Resolving heterogeneous infection states within a virus microenvironment

Viral infections spread within complex cellular microenvironments that shape the outcome of infection. As such, intra-cellular communication cascades, as well as communication between infected cells and cells in the surrounding tissue create a virus microenvironment. Here, we describe our efforts to characterize communication at the intra- and inter-cellular levels. At the intra-cellular level, we consider the dynamic remodeling of organelles and organelle-organelle contacts, investigating their effects on metabolism and immune responses during infections with several DNA and RNA viruses. Upon infection with human cytomegalovirus (HCMV), we investigate the paradox of infection-induced mitochondria fragmentation concurrent with increased respiration. We integrate super-resolution microscopy, cryotomography, proteomics, and molecular assays to show how HCMV infection drives the formation of mitochondrial-ER encapsulations that protect fragmented mitochondria against mitophagy and promote bioenergetics. At the interface between this metabolic rewiring and immune signaling, we consider that several human viruses, including HCMV, induce a Warburg-like effect that results in increased lactate production. We show the effect of lactate on host immune factors during infections with HCMV and herpes simplex virus 1 (HSV-1). At the inter-cellular level, we establish an approach to characterize a virus microenvironment, separating cell populations based on their proximity to an infection site. We show that HCMV infection primes uninfected, proximal cells for infections with HCMV, HSV-1, and influenza A. To resolve heterogeneous infection states within this virus microenvironment, we design a single cell proteomics workflow. We uncover distinct infection trajectories and terminal states, as well as decision points that control immune responses. Overall, these findings demonstrate how infection reshapes a microenvironment and how spatial proximity to infection guides cell fate.