Single cell analysis reveals human cytomegalovirus drives latently infected cells towards an anergic-like monocyte state

Human cytomegalovirus (HCMV) causes a lifelong infection through establishment of latency. Although reactivation from latency can cause life-threatening disease, our molecular understanding of HCMV latency is incomplete. Here we use single cell RNA-seq analysis to characterize latency in monocytes and hematopoietic stem and progenitor cells (HSPCs). In monocytes, we identify host cell surface markers that enable enrichment of latent cells harboring higher viral transcript levels, which can reactivate more efficiently, and are characterized by reduced intrinsic immune response that is important for viral gene expression. Significantly, in latent HSPCs, viral transcripts could be detected only in monocyte progenitors and were also associated with reduced immune-response. Overall, our work indicates that regardless of the developmental stage in which HCMV infects, HCMV drives hematopoietic cells towards a weaker immune-responsive monocyte state and that this anergic-like state is crucial for the virus ability to express its transcripts and to eventually reactivate

Defining the Transcriptional Landscape during Cytomegalovirus Latency with Single-Cell RNA Sequencing

Primary infection with human cytomegalovirus (HCMV) results in a lifelong infection due to its ability to establish latent infection, with one characterized viral reservoir being hematopoietic cells. Although reactivation from latency causes serious disease in immunocompromised individuals, our molecular understanding of latency is limited. Here, we delineate viral gene expression during natural HCMV persistent infection by analyzing the massive RNA-seq atlas generated by the Genotype-Tissue Expression (GTEx) project. This systematic analysis reveals that HCMV persistence in-vivo is prevalent in diverse tissues. Unexpectedly, we find only viral transcripts that resemble gene expression during various stages of lytic infection with no evidence of any highly restricted latency-associated viral gene expression program. To further define the transcriptional landscape during HCMV latent infection, we also used single cell RNA-seq and a tractable experimental latency model. In contrast to some current views on latency, we also find no evidence for any highly restricted latency-associated viral gene expression program. Instead, we reveal that latency-associated gene expression largely mirrors a late lytic viral program albeit at much lower levels of expression. Overall, our work has the potential to revolutionize our understanding of HCMV persistence and suggests that latency is governed mainly by quantitative changes, with a limited number of qualitative changes, in viral gene expression.This research was supported by the EU-FP7-PEOPLE career integration grant, the Israeli Science Foundation (1073/14; N.S.-G.), Infect-ERA (TANKACY; N.S.-G.), the European Research Council starting grant (StG-2014-638142; N.S.-G.), the British Medical Research Programme (grant G0701279; J.S.), a Wellcome Research Studentship Grant (B.K.), and the Cambridge NIHR BRC Cell Phenotyping Hub. N.S.-G. is incumbent of the Skirball career development chair in new scientist

Defining the Transcriptional Landscape during Cytomegalovirus Latency with Single-Cell RNA Sequencing

Primary infection with human cytomegalovirus (HCMV) results in a lifelong infection due to its ability to establish latent infection, with one characterized viral reservoir being hematopoietic cells. Although reactivation from latency causes serious disease in immunocompromised individuals, our molecular understanding of latency is limited. Here, we delineate viral gene expression during natural HCMV persistent infection by analyzing the massive transcriptome RNA sequencing (RNA-seq) atlas generated by the Genotype-Tissue Expression (GTEx) project. This systematic analysis reveals that HCMV persistence in vivo is prevalent in diverse tissues. Notably, we find only viral transcripts that resemble gene expression during various stages of lytic infection with no evidence of any highly restricted latency-associated viral gene expression program. To further define the transcriptional landscape during HCMV latent infection, we also used single-cell RNA-seq and a tractable experimental latency model. In contrast to some current views on latency, we also find no evidence for any highly restricted latency-associated viral gene expression program. Instead, we reveal that latency-associated gene expression largely mirrors a late lytic viral program, albeit at much lower levels of expression. Overall, our work has the potential to revolutionize our understanding of HCMV persistence and suggests that latency is governed mainly by quantitative changes, with a limited number of qualitative changes, in viral gene expression

An iPSC-Derived Myeloid Lineage Model of Herpes Virus Latency and Reactivation.

Herpesviruses undergo life-long latent infection which can be life-threatening in the immunocompromised. Models of latency and reactivation of human cytomegalovirus (HCMV) include primary myeloid cells, cells known to be important for HCMV latent carriage and reactivation in vivo. However, primary cells are limited in availability, and difficult to culture and to genetically modify; all of which have hampered our ability to fully understand virus/host interactions of this persistent human pathogen. We have now used iPSCs to develop a model cell system to study HCMV latency and reactivation in different cell types after their differentiation down the myeloid lineage. Our results show that iPSCs can effectively mimic HCMV latency/reactivation in primary myeloid cells, allowing molecular interrogations of the viral latent/lytic switch. This model may also be suitable for analysis of other viruses, such as HIV and Zika, which also infect cells of the myeloid lineage