Host signaling and EGR1 transcriptional control of human cytomegalovirus replication and latency

Abstract

Sustained phosphotinositide3-kinase (PI3K) signaling is critical to the maintenance of alpha and beta herpesvirus latency. We have previously shown that the beta-herpesvirus, human cytomegalovirus (CMV), regulates epidermal growth factor receptor (EGFR), upstream of PI3K, to control states of latency and reactivation. How signaling downstream of EGFR is regulated and how this impacts CMV infection and latency is not fully understood. We demonstrate that CMV downregulates EGFR early in the productive infection, which blunts the activation of EGFR and its downstream pathways in response to stimuli. However, CMV infection sustains basal levels of EGFR and downstream pathway activity in the context of latency in CD34+ hematopoietic progenitor cells (HPCs). Inhibition of MEK/ERK, STAT or PI3K/AKT pathways downstream of EGFR increases viral reactivation from latently infected CD34+ HPCs, defining a role for these pathways in latency. We hypothesized that CMV modulation of EGFR signaling might impact viral transcription important to latency. Indeed, EGF-stimulation increased expression of the UL138 latency gene, but not immediate early or early viral genes, suggesting that EGFR signaling promotes latent gene expression. The early growth response-1 (EGR1) transcription factor is induced downstream of EGFR signaling through the MEK/ERK pathway and is important for the maintenance of hematopoietic stemness. We demonstrate that EGR1 binds the viral genome upstream of UL138 and is sufficient to promote UL138 expression. Further, disruption of EGR1 binding upstream of UL138 prevents the establishment of latency in CD34+ HPCs. Our results indicate a model whereby UL138 modulation of EGFR signaling feeds back to promote UL138 gene expression and suppression of replication for latency. By this mechanism, the virus has hardwired itself into host cell biology to sense and respond to changes in homeostatic host cell signaling.

Cell signaling and cytomegalovirus reactivation: what do Src family kinases have to do with it?

“…This protein has been reported to have an antagonistic relationship with another viral protein, UL135 [36]. During latency, UL138 regulates epidermal growth factor receptor (EGFR) localization to ensure Pi3K signaling is active in the cell which is hypothesized to help maintain latency via the Egr-1 transcription factor [37] an event also targeted by a virally encoded miRNA [38]. Consistent with this is the observation that Pi3K inhibitors trigger HCMV reactivation in vitro suggesting that this pathway actively supports latency [37].…”

“…During latency, UL138 regulates epidermal growth factor receptor (EGFR) localization to ensure Pi3K signaling is active in the cell which is hypothesized to help maintain latency via the Egr-1 transcription factor [37] an event also targeted by a virally encoded miRNA [38]. Consistent with this is the observation that Pi3K inhibitors trigger HCMV reactivation in vitro suggesting that this pathway actively supports latency [37]. Interestingly, work from the Trono laboratory demonstrated that chloroquine also drove HCMV reactivation by modulating the activity of the transcriptional repressor, TRIM28/KAP1 [39].…”

Bright and Early: Inhibiting Human Cytomegalovirus by Targeting Major Immediate-Early Gene Expression or Protein Function

“…Initial PI3K/AKT activation is required for efficient viral entry as well as optimal replication in fibroblasts and establishment of latency in monocytes [156,158,[160][161][162][163]. However, at later times during infection inhibition of EGFR or PI3K seems to favour viral replication and reactivation from latency suggesting a negative regulatory role at this point [164][165][166][167]. Besides PI3K/AKT signalling, various other kinase pathways are known to be activated very early during HCMV infection.…”

FOXO Transcription Factors Activate Alternative Major Immediate Early Promoters to Induce Human Cytomegalovirus Reactivation

“…While iP1 and iP2 promoters are necessary for efficient reactivation, the factors regulating their activity are unknown. Reactivation is induced by differentiation of monocytes into macrophages, a process that is mimicked in experimental latency models by treating latently infected cells with phorbol esters (e.g., TPA) (10) or LY294002 (20), a chemical inhibitor of the PI3K/mTOR pathway. A literature review found that phorbol esters, LY294002, and monocyte differentiation all activate FOXO family of transcription factors (21).…”