Magee, J. A. | bioRxiv Science

bioRxiv2021

Single-cell RNA transcriptome analysis reveals CXCL16/CXCR6 as maintenance factors for CNS tissue resident T cells that drive synapse elimination during viral recovery

BackgroundEmerging RNA viruses that target the central nervous system (CNS) lead to cognitive sequelae in survivors. Studies in humans and mice infected with West Nile virus (WNV), a re-emerging RNA virus associated with learning and memory deficits, revealed microglial-mediated synapse elimination within the hippocampus. Moreover, CNS resident memory T (TRM) cells activate microglia, limiting synapse recovery and inducing spatial learning defects in WNV-recovered mice. The signals involved in T cell-microglia interactions are unknown. MethodsHere, we examined the murine WNV-recovered forebrain using single-cell RNA sequencing to identify putative ligand-receptor pairs involved in intercellular communication between T cells and microglia. Clustering and differential gene analyses were followed by protein validation, genetic and antibody-based approaches utilizing an established murine model of WNV recovery in which microglia and complement promote ongoing hippocampal synaptic loss. ResultsProfiling of host transcriptome at 25 days post-infection revealed a shift in forebrain homeostatic microglia to activated subpopulations with transcriptional signatures that have previously been observed in studies of neurodegenerative diseases. Importantly, CXCL16/CXCR6, a chemokine signaling pathway involved in TRM cell biology, was identified as critically regulating CXCR6 expressing CD8+ TRM cell numbers within the WNV-recovered forebrain. We demonstrate that CXCL16 is highly expressed by all myeloid cells, and its unique receptor, CXCR6, is highly expressed on all CD8+ T cells. Using genetic and pharmacological approaches, we demonstrate that CXCL16/CXCR6 is required not only for the maintenance of WNV-specific, CD8 TRM cells in the post-infectious CNS, but also contributes to their expression of TRM cell markers. Moreover, CXCR6+CD8+ T cells are required for glial activation and ongoing synapse elimination. ConclusionsWe provide a comprehensive assessment of the role of CXCL16/CXCR6 as an interaction link between microglia and CD8+ T cells that maintains forebrain TRM cells, microglial and astrocyte activation, and ongoing synapse elimination in virally recovered animals. We also show that therapeutic targeting of CXCL16 during recovery may reduce CNS CD8+ TRM cells.

immunology↗

bioRxiv2020

Developmental stage-specific changes in protein synthesis differentially sensitize hematopoietic stem cells and erythroid progenitors to impaired ribosome biogenesis

Ribosomopathies encompass a collection of human genetic disorders that often arise from mutations in ribosomal proteins or ribosome biogenesis factors. Despite ubiquitous requirement of ribosomes for protein synthesis, ribosomopathies present with tissue- and cell-type-specific disorders, and blood is particularly affected. Several ribosomopathies present with congenital anemias and bone marrow failure, and accordingly, erythroid lineage cells and hematopoietic stem cells (HSCs) are preferentially impaired by ribosomal dysfunction. However, the factors that influence this cell-type-specific sensitivity are incompletely understood. Here, we show that protein synthesis rates change during HSC and erythroid progenitor ontogeny. Fetal HSCs exhibit significantly higher protein synthesis than adult HSCs. Despite protein synthesis differences, reconstituting activity of both fetal and adult HSCs is severely disrupted by a ribosomal mutation (Rpl24Bst/+). In contrast, fetal erythroid lineage progenitors exhibit significantly lower protein synthesis than their adult counterparts. Protein synthesis declines during erythroid differentiation, but the decline starts earlier in fetal differentiation than in adults. Strikingly, the Rpl24Bst/+ mutation impairs fetal, but not adult erythropoiesis, by impairing proliferation at fetal erythroid progenitor stages with the lowest protein synthesis relative to their adult counterparts. Thus, developmental and cell-type-specific changes in protein synthesis can sensitize hematopoietic cells to impaired ribosome biogenesis. Key PointsO_LIFetal HSCs synthesize much more protein per hour than young adult HSCs in vivo C_LIO_LIFetal erythroid progenitors synthesize much less protein per hour than young adult erythroid progenitors in vivo C_LIO_LIDifferences in protein synthesis dynamics distinguish fetal and adult erythroid differentiation C_LIO_LIA ribosomal mutation that reduces protein synthesis impairs fetal and adult HSCs C_LIO_LIReduced protein synthesis impairs fetal but not adult erythroid progenitors C_LI

developmental biology↗