Abstract
Human cytomegalovirus (HCMV) encodes four homologs of G protein coupled receptors (vGPCRs), of which two, designated UL33 and US28, signal constitutively. UL33 and US28 are also conserved with chemokine receptors: US28 binds numerous chemokine classes, including the membrane bound chemokine, fractalkine; whereas UL33 remains an orphan receptor. There is emerging data that UL33 and US28 each contribute to HCMV associated disease, although no studies to date have reported their potential contribution to aberrant placental physiology that has been detected with HCMV congenital infection. We investigated the signaling repertoire of UL33 and US28 and their potential to enable trophoblast mobilization in vitro. Results demonstrate the constitutive activation of CREB by each vGPCR in ACIM-88 and HTR-8SVneo trophoblasts; constitutive NF-kB activation was detected for US28 only. Constitutive signaling by each vGPCR enabled trophoblast migration. For US28, fractalkine exhibited inverse agonist activity and dampened trophoblast migration. UL33 stimulated expression of both p38 mitogen activated (MAP) and Jun N-terminal (JNK) kinases; while p38 MAP kinase stimulated CREB, JNK was inhibitory, suggesting that UL33 dependent CREB activation was regulated by p38/JNK crosstalk. Given that chemokines and their receptors are important for placental development, these data point to the potential of HCMV UL33 and US28 to interfere with trophoblast responses which are important for normal placental development.
Highlights
Herpesvirus genomes show evidence of host gene capture events that have evolved to facilitate key aspects of the virus lifecycle
Subsequent studies have been conducted in cells or tissues of relevance to herpesvirus infection, revealing viral G protein coupled receptors (GPCRs) signaling characteristics that have the potential to contribute to pathological outcomes
The total protein expression of wild type (WT) and mutated derivatives of US28 and UL33 was evaluated in transfected HEK293 cells by ELISA (Figure 1A)
Summary
Herpesvirus genomes show evidence of host gene capture events that have evolved to facilitate key aspects of the virus lifecycle. Homologues of G protein coupled receptors (GPCRs), the largest and most diverse membrane receptors in eukaryotes, have been identified in both beta- and gamma-herpesviridae [1,2,3,4] A number of the viral GPCRs (vGPCRs) show homology to chemokine receptors and exhibit constitutive G protein dependent signaling [4]. Subsequent studies have been conducted in cells or tissues of relevance to herpesvirus infection, revealing viral GPCR (vGPCR) signaling characteristics that have the potential to contribute to pathological outcomes (reviewed by [5]). In these studies, we sought to evaluate the ability of HCMV US28 and UL33 to produce quantifiable, constitutive signaling in trophoblasts and to evaluate their contribution to trophoblast mobilization
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