A Critical Analysis of the Evidence for the SARS-CoV-2 Origin Hypotheses.

When humans experience a new, devastating viral infection such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), significant challenges arise. How should individuals as well as societies respond to the situation? One of the primary questions concerns the origin of the SARS-CoV-2 virus that infected and was transmitted efficiently among humans, resulting in a pandemic. At first glance, the question appears straightforward to answer. However, the origin of SARS-CoV-2 has been the topic of substantial debate primarily because we do not have access to some relevant data. At least two major hypotheses have been suggested: a natural origin through zoonosis followed by sustained human-to-human spread or the introduction of a natural virus into humans from a laboratory source. Here, we summarize the scientific evidence that informs this debate to provide our fellow scientists and the public with the tools to join the discussion in a constructive and informed manner. Our goal is to dissect the evidence to make it more accessible to those interested in this important problem. The engagement of a broad representation of scientists is critical to ensure that the public and policy-makers can draw on relevant expertise in navigating this controversy.

Human cytomegalovirus attenuates AKT activity by destabilizing insulin receptor substrate proteins.

IMPORTANCE: Human cytomegalovirus (HCMV) requires inactivation of AKT to efficiently replicate, yet how AKT is shut off during HCMV infection has remained unclear. We show that UL38, an HCMV protein that activates mTORC1, is necessary and sufficient to destabilize insulin receptor substrate 1 (IRS1), a model insulin receptor substrate (IRS) protein. Degradation of IRS proteins in settings of excessive mTORC1 activity is an important mechanism for insulin resistance. When IRS proteins are destabilized, PI3K cannot be recruited to growth factor receptor complexes, and hence, AKT membrane recruitment, a rate limiting step in its activation, fails to occur. Despite its penchant for remodeling host cell signaling pathways, our results reveal that HCMV relies upon a cell-intrinsic negative regulatory feedback loop to inactivate AKT. Given that pharmacological inhibition of PI3K/AKT potently induces HCMV reactivation from latency, our findings also imply that the expression of UL38 activity must be tightly regulated within latently infected cells to avoid spontaneous reactivation.

Field Research Is Essential to Counter Virological Threats.

The interface between humans and wildlife is changing and, with it, the potential for pathogen introduction into humans has increased. Avian influenza is a prominent example, with an ongoing outbreak showing the unprecedented expansion of both geographic and host ranges. Research in the field is essential to understand this and other zoonotic threats. Only by monitoring dynamic viral populations and defining their biology in situ can we gather the information needed to ensure effective pandemic preparation.

CD34+ Hematopoietic Progenitor Cell Subsets Exhibit Differential Ability To Maintain Human Cytomegalovirus Latency and Persistence.

In human cytomegalovirus (HCMV)-seropositive patients, CD34+ hematopoietic progenitor cells (HPCs) provide an important source of latent virus that reactivates following cellular differentiation into tissue macrophages. Multiple groups have used primary CD34+ HPCs to investigate mechanisms of viral latency. However, analyses of mechanisms of HCMV latency have been hampered by the genetic variability of CD34+ HPCs from different donors, availability of cells, and low frequency of reactivation. In addition, multiple progenitor cell types express surface CD34, and the frequencies of these populations differ depending on the tissue source of the cells and culture conditions in vitro In this study, we generated CD34+ progenitor cells from two different embryonic stem cell (ESC) lines, WA01 and WA09, to circumvent limitations associated with primary CD34+ HPCs. HCMV infection of CD34+ HPCs derived from either WA01 or WA09 ESCs supported HCMV latency and induced myelosuppression similar to infection of primary CD34+ HPCs. Analysis of HCMV-infected primary or ESC-derived CD34+ HPC subpopulations indicated that HCMV was able to establish latency and reactivate in CD38+ CD90+ and CD38+/low CD90- HPCs but persistently infected CD38- CD90+ cells to produce infectious virus. These results indicate that ESC-derived CD34+ HPCs can be used as a model for HCMV latency and that the virus either latently or persistently infects specific subpopulations of CD34+ cells.IMPORTANCE Human cytomegalovirus infection is associated with severe disease in transplant patients and understanding how latency and reactivation occur in stem cell populations is essential to understand disease. CD34+ hematopoietic progenitor cells (HPCs) are a critical viral reservoir; however, these cells are a heterogeneous pool with donor-to-donor variation in functional, genetic, and phenotypic characteristics. We generated a novel system using embryonic stem cell lines to model HCMV latency and reactivation in HPCs with a consistent cellular background. Our study defined three key stem cell subsets with differentially regulated latent and replicative states, which provide cellular candidates for isolation and treatment of transplant-mediated disease. This work provides a direction toward developing strategies to control the switch between latency and reactivation.

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.

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+ 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+ 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+ 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 mock-infected 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+ 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 receptor, EGFR, in CD34+ HPCs initiates early events necessary for successful latent infection of this cell type. EGFR and associated signaling players may therefore represent promising targets for mitigating HCMV persistence.

Antagonistic determinants controlling replicative and latent states of human cytomegalovirus infection.

UNLABELLED: The mechanisms by which viruses persist and particularly those by which viruses actively contribute to their own latency have been elusive. Here we report the existence of opposing functions encoded by genes within a polycistronic locus of the human cytomegalovirus (HCMV) genome that regulate cell type-dependent viral fates: replication and latency. The locus, referred to as the UL133-UL138 (UL133/8) locus, encodes four proteins, pUL133, pUL135, pUL136, and pUL138. As part of the ULb' region of the genome, the UL133/8 locus is lost upon serial passage of clinical strains of HCMV in cultured fibroblasts and is therefore considered dispensable for replication in this context. Strikingly, we could not reconstitute infection in permissive fibroblasts from bacterial artificial chromosome clones of the HCMV genome where UL135 alone was disrupted. The loss of UL135 resulted in complex phenotypes and could ultimately be overcome by infection at high multiplicities. The requirement for UL135 but not the entire locus led us to hypothesize that another gene in this locus suppressed virus replication in the absence of UL135. The defect associated with the loss of UL135 was largely rescued by the additional disruption of the UL138 latency determinant, indicating a requirement for UL135 for virus replication when UL138 is expressed. In the CD34(+) hematopoietic progenitor model of latency, viruses lacking only UL135 were defective for viral genome amplification and reactivation. Taken together, these data indicate that UL135 and UL138 comprise a molecular switch whereby UL135 is required to overcome UL138-mediated suppression of virus replication to balance states of latency and reactivation. IMPORTANCE: Mechanisms by which viruses persist in their host remain one of the most poorly understood phenomena in virology. Herpesviruses, including HCMV, persist in an incurable, latent state that has profound implications for immunocompromised individuals, including transplant patients. Further, the latent coexistence of HCMV may increase the risk of age-related pathologies, including vascular disease. The key to controlling or eradicating HCMV lies in understanding the molecular basis for latency. In this work, we describe the complex interplay between two viral proteins, pUL135 and pUL138, which antagonize one another in infection to promote viral replication or latency, respectively. We previously described the role of pUL138 in suppressing virus replication for latency. Here we demonstrate a role of pUL135 in overcoming pUL138-mediated suppression for viral reactivation. From this work, we propose that pUL135 and pUL138 constitute a molecular switch balancing states of latency and reactivation.

An epistatic relationship between the viral protein kinase UL97 and the UL133-UL138 latency locus during the human cytomegalovirus lytic cycle.

UNLABELLED: We report that UL133-UL138 (UL133/8), a transcriptional unit within the ULb' region (ULb') of the human cytomegalovirus (HCMV) genome, and UL97, a viral protein kinase encoded by HCMV, play epistatic roles in facilitating progression of the viral lytic cycle. In studies with HCMV strain TB40/E, pharmacological blockade or genetic ablation of UL97 significantly reduced the levels of mRNA and protein for IE2 and viral early and early-late genes during a second wave of viral gene expression that commenced at between 24 and 48 h postinfection. These effects were accompanied by significant defects in viral DNA synthesis and viral replication. Interestingly, deletion of UL133/8 likewise caused significant defects in viral DNA synthesis, viral gene expression, and viral replication, which were not exacerbated upon UL97 inhibition. When UL133/8 was restored to HCMV laboratory strain AD169, which otherwise lacks the locus, the resulting recombinant virus replicated similarly to the parental virus. However, during UL97 inhibitor treatment, the virus in which UL133/8 was restored showed significantly exacerbated defects in viral DNA synthesis, viral gene expression, and production of infectious progeny virus, thus recapitulating the differences between wild-type TB40/E and its UL133/8-null derivative. Phenotypic evaluation of mutants null for specific open reading frames within UL133/8 revealed a role for UL135 in promoting viral gene expression, viral DNA synthesis, and viral replication, which depended on UL97. Taken together, our findings suggest that UL97 and UL135 play interdependent roles in promoting the progression of a second phase of the viral lytic cycle and that these roles are crucial for efficient viral replication. IMPORTANCE: A unique feature of the herpesviruses, such as human cytomegalovirus (HCMV), is that they can undergo latency, a state during which the virus silences its gene expression, which allows lifelong viral persistence in immunocompetent hosts. We have uncovered an unexpected link between a cluster of HCMV genes involved in latency, UL133-UL138, and a virally encoded protein kinase, UL97, which plays crucial roles in manipulating the cell cycle during HCMV lytic replication. Although viral immediate early (IE) gene expression is essential for HCMV lytic replication, the activation of IE gene expression in latently infected cells is not sufficient to result in production of infectious virus. Our findings here and in an accompanying study (M. Umashankar, M. Rak, F. Bughio, P. Zagallo, K. Caviness, and F. D. Goodrum, J. Virol. 88:5987-6002, 2014) show that proteins expressed from the UL133-UL138 latency locus and UL97 play interdependent roles in overcoming checkpoints that restrict the viral lytic replication cycle, findings which suggest intriguing implications for establishment of and reactivation from HCMV latency.

Viral and host network analysis of the human cytomegalovirus transcriptome in latency.

HCMV genes UL135 and UL138 play opposing roles regulating latency and reactivation in CD34+ human progenitor cells (HPCs). Using the THP-1 cell line model for latency and reactivation, we designed an RNA sequencing study to compare the transcriptional profile of HCMV infection in the presence and absence of these genes. The loss of UL138 results in elevated levels of viral gene expression and increased differentiation of cell populations that support HCMV gene expression and genome synthesis. The loss of UL135 results in diminished viral gene expression during an initial burst that occurs as latency is established and no expression of eleven viral genes from the ULb' region even following stimulation for differentiation and reactivation. Transcriptional network analysis revealed host transcription factors with potential to regulate the ULb' genes in coordination with pUL135. These results reveal roles for UL135 and UL138 in regulation of viral gene expression and potentially hematopoietic differentiation.

Human cytomegalovirus attenuates AKT activity by destabilizing insulin receptor substrate proteins.

The phosphoinositide 3-kinase (PI3K)/AKT pathway plays crucial roles in cell viability and protein synthesis and is frequently co-opted by viruses to support their replication. Although many viruses maintain high levels of AKT activity during infection, other viruses, such as vesicular stomatitis virus and human cytomegalovirus (HCMV), cause AKT to accumulate in an inactive state. To efficiently replicate, HCMV requires FoxO transcription factors to localize to the infected cell nucleus (Zhang et. al. mBio 2022), a process directly antagonized by AKT. Therefore, we sought to investigate how HCMV inactivates AKT to achieve this. Subcellular fractionation and live cell imaging studies indicated that AKT failed to recruit to membranes upon serum-stimulation of infected cells. However, UV-inactivated virions were unable to render AKT non-responsive to serum, indicating a requirement for de novo viral gene expression. Interestingly, we were able to identify that UL38 (pUL38), a viral activator of mTORC1, is required to diminish AKT responsiveness to serum. mTORC1 contributes to insulin resistance by causing proteasomal degradation of insulin receptor substrate (IRS) proteins, such as IRS1, which are necessary for the recruitment of PI3K to growth factor receptors. In cells infected with a recombinant HCMV disrupted for UL38 , AKT responsiveness to serum is retained and IRS1 is not degraded. Furthermore, ectopic expression of UL38 in uninfected cells induces IRS1 degradation, inactivating AKT. These effects of UL38 were reversed by the mTORC1 inhibitor, rapamycin. Collectively, our results demonstrate that HCMV relies upon a cell-intrinsic negative feedback loop to render AKT inactive during productive infection.

A Critical Analysis of the Evidence for the SARS-CoV-2 Origin Hypotheses.

When humans experience a new, devastating viral infection such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), significant challenges arise. How should individuals as well as societies respond to the situation? One of the primary questions concerns the origin of the SARS-CoV-2 virus that infected and was transmitted efficiently among humans, resulting in a pandemic. At first glance, the question appears straightforward to answer. However, the origin of SARS-CoV-2 has been the topic of substantial debate primarily because we do not have access to some relevant data. At least two major hypotheses have been suggested: a natural origin through zoonosis followed by sustained human-to-human spread or the introduction of a natural virus into humans from a laboratory source. Here, we summarize the scientific evidence that informs this debate to provide our fellow scientists and the public with the tools to join the discussion in a constructive and informed manner. Our goal is to dissect the evidence to make it more accessible to those interested in this important problem. The engagement of a broad representation of scientists is critical to ensure that the public and policy-makers can draw on relevant expertise in navigating this controversy.

A Critical Analysis of the Evidence for the SARS-CoV-2 Origin Hypotheses.

When humans experience a new, devastating viral infection such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), significant challenges arise. How should individuals as well as societies respond to the situation? One of the primary questions concerns the origin of the SARS-CoV-2 virus that infected and was transmitted efficiently among humans, resulting in a pandemic. At first glance, the question appears straightforward to answer. However, the origin of SARS-CoV-2 has been the topic of substantial debate primarily because we do not have access to some relevant data. At least two major hypotheses have been suggested: a natural origin through zoonosis followed by sustained human-to-human spread or the introduction of a natural virus into humans from a laboratory source. Here, we summarize the scientific evidence that informs this debate to provide our fellow scientists and the public with the tools to join the discussion in a constructive and informed manner. Our goal is to dissect the evidence to make it more accessible to those interested in this important problem. The engagement of a broad representation of scientists is critical to ensure that the public and policy-makers can draw on relevant expertise in navigating this controversy.

The Akt Forkhead Box O Transcription Factor Axis Regulates Human Cytomegalovirus Replication.

The protein kinase Akt broadly impacts many cellular processes, including mRNA translation, metabolism, apoptosis, and stress responses. Inhibition of phosphatidylinositol 3-kinase (PI3K), a lipid kinase pivotal to Akt activation, triggers various herpesviruses to reactivate from latency. Hence, decreased Akt activity may promote lytic replication. Here, we show that Akt accumulates in an inactive form during human cytomegalovirus (HCMV) infection of permissive fibroblasts, as indicated by hypophosphorylation of sites that activate Akt, decreased phosphorylation of PRAS40, and pronounced nuclear localization of FoxO3a, a substrate that remains cytoplasmic when Akt is active. HCMV strongly activates mTORC1 during lytic infection, suggesting a potential mechanism for Akt inactivation, since mTORC1 negatively regulates PI3K. However, we were surprised to observe that constitutive Akt activity, provided by expression of Akt fused to a myristoylation signal (myr-Akt), caused a 1-log decrease in viral replication, accompanied by defects in viral DNA synthesis and late gene expression. These results indicated that Akt inactivation is required for efficient viral replication, prompting us to address which Akt substrates underpin this requirement. Interestingly, we found that short interfering RNA knockdown of FoxO3a, but not FoxO1, phenocopied the defects caused by myr-Akt, corroborating a role for FoxO3a. Accordingly, a chimeric FoxO3a-estrogen receptor fusion protein, in which nuclear localization is regulated by 4-hydroxytamoxifen instead of Akt, reversed the replication defects caused by myr-Akt. Collectively, our results reveal a role for FoxO transcription factors in HCMV lytic replication and argue that this single class of Akt substrates underpins the requirement for Akt inactivation during productive infection. IMPORTANCE Evidence from diverse herpesvirus infection models suggests that the PI3K/Akt signaling pathway suppresses reactivation from latency and that inactivation of the pathway stimulates viral lytic replication. Here, we show that Akt accumulates in an inactive state during HCMV infection of lytically permissive cells while the presence of constitutive Akt activity causes substantial viral replication defects. Although Akt phosphorylates a diverse array of cellular substrates, we identify an important role for the Forkhead box class O transcription factors. Our findings show that when FoxO3a nuclear localization is decoupled from its negative regulation by Akt, the viral replication defects observed in the presence of constitutively active Akt are reversed. Collectively, our results reveal that HCMV inactivates Akt to promote the nuclear localization of FoxO transcription factors, which strongly implies that FoxOs play critical roles in transactivating cellular and/or viral genes during infection.

Human Cytomegalovirus Host Interactions: EGFR and Host Cell Signaling Is a Point of Convergence Between Viral Infection and Functional Changes in Infected Cells.

Viruses have evolved diverse strategies to manipulate cellular signaling pathways in order to promote infection and/or persistence. Human cytomegalovirus (HCMV) possesses a number of unique properties that allow the virus to alter cellular events required for infection of a diverse array of host cell types and long-term persistence. Of specific importance is infection of bone marrow derived and myeloid lineage cells, such as peripheral blood monocytes and CD34+ hematopoietic progenitor cells (HPCs) because of their essential role in dissemination of the virus and for the establishment of latency. Viral induced signaling through the Epidermal Growth Factor Receptor (EGFR) and other receptors such as integrins are key control points for viral-induced cellular changes and productive and latent infection in host organ systems. This review will explore the current understanding of HCMV strategies utilized to hijack cellular signaling pathways, such as EGFR, to promote the wide-spread dissemination and the classic life-long herpesvirus persistence.

E4orf1 limits the oncolytic potential of the E1B-55K deletion mutant adenovirus.

Clinical trials have shown oncolytic adenoviruses to be tumor selective with minimal toxicity toward normal tissue. The virus ONYX-015, in which the gene encoding the early region 1B 55-kDa (E1B-55K) protein is deleted, has been most effective when used in combination with either chemotherapy or radiation therapy. Therefore, improving the oncolytic nature of tumor-selective adenoviruses remains an important objective for improving this form of cancer therapy. Cells infected during the G(1) phase of the cell cycle with the E1B-55K deletion mutant virus exhibit a reduced rate of viral late protein synthesis, produce fewer viral progeny, and are less efficiently killed than cells infected during the S phase. Here we demonstrate that the G(1) restriction imposed on the E1B-55K deletion mutant virus is due to the viral oncogene encoded by open reading frame 1 of early region 4 (E4orf1). E4orf1 has been reported to signal through the phosphatidylinositol 3'-kinase pathway leading to the activation of Akt, mTOR, and p70 S6K. Evidence presented here shows that E4orf1 may also induce the phosphorylation of Akt and p70 S6K in a manner that depends on Rac1 and its guanine nucleotide exchange factor Tiam1. Accordingly, agents that have been reported to disrupt the Tiam1-Rac1 interaction or to prevent phosphorylation of the ribosomal S6 kinase partially alleviated the E4orf1 restriction to late viral protein synthesis and enhanced tumor cell killing by the E1B-55K mutant virus. These results demonstrate that E4orf1 limits the oncolytic nature of a conditionally replicating adenovirus such as ONYX-015. The therapeutic value of similar oncolytic adenoviruses may be improved by abrogating E4orf1 function.

Human Cytomegalovirus miR-US5-2 Downregulation of GAB1 Regulates Cellular Proliferation and UL138 Expression through Modulation of Epidermal Growth Factor Receptor Signaling Pathways.

Regulation of epidermal growth factor (EGF) receptor (EGFR) signaling is critical for the replication of human cytomegalovirus (HCMV) as well as latency and reactivation in CD34+ hematopoietic progenitor cells. HCMV microRNAs (miRNAs) provide a means to modulate the signaling activated by EGF through targeting components of the EGFR signaling pathways. Here, we demonstrate that HCMV miR-US5-2 directly downregulates the critical EGFR adaptor protein GAB1 that mediates activation and sustained signaling through the phosphatidylinositol 3-kinase (PI3K) and MEK/extracellular signal-regulated kinase (ERK) pathways and cellular proliferation in response to EGF. Expression of HCMV UL138 is regulated by the transcription factor early growth response gene 1 (EGR1) downstream of EGFR-induced MEK/ERK signaling. We show that by targeting GAB1 and attenuating MEK/ERK signaling, miR-US5-2 indirectly regulates EGR1 and UL138 expression, which implicates the miRNA in critical regulation of HCMV latency.IMPORTANCE Human cytomegalovirus (HCMV) causes significant disease in immunocompromised individuals, including transplant patients. HCMV establishes latency in hematopoietic stem cells in the bone marrow. The mechanisms governing latency and reactivation of viral replication are complex and not fully understood. HCMV-encoded miRNAs are small regulatory RNAs that reduce protein expression. In this study, we found that the HCMV miRNA miR-US5-2 targets the epidermal growth factor receptor (EGFR) adaptor protein GAB1 which directly affects downstream cellular signaling pathways activated by EGF. Consequently, miR-US5-2 blocks the EGF-mediated proliferation of human fibroblasts. Early growth response gene 1 (EGR1) is a transcription factor activated by EGFR signaling that regulates expression of HCMV UL138. We show that miR-US5-2 regulates UL138 expression through GAB1-mediated downregulation of the signaling pathways that lead to EGR1 expression. These data suggest that miR-US5-2, through downregulation of GAB1, could play a critical role during reactivation from latency by reducing proliferation and UL138 expression.

The Role of the Human Cytomegalovirus UL133-UL138 Gene Locus in Latency and Reactivation.

Human cytomegalovirus (HCMV) latency, the means by which the virus persists indefinitely in an infected individual, is a major frontier of current research efforts in the field. Towards developing a comprehensive understanding of HCMV latency and its reactivation from latency, viral determinants of latency and reactivation and their host interactions that govern the latent state and reactivation from latency have been identified. The polycistronic UL133-UL138 locus encodes determinants of both latency and reactivation. In this review, we survey the model systems used to investigate latency and new findings from these systems. Particular focus is given to the roles of the UL133, UL135, UL136 and UL138 proteins in regulating viral latency and how their known host interactions contribute to regulating host signaling pathways towards the establishment of or exit from latency. Understanding the mechanisms underlying viral latency and reactivation is important in developing strategies to block reactivation and prevent CMV disease in immunocompromised individuals, such as transplant patients.

Human Cytomegalovirus Infection Suppresses CD34+ Progenitor Cell Engraftment in Humanized Mice.

Human cytomegalovirus (HCMV) infection is a serious complication in hematopoietic stem cell transplant (HSCT) recipients due to virus-induced myelosuppression and impairment of stem cell engraftment. Despite the clear clinical link between myelosuppression and HCMV infection, little is known about the mechanism(s) by which the virus inhibits normal hematopoiesis because of the strict species specificity and the lack of surrogate animal models. In this study, we developed a novel humanized mouse model system that recapitulates the HCMV-mediated engraftment failure after hematopoietic cell transplantation. We observed significant alterations in the hematopoietic populations in peripheral lymphoid tissues following engraftment of a subset of HCMV+ CD34+ hematopoietic progenitor cells (HPCs) within the transplant, suggesting that a small proportion of HCMV-infected CD34+ HPCs can profoundly affect HPC differentiation in the bone marrow microenvironment. This model will be instrumental to gain insight into the fundamental mechanisms of HCMV myelosuppression after HSCT and provides a platform to assess novel treatment strategies.