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ABSTRACT Astrocytes provide physical and metabolic support for neurons, regulate the blood–brain barrier, and react to injury, infection, and disease. When astrocytes become reactive, maintenance of the inflammatory state and its functional implications throughout chronic neuroinflammation are all poorly understood. Several models of acute inflammation have revealed astrocyte subpopulations that go beyond a two‐activation state model, instead encompassing distinct functional subsets. However, how reactive astrocyte (RA) subsets evolve over time during chronic inflammatory disease or infection has been difficult to address. Here we use a prolific human pathogen,Toxoplasma gondii, that causes lifelong infection in the brain alongside aLcn2CreERT2reporter mouse line to examine reactive astrocyte subsets during chronic neuroinflammation. Single‐cell RNA sequencing revealed diverse astrocyte populations including transcriptionally uniqueLcn2CreERT2+ RAs which change over the course of infection in a subset‐dependent manner. In addition to an immune‐regulatingLcn2CreERT2+ astrocyte population enriched with gene transcripts encoding chemokines CCL5, CXCL9, CXCL10, and receptors CCR7 and IL7R, a specific subset ofLcn2CreERT2+ astrocytes highly expressedtransthyretin(Ttr), a secreted carrier protein involved in glycolytic enzyme activation and potential vasculature regulation and angiogenesis. These findings provide novel information about the evolution and diversity of reactive astrocyte subtypes and functional signatures at different stages of infection, revealing an undocumented role for transthyretin‐expressing astrocytes in immune regulation at the central nervous system (CNS) vasculature. 

Mical family enzymes are unusual actin regulators that prime filaments (F-actin) for disassembly via the site-specific oxidation of M44/M47. Filamentous actin acts as a substrate of Mical enzymes, as well as an activator of their NADPH oxidase activity, which leads to hydrogen peroxide generation. Mical enzymes are required for cytokinesis, muscle and heart development, dendritic pruning, and axonal guidance, among other processes. Thus, it is critical to understand how this family of actin regulators functions in different cell types. Vertebrates express six actin isoforms in a cell-specific manner, but MICALs’ impact on their intrinsic properties has never been systematically investigated. Our data reveal the differences in the intrinsic dynamics of Mical-oxidized actin isoforms. Furthermore, our results connect the intrinsic dynamics of actin isoforms and their redox state with the patterns of hydrogen peroxide (H2O2) generation by MICALs. We documented that the differential properties of actin isoforms translate into the distinct patterns of hydrogen peroxide generation in Mical/NADPH-containing systems. Moreover, our results establish a conceptual link between actin stabilization by interacting factors and its ability to activate MICALs’ NADPH oxidase activity. Altogether, our results suggest that the regulatory impact of MICALs may differ depending on the isoform-related identities of local actin networks.