene in herpesvirus species identification and phylogenetic analysis [26][27][28]. Here, in this study, the nucleotide and amino acid sequences comparison demonstrated that DPOL gene sequences in different PCMV isolates are highly conserved, with no significant variation between DPOL nucleotide and amino acid sequences from different PCMV isolates.…”

Human Cytomegalovirus Primary Infection and Reactivation: Insights From Virion-Carried Molecules

“…gB1 and gB2 types are often seen in patients with transplantation and infection with human immunodeficiency virus (HIV), respectively ( Fries et al, 1994 ; Shepp et al, 1996 ). The 275Y variants of gB expressed on the AD169 strain promote fusogenicity with activation of caspase 2 and the DNA damage response since the 275D variants show less syncytium formation and an inability to trigger caspase 2 ( Tang et al, 2019 ). Whether gB genotypes are associated with cell tropism are largely elusive.…”

Human Cytomegalovirus Cell Tropism and Host Cell Receptors

“…However, the mechanism of entry into fibroblasts is still under discussion. Recent observations suggest that it may occur via a rapid micropinocytosis [54] or even by direct fusion with the outer plasma membrane through the fusogenic activity of gB [55].…”

“…The discovery that HCMV uses different glycoprotein complexes for entry into endothelial cells/epithelial cells [42,43] and into fibroblasts [52] prompted us to investigate whether human antibody response to natural HCMV infection displayed different levels of neutralization potential against the infection of these two cell types. Results showed that in the convalescent phase of primary HCMV infection, the reciprocal geometric mean neutralizing antibody titer was >30-times more potent in neutralizing infection of epithelial cells/endothelial cells than that of fibroblasts [55]. The primary candidates for this differential neutralizing activity appeared to be antibodies directed against UL128L proteins, as indirectly suggested by murine and rabbit polyclonal antibodies raised against the individual components of pUL128L [44,56,57].…”