The lipid-sensor TREM2 aggravates disease in a model of LCMV-induced hepatitis

textabstractLipid metabolism is increasingly being appreciated to affect immunoregulation, inflammation and pathology. In this study we found that mice infected with lymphocytic choriomeningitis virus (LCMV) exhibit global perturbations of circulating serum lipids. Mice lacking the lipid-sensing surface receptor triggering receptor expressed on myeloid cells 2 (Trem2 -/-) were protected from LCMV-induced hepatitis and showed improved virus control despite comparable virus-specific T cell responses. Non-hematopoietic expression of TREM2 was found to be responsible for aggravated hepatitis, indicating a novel role for TREM2 in the non-myeloid compartment. These results suggest a link between virus-perturbed lipids and TREM2 that modulates liver pathogenesis upon viral infection. Targeted interventions of this immunoregulatory axis may ameliorate tissue pathology in hepatitis

Characterization of host proteins interacting with the lymphocytic choriomeningitis virus L protein

<div><p>RNA-dependent RNA polymerases (RdRps) play a key role in the life cycle of RNA viruses and impact their immunobiology. The arenavirus lymphocytic choriomeningitis virus (LCMV) strain Clone 13 provides a benchmark model for studying chronic infection. A major genetic determinant for its ability to persist maps to a single amino acid exchange in the viral L protein, which exhibits RdRp activity, yet its functional consequences remain elusive. To unravel the L protein interactions with the host proteome, we engineered infectious L protein-tagged LCMV virions by reverse genetics. A subsequent mass-spectrometric analysis of L protein pulldowns from infected human cells revealed a comprehensive network of interacting host proteins. The obtained LCMV L protein interactome was bioinformatically integrated with known host protein interactors of RdRps from other RNA viruses, emphasizing interconnected modules of human proteins. Functional characterization of selected interactors highlighted proviral (DDX3X) as well as antiviral (NKRF, TRIM21) host factors. To corroborate these findings, we infected <i>Trim21</i>^-/- mice with LCMV and found impaired virus control in chronic infection. These results provide insights into the complex interactions of the arenavirus LCMV and other viral RdRps with the host proteome and contribute to a better molecular understanding of how chronic viruses interact with their host.</p></div

Characterization of host proteins interacting with the lymphocytic choriomeningitis virus L protein

<div><p>RNA-dependent RNA polymerases (RdRps) play a key role in the life cycle of RNA viruses and impact their immunobiology. The arenavirus lymphocytic choriomeningitis virus (LCMV) strain Clone 13 provides a benchmark model for studying chronic infection. A major genetic determinant for its ability to persist maps to a single amino acid exchange in the viral L protein, which exhibits RdRp activity, yet its functional consequences remain elusive. To unravel the L protein interactions with the host proteome, we engineered infectious L protein-tagged LCMV virions by reverse genetics. A subsequent mass-spectrometric analysis of L protein pulldowns from infected human cells revealed a comprehensive network of interacting host proteins. The obtained LCMV L protein interactome was bioinformatically integrated with known host protein interactors of RdRps from other RNA viruses, emphasizing interconnected modules of human proteins. Functional characterization of selected interactors highlighted proviral (DDX3X) as well as antiviral (NKRF, TRIM21) host factors. To corroborate these findings, we infected <i>Trim21</i>^-/- mice with LCMV and found impaired virus control in chronic infection. These results provide insights into the complex interactions of the arenavirus LCMV and other viral RdRps with the host proteome and contribute to a better molecular understanding of how chronic viruses interact with their host.</p></div

Identification of L protein interactome.

<p><b>(A)</b> GO enrichment analyses for the L protein interactome based on the molecular functions (light grey) and biological processes (dark grey) of interactors followed by visualization with ReviGO. <b>(B)</b> Overview of L protein interactomes classified based on the protein functions and visualized in Cytoscape. The data is based on the mass-spectrometry derived list of proteins identified in L protein pulldowns after filtration using Top3 quantitation and SAINTexpress software as detailed in Materials and Methods.</p

Generation and characterization of LCMV strains expressing a tagged L protein.

<p><b>(A)</b> Viral titer of N- and C-terminal L protein-tagged Cl13 LCMV and WT Cl13 LCMV measured by focus forming assay at 72 hours post infection after reverse genetic rescue on BHK21 cells. (<b>B</b>) HEK293T cells were infected at a MOI of 0.01 with either Cl13_L-HA or with untagged Cl13. Supernatant was harvested and viral loads were measured at the indicated time points by focus forming assay. <b>(C</b> and <b>D)</b> C57BL/6J mice were infected with 2x10⁶ FFU of the indicated viruses. Viral titers were determined in <b>(C)</b> blood at indicated time points and in <b>(D)</b> organs 20 days post infection. <b>(E)</b> C57BL/6J mice were infected with 2x10⁶ FFU of the indicated viruses and the percentage of GP33-specific-tetramer⁺ CD8⁺ T cells was quantified in the spleen at 8 days post infection. Each symbol and bar represents the mean ± SEM of three to five mice. Statistical significance was calculated by Two-way ANOVA (B-<b>C</b>) or unpaired t-test (<b>D-E</b>). Significant p values were indicated as follows: ns—non significant, * p≤0.05,: ** p≤0.01.</p

Viral RNA-dependent RNA-polymerases target host proteome by common and virus-specific strategies.

<p><b>(A)</b> Integrated interactome of viral RdRp targets. Host proteins interacting with viral RdRps are highlighted in blue, the rest of the human proteome—in grey. <b>(B)</b> Largest connected component (LCC) analyses for global RdRps and LCMV only datasets. <b>(C)</b> Functional protein modules targeted by RdRps based on the community detection method. <b>(D)</b> Heat map representing virus-specific targeting of protein functional modules.</p

Functional screening for L protein interactors involved in LCMV life cycle.

<p>(<b>A</b>) Two independently generated HeLa S3 CRISPR-Cas9 targeted cell pools per gene of interest for 5 genes were infected in triplicate wells with LCMV Cl13 WT at a MOI of 0.01 and viral loads were measured at 36 hours post infection by focus forming assay. The obtained data were normalized to the non-target control and log2 transformed. (<b>B</b>) Two HeLa S3 CRISPR-Cas9 TRIM21-targeted cell pools were reconstituted either with TRIM21-expressing plasmid or with non-target control and 36 hour post transfection were infected in triplicate wells with LCMV Cl13 WT at a MOI of 0.01. Viral loads were measured at 36 hours post infection by focus forming assay. The obtained data were normalized to the non-target control and log2 transformed. (<b>C</b>-<b>D</b>) C57BL/6 and <i>Trim21</i>^-/- mice were infected with 2x10⁶ FFU of the indicated viruses. Viral titers were determined in (<b>C</b>) blood at indicated time points and in (<b>D</b>) organs at 21 days post infection. The data shown in (<b>C</b>) is representative of two similar experiments. Each symbol and bar represents the mean ± SEM of three to five mice. Statistical significance was calculated by unpaired t-test (<b>A, B, D</b>) or by Two-way ANOVA (<b>C</b>). Significant p values were indicated as follows: ns—non significant, * p≤0.05, ** p≤0.01, *** p≤0.001, **** p≤0.0001.</p

Comparative analysis of genome-scale, base-resolution DNA methylation profiles across 580 animal species

DNA methylation is involved in regulatory processes throughout the animal kingdom. Here, the authors map DNA methylation in 535 vertebrates and 45 invertebrates, establishing a reference dataset for cross-species analysis and exploring epigenetic variation across vertebrate evolution

The methyltransferase Setdb2 mediates virus-induced susceptibility to bacterial superinfection

Immune responses are tightly regulated to ensure efficient pathogen clearance while avoiding tissue damage. Here we report that SET domain bifurcated 2 (Setdb2) was the only protein lysine methyltransferase induced during influenza virus infection. Setdb2 expression depended on type-I interferon signaling and it repressed the expression of the neutrophil attractant Cxcl1 and other NF-κB target genes. This coincided with Setdb2 occupancy at the Cxcl1 promoter, which in the absence of Setdb2 displayed reduced H3K9 tri-methylation. Setdb2 hypomorphic gene-trap mice exhibited increased neutrophil infiltration in sterile lung inflammation and were less sensitive to bacterial superinfection upon influenza virus infection. This suggests that a Setdb2-mediated regulatory crosstalk between the type-I interferon and NF-κB pathways represents an important mechanism for virus-induced susceptibility to bacterial superinfection

Type I Interferon Signaling Disrupts the Hepatic Urea Cycle and Alters Systemic Metabolism to Suppress T Cell Function.

Infections induce complex host responses linked to antiviral defense, inflammation, and tissue damage and repair. We hypothesized that the liver, as a central metabolic hub, may orchestrate systemic metabolic changes during infection. We infected mice with chronic lymphocytic choriomeningitis virus (LCMV), performed RNA sequencing and proteomics of liver tissue, and integrated these data with serum metabolomics at different infection phases. Widespread reprogramming of liver metabolism occurred early after infection, correlating with type I interferon (IFN-I) responses. Viral infection induced metabolic alterations of the liver that depended on the interferon alpha/beta receptor (IFNAR1). Hepatocyte-intrinsic IFNAR1 repressed the transcription of metabolic genes, including Otc and Ass1, which encode urea cycle enzymes. This led to decreased arginine and increased ornithine concentrations in the circulation, resulting in suppressed virus-specific CD8+ T cell responses and ameliorated liver pathology. These findings establish IFN-I-induced modulation of hepatic metabolism and the urea cycle as an endogenous mechanism of immunoregulation. VIDEO ABSTRACT