Abstract
Unraveling the heterogeneity in biological systems provides the key to understanding of the fundamental dynamics that regulate host pathogen relationships at the single cell level. While most studies have determined virus-host cell interactions using cultured cells in bulk, recent advances in deep protein profiling from single cells enable the understanding of the dynamic response equilibrium of single cells even within the same cell types. Mass cytometry allows the simultaneous detection of multiple proteins in single cells, which helps to evaluate alterations in multiple signaling networks that work in tandem in deciding the response of a cell to the presence of a pathogen or other stimulus. In applying this technique to studying varicella zoster virus (VZV), it was possible to better understand the molecular basis for lymphotropism of the virus and how virus-induced effects on T cells promoted skin tropism. While the ability of VZV to manifest itself in the skin is well established, how the virus is transported to the skin and causes the characteristic VZV skin lesions was not well elucidated. Through mass cytometry analysis of VZV-infected tonsil T cells, we were able to observe that VZV unleashes a “remodeling” program in the infected T cells that not only makes these T cells more skin tropic but also at the same time induces changes that make these T cells unlikely to respond to immune stimulation during the journey to the skin.
Highlights
Varicella zoster virus (VZV) is a medically important human alphaherpesvirus that causes varicella as the manifestation of primary infection and herpes zoster when it reactivates from latency (Figure 1; Zerboni et al, 2014)
The lymphotropism of VZV is essential for the pathogenesis of primary infection as well as for the establishment of latency in sensory ganglion neurons since VZV is delivered to skin sites of replication by T cells during primary infection and to sensory ganglia either by retrograde transport along neuronal axons from varicella skin lesions or directly to ganglia by VZV-infected T cells (Ku et al, 2002; Schaap et al, 2005; Zerboni et al, 2005)
Our work on VZV T cell tropism substantiates major advantages of this approach as follows: (a) testing single cells revealed that VZV orchestrates a continuum of changes within heterogeneous host cells, regardless of their basal state, which could not be shown by averaged measurements and (b) multi-parametric analysis of single cells was vital because there is no one “skin homing” marker on human T cells which could be measured to prove this functional consequence of VZV infection
Summary
Varicella zoster virus (VZV) is a medically important human alphaherpesvirus that causes varicella (chickenpox) as the manifestation of primary infection and herpes zoster (shingles) when it reactivates from latency (Figure 1; Zerboni et al, 2014). In our studies of VZV pathogenesis, single cell mass cytometry made it possible to overcome major challenges in the investigation of VZV T cell tropism, including that tonsil T cells are heterogeneous differentiated primary cells, signaling pathways in these cells are in varying states of activation and deactivation, and transformed T cell lines do not reproduce the diversity of primary T cells encountered by the virus in the natural host. The more scattered distribution of the VZV-infected T cells (green dots) showed that VZV induced remodeling directs alterations, resulting in a widely heterogenous population of T cells; the heterogeneity is most likely contributed by the fact that these cells are at different stages of virus infection/ replication This remodeling was a direct consequence of VZV induced changes in expression of several surface proteins of the naïve T cells to impart an activated, skin tropic effector phenotype without which, naïve T cells would not be expected to leave the tonsils and traffic to the skin (Figure 5B)
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