The human cytomegalovirus US27 gene product constitutively activates antioxidant response element (ARE)-mediated transcription through Gβγ, phosphoinositide 3-kinase (PI3K), and nuclear respiratory factor 1 (NRF-1)

Jordan M. Boeck, Gregory A. Stowell, Christine M. O'Connor, Juliet V. Spencer
2018 Journal of Virology  
Human cytomegalovirus (HCMV) is a widespread pathogen that modulates host chemokine signaling during persistent infection in the host. HCMV encodes four proteins with homology to the chemokine receptor family of G protein-coupled receptors (GPCRs): US27, US28, UL33, and UL78. Each of the four receptors modulates host CXCR4 signaling. US28, UL33 and UL78 impair CXCR4 signaling outcomes while US27 enhances signaling, as evidenced by increased calcium mobilization and cell migration to CXCL12. To
more » ... nvestigate effects of US27 on CXCR4 during virus infection, fibroblasts were infected with BAC-derived clinical strain HCMV TB40/E-mCherry (WT) or mutants lacking US27 (TB40/E-mCherry-US27Δ) or all four GPCRs (TB40 E-mCherry-allΔ), or expressing only US27, but not US28, UL33, or UL78 (TB40/E-mCherry-US27wt). CXCR4 gene expression was significantly higher in WT and US27wt-infected fibroblasts. This effect was evident at 3 hours post-infection, suggesting that US27 derived from the parental virion enhanced CXCR4 expression. Reporter gene assays demonstrated US27 increased transcriptional activity regulated by the antioxidant response element (ARE), and siRNA treatment indicated this effect was mediated by NRF-1, the primary transcription factor for CXCR4. Increased translocation of NRF-1 into the nucleus of WT-infected cells compared to mock- or US27Δ-infected cells was confirmed by immunofluorescence microscopy. Chemical inhibitors targeting Gβγ and phosphoinositide 3-kinase (PI3K) ablated the increase in ARE-driven transcription, implicating these proteins as mediators of US27-stimulated gene transcription. This work identifies the first signaling pathway activated by HCMV US27 and may reveal a novel regulatory function for this orphan viral receptor in stimulating stress response genes during infection. IMPORTANCE Human cytomegalovirus (HCMV) is the most common congenital infection worldwide, causing deafness, blindness, and other serious birth defects. CXCR4 is a human chemokine receptor that is crucial for both fetal development and immune responses. We found that the HCMV protein US27 stimulates increased expression of CXCR4 through activation of the transcription factor nuclear respiratory factor 1 (NRF-1). NRF-1 regulates stress response genes that contain the antioxidant response element (ARE), and HCMV infection is associated with increased expression of many stress response genes when US27 is present. Our results show that the US27 protein activates the NRF-1/ARE pathway, stimulating higher expression of CXCR4 and other stress response genes, which is likely to be beneficial for virus replication and/or immune evasion.
doi:10.1128/jvi.00644-18 fatcat:cxgoqmzbeng2hffogmcp5ouiyq