Cyclophilin A is required for efficient human cytomegalovirus DNA replication and reactivation

Abstract

Human cytomegalovirus (HCMV) is a large DNA virus belonging to the subfamily Betaherpesvirinae. Haematopoietic cells of the myeloid lineage have been shown to harbour latent HCMV. However, following terminal differentiation of these cells, virus is reactivated, and in an immunocompromised host acute infection can occur. It is currently unknown which viral and cellular factors are involved in regulating the switch between lytic and latent infections. Cyclophilin A (CyPA) is a cellular protein that acts as a major factor in virus replication and/or virion maturation for a number of different viruses, including human immunodeficiency virus, hepatitis C virus, murine cytomegalovirus, influenza A virus and vaccinia virus. This study investigated the role of CyPA during HCMV infection. CyPA expression was silenced in human foreskin fibroblast (HF) and THP-1 cells using small interfering RNA (siRNA) technology, or the cells were treated with cyclosporin A (CsA) to inhibit CyPA activity. Silencing CyPA in HF cells with siRNA resulted in an overall reduction in virus production characterized by delayed expression of immediate-early (IE) proteins, decreased viral DNA loads and reduced titres. Furthermore, silencing of CyPA in THP-1 cells pre-and postdifferentiation prevented IE protein expression and virus reactivation from a non-productive state. Interestingly, it was observed that treatment of THP-1 cells with CsA prevented the cells from establishing a fully latent infection. In summary, these results demonstrate that CyPA expression is an important factor in HCMV IE protein expression and virus production in lytically infected HF cells, and is a major component in virus reactivation from infected THP-1 cells.

Human cytomegalovirus miR-UL112-1 promotes the down-regulation of viral immediate early-gene expression during latency to prevent T-cell recognition of latently infected cells

“…As observed by others (Keyes et al, 2012a), THP-1 cells acted as a tractable model of latent infection. However, we found that growing cells in 2 % FCS, rather than 10 % FCS (as is standard), after infection reduced large-scale expansion of the cell population and the associated loss of viral genomes (see Fig.…”

“…A number of models of HCMV latency that use established myeloid cell lines have been described and these include myeloid cells expressing the CD34 + marker, such as Kasumi-3 (O'Connor & Murphy, 2012), or CD14 + monocytic cells, such as THP-1s (Keyes et al, 2012a), which can recapitulate many aspects of HCMV latency and reactivation observed in primary cells (Albright & Kalejta, 2013). The ability to use such established cell line model systems in the analysis of HCMV latency has a number of advantages, which include their ease of use compared with primary cells, the lack of a requirement to recruit donors and, importantly, there would be no need to tissue type match each individual donor as with, for example, immunological assays.…”

Inhibition of cyclophilin A suppresses H2O2‐enhanced replication of HCMV through the p38 MAPK signaling pathway

“…This is may be the reason why silencing CyPA could inhibit the HCMV replication in the absence of H 2 O 2 20. Furthermore, this study has demonstrated that CsA could inhibit MCMV replication in neural stem/progenitor cells while it has little impact in MEF cells 19.…”

Human Cytomegalovirus Latency and Reactivation in and Beyond the Myeloid Lineage

“…However, the exact myeloid cell types that efficiently carry and reactivate HCMV in vivo will require a much more detailed analysis of naturally latent myeloid cells, perhaps using combinations of specific myeloid cell markers. Unfortunately, as discussed above, the frequency of naturally latent cells in the peripheral blood makes these types of analyses beyond our current technology and so many laboratories have resorted to the use of infection of primary cells [21,[23][24][25][26][27][28], as well as established cells lines [29][30][31] in culture as models of experimentally latent infection. In all cases, these models allow the long-term culture (up to 20 days for primary myeloid cells) of cells carrying quiescent viral genomes from which virus can be reactivated after differentiation along the myeloid lineage.…”