The enhanced host-cell permissiveness of human cytomegalovirus is mediated by the Ras signaling pathway

Abstract

Human cytomegalovirus utilizes cellular signal transduction pathways to activate viral or cellular transcription factors involved in the control of viral gene expression and DNA replication. In the present study, we demonstrate that Harvey-ras-transformed cells show increased permissiveness to human cytomegalovirus when compared to their parental non-transformed cells. Both the progeny viral yield and the protein levels were elevated in the human cytomegalovirus-infected Harvey-ras-transformed cells requiring active viral gene replication, as shown by the infection with UV-inactivated human cytomegalovirus. Inhibition of Ras or of key molecules of the Ras pathway, effectively suppressed viral infection in the Harvey-ras-transformed cells. On a cellular level, the human cytomegalovirus-infected Harvey-ras-transformed cells formed larger cellular foci, which were significantly higher in number, compared to the uninfected cells and preferentially recruited human cytomegalovirus virions, thereby incriminating human cytomegalovirus infection for the increased transformation of these cells. Furthermore, proliferation assays revealed a higher rate for the human cytomegalovirus-infected Harvey-ras-transformed cells compared to mock-infected cells, whereas human cytomegalovirus infection had no considerable effect on the proliferation of the non-transformed cells. Higher susceptibility to apoptosis was also detected in the human cytomegalovirus-infected ras-transformed cells, which in combination with the higher progeny virus reveals a mode by which human cytomegalovirus achieves efficient spread of infection in the cells expressing the oncogenic Harvey-ras (12V) gene. Collectively, our data suggest that human cytomegalovirus employs the host-cell Ras signaling pathway to ensue viral expression and ultimately successful propagation. Transformed cells with an activated Ras signaling pathway are therefore particularly susceptible to human cytomegalovirus infection.

The Role of RhoA, RhoB and RhoC GTPases in Cell Morphology, Proliferation and Migration in Human Cytomegalovirus (HCMV) Infected Glioblastoma Cells

“…The wild-type HCMV AD169 strain was used in this study. The virus stocks were propagated and titrated on HFF cells according to standard protocols [56]. For viral infections, the cells were infected with HCMV at the indicated MOI for 2 hours and then the inoculum was removed and replaced by fresh medium.…”

RhoB is a component of the human cytomegalovirus assembly complex and is required for efficient viral production

“…The conditions for fixation and immunodetection of viral and cellular proteins, were as previously described. 86 For microtubule staining, cells were fixed in 4% paraformaldehyde in cytoskeleton buffer (1.1 M Na2HPO4, 0.4 M KH2PO4, 137 mM NaCl, 5 mM KCl, 2 mM MgCl2, 2 mM EGTA, 5mM PIPES, 5 mM Glucose, pH 6.1) for 5 min at room temperature.…”

Intermittent Bulk Release of Human Cytomegalovirus

“…Tipifarnib has been shown to reduce Rab27a, nSMase2, and Alix levels, which might result in an effect on trafficking of viral components to assembly sites or membrane remodeling during secondary envelopment [42]. Alternatively, inhibition of the cellular farnesyltransferase by Tipifarnib might also act on Ras signaling pathways, which have been reported to positively influence HCMV, HSV-1, and other herpesvirus infections [69][70][71]. An effect of Tipifarnib on transcription would be the simplest explanation for the reduced pp150 levels at 3 and 4 dpi.…”