Human Cytomegalovirus Requires Epidermal Growth Factor Receptor Signaling To Enter and Initiate the Early Steps in the Establishment of Latency in CD34 ⁺ Human Progenitor Cells

Abstract

The establishment of human cytomegalovirus (HCMV) latency and persistence relies on the successful infection of hematopoietic cells, which serve as sites of viral persistence and contribute to viral spread. Here, using blocking antibodies and pharmacological inhibitors, we document that HCMV activation of the epidermal growth factor receptor (EGFR) and downstream phosphatidylinositol 3-kinase (PI3K) mediates viral entry into CD34 ϩ human progenitor cells (HPCs), resulting in distinct cellular trafficking and nuclear translocation of the virus compared to that in other immune cells, such as we have documented in monocytes. We argue that the EGFR allows HCMV to regulate the cellular functions of these replication-restricted cells via its signaling activity following viral binding. In addition to regulating HCMV entry/ trafficking, EGFR signaling may also shape the early steps required for the successful establishment of viral latency in CD34 ϩ cells, as pharmacological inhibition of EGFR increases the transcription of lytic IE1/IE2 mRNA while curbing the expression of latency-associated UL138 mRNA. EGFR signaling following infection of CD34 ϩ HPCs may also contribute to changes in hematopoietic potential, as treatment with the EGFR kinase (EGFRK) inhibitor AG1478 alters the expression of the cellular hematopoietic cytokine interleukin 12 (IL-12) in HCMV-infected cells but not in mockinfected cells. These findings, along with our previous work with monocytes, suggest that EGFR likely serves as an important determinant of HCMV tropism for select subsets of hematopoietic cells. Moreover, our new data suggest that EGFR is a key receptor for efficient viral entry and that the ensuing signaling regulates important early events required for successful infection of CD34 ϩ HPCs by HCMV.IMPORTANCE HCMV establishes lifelong persistence within the majority of the human population without causing overt pathogenesis in healthy individuals. Despite this, reactivation of HCMV from its latent reservoir in the bone marrow causes significant morbidity and mortality in immunologically compromised individuals, such as bone marrow and solid organ transplant patients. Lifelong persistent infection has also been linked with the development of various cardiovascular diseases in otherwise healthy individuals. Current HCMV therapeutics target lytic replication, but not the latent viral reservoir; thus, an understanding of the molecular basis for viral latency and persistence is paramount to controlling or eliminating HCMV infection. Here, we show that the viral signalosome activated by HCMV binding to its entry re-

The Role of ErbB Receptors in Infection

“…Immune evasion [69] HPV Induces EGFR-dependent translocation of AnxA2, results in formation of the HPV-AnxA2-S100A10 binding complex required for viral entry Campylobacter jejuni Induces lipid-raft clustering and EGFR activation Host cell entry [44] Chlamydia pneumoniae Bacterial Pmp21 adhesin directly binds EGFR Host cell entry [10] Escherichia coli Disrupts endosome trafficking, resulting in diminished surface EGFR Unconfirmed [62] Helicobacter pylori Induces EGFR-dependent b-catenin nuclear translocation and PI3K/Akt signalling Host cell survival [16] Dephosphorylates EGFR to inhibit hBD3 expression and promote infection Immune modulation [104] Klebsiella pneumoniae Induces EGFR dependent inhibition of NF-kB translocation Immune modulation [65,66] Neisseria gonorrhoeae Induces redistribution of b-catenin from apical junction to cytoplasm through EGFR activation Host cell entry [51] Induces translocation of ErbB2 and ErbB3 to apical surfaces [61] Neisseria meningitidis Recruits and activates ErbB2 receptors Host cell entry [2] Mycobacterium leprae Directly binds ErbB2 receptors Host cell entry [14] Mycobacterium tuberculosis Prevent proper macrophage function resulting in enhanced infection Immune modulation [70] Salmonella Typhimurium Induces Claudin 2 expression through EGFR and downstream JNK activation Host cell entry [50] Shigella flexneri Induces PI5P production to regulate EGFR trafficking Host cell survival [17] Staphylococcus aureus Cleaves junction proteins occludin and E-cadherin to facilitate transmigration. EGFR activation is required Host cell entry [13] Fungi…”

“…HCMV-induced latency is another interesting tactic providing immune evasion and is also governed by EGFR activity. During infection, the ability to induce latency-associated UL138, while suppressing IE1/IE2 lytic genes, was shown to be EGFR dependent, by the use of EGFR-specific inhibitors [69]. Additionally, Mycobacterium tuberculosis can utilise EGFR-induced p38/MAPK signalling pathways within macrophages to prevent proper antimicrobial macrophage responses, resulting in enhanced murine infection [70].…”

Infected T98G glioblastoma cells support human cytomegalovirus reactivation from latency

“…Some cellular signaling pathway have been demonstrated to involve in HCMV latency and reactivation. Epidermal growth factor receptor (EGFR), its downstream phosphatidylinositol-3-kinase (PI(3) K) Kim et al, 2016), and HCMV-encoded miR-UL148D mediated immediate early response gene 5 (IER5)-cell division cycle 25B (CDC25B) (Pan et al, 2016) promote HCMV latency in CD34 + cells. PKA-CREB-TORC2 signaling cascade (Yuan et al, 2009), PKC mediated cellular CREB and NF-κB (Liu et al, 2010), and mitogen and stress activated kinases coupled with CREB (Kew et al, 2014) have been reported to activate IE gene expression from latency.…”

Targeted Therapies: Immunologic Effects and Potential Applications Outside of Cancer

“…For example, the principal envelope glycoprotein of human CMV preferentially binds to an EGFR monomer and oligomerizes with ErbB3, which then allows the virus to infect human fibroblast HEL cells . Recently, Kim et al demonstrated that human CMV binds to the EGFR and induces signaling via PI3K in CD34+ human progenitor cells, which mediate HCMV entry and viral trafficking as well as the establishment of viral latency . IAV also uses the EGFR as a coreceptor for binding to host cells.…”