Human cytomegalovirus (HCMV) is a ?-herpesvirus infecting 44-100% of the population and remains a significant cause of morbidity and mortality in solid organ transplant (SOT) and allogeneic hematopoietic stem cell transplant (SCT) recipients. Infection in SCT patients is often associated with myelosuppression and graft failure due to virus reactivation from latency but the associated mechanisms are still largely unknown. We have recently identified a secreted viral factor, UL7, containing an Ig-like domain with structural and functional homology to carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1), a protein with multiple cellular activities including differentiation. We recently demonstrated that UL7 stimulates an inflammatory response in endothelial cells, inducing the secretion of IL-6 and other cytokines mediated in part through the phosphorylation of STAT3 and ERK1/2 in the JAK/STAT and MAP kinase pathways. Secretion of IL-6 and activation of the ERK-MAPK signaling pathway have been reported to stimulate viral reactivation. Importantly, we have shown that although UL7 is not expressed during latency, HCMV with a deletion of UL7 fails to reactivate in latently infected CD34+ HPCs in vitro and in vivo although the virus is replication competent in fibroblasts. These data indicate that UL7 is acting after the initiation of reactivation in latently infected CD34+ HPCs but before the lytic program is initiated, suggesting that UL7 reprogramming of the cell is essential for the reactivation event. In addition, we have shown that UL7 is sufficient to promote CD34+ HPC differentiation into myeloid lineage cells. Therefore, based upon our exciting new findings, we hypothesize that UL7 modulates the JAK-STAT-AKT pathways downstream of the Epidermal Growth Factor Receptor (EGFR) to promote reactivation from latency and cellular differentiation for viral dissemination. In this project we will examine UL7 signaling in CD34+ HPCs and identify the cellular pathways necessary for viral reactivation and hematopoiesis in the context of infection. We will also determine whether these UL7 pathways cross-talk with pathways modulated by US28 (Project 3) and UL133/8 (Project 1) or HCMV miRNAs (Project 2), other HCMV factors important to latency and differentiation that modulate host signaling downstream of EGFR (Project 5). We propose the following specific aims: 1) To determine how UL7 signaling in CD34+ HPCs intersects with EGFR signaling pathways using a multi-omics approach; 2) To determine what molecular characteristics of UL7 mediate HCMV latency and reactivation using both in vitro CD34+ HPCs and in vivo humanized BLT mouse model; 3) To determine what UL7 signaling pathways promote CD34+ HPC hematopoiesis also using both in vitro CD34+ HPC and in vivo huBLT models. Results of this study will generate new virus latency and reactivation paradigms promoting the development of novel therapies to prevent virus reactivation and to treat HCMV-mediated myelosuppression.
PROJECT 4 NARRATIVE Human Cytomegalovirus (HCMV) remains a significant cause of morbidity and mortality after Solid Organ Transplantation (SOT) and Hematopoietic Stem Cell Transplantation (HSCT), and myelosuppression is a common clinical manifestation of HCMV infection in these patients. Recently, we have determined that HCMV UL7 targets signaling pathways involved in cellular differentiation that are important for viral reactivation and hematopoiesis. The goal of this project is, together with the other projects that form this Program Project Grant (PPG) proposal, to determine mechanisms involved in HCMV-encoded UL7 in viral latency and reactivation as well as bone marrow graft rejection and hematopoiesis.
|Collins-McMillen, Donna; Chesnokova, Liudmila; Lee, Byeong-Jae et al. (2018) HCMV Infection and Apoptosis: How Do Monocytes Survive HCMV Infection? Viruses 10:|
|Crawford, Lindsey B; Kim, Jung Heon; Collins-McMillen, Donna et al. (2018) Human Cytomegalovirus Encodes a Novel FLT3 Receptor Ligand Necessary for Hematopoietic Cell Differentiation and Viral Reactivation. MBio 9:|
|Collins-McMillen, Donna; Stevenson, Emily V; Kim, Jung Heon et al. (2017) HCMV utilizes a non-traditional STAT1 activation cascade via signaling through EGFR and integrins to efficiently promote the motility, differentiation, and polarization of infected monocytes. J Virol :|