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, 51 (6), 1074-1087.e9

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

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

Alexander Lercher et al. Immunity.

Abstract

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.

Keywords: CD8 T cells; hepatitis; hepatocyte; immunometabolism; infection; inflammation; interferons; liver; urea cycle; virus.

Conflict of interest statement

P.N.C. is the founder of Biocancer Treatment International, and G.S. is scientific advisor of Biocancer Treatment International.

Figures

None
Figure 1
Figure 1
LCMV Cl13 Induces Hepatic Metabolic Reprogramming, Translating to Changes in Systemic Metabolism during the Course of Infection (A and B) Viremia and RNemia of LCMV-clone-13-infected liver tissue (A; n = 3) and serum alanine transferase (ALT) and aspartate aminotransferase (AST) levels upon LCMV clone 13 infection up to 60 days after infection (B; n = 4–12). (C) Principal-component analyses (PCA) of liver tissue transcriptomes at 0, 2, 8, 30, and 60 days after infection (n = 3). (D) Hierarchical clustering (fragments per kilobase of transcript per million [FPKM]; k-means; Pearson’s correlation) of significantly changed genes at any indicated time point (n = 3). (E) Regulation of detected proteins in liver tissue at the corresponding time points (n = 3). (F) Enriched GO terms and pathways (ClueGO) of transcripts identified in the groups of clusters shown in (D). (G) Enrichment of the union of significantly regulated metabolic transcripts and proteins on KEGG metabolic database at any time point. (H) Hierarchical clustering of significantly regulated serum metabolites (n = 4; k-means; Pearson’s correlation). dpi, days post-infection. For (A), (B), and (H), one out of at least two representative experiments is shown. For (C)–(G), transcriptomic and proteomic data are derived from one experiment. Symbols represent the arithmetic mean ± SEM; ns, not significant; p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001 (Student’s t test). See also Figure S1.
Figure 2
Figure 2
Reduced Food Intake upon LCMV Infection Mainly Affects Lipid-Metabolism-Associated Genes in the Liver (A) PCA of liver transcriptomes of naive, 8 days after LCMV infection, and pair-fed (8 days) mice (n = 3). (B) Significantly deregulated transcripts in liver tissue upon LCMV infection at 8 days after infection compared to naive and pair-fed and between pair-fed and naive animals (n = 3). (C) Enriched GO terms and pathways (ClueGO) of significantly up- and downregulated genes of pair-fed versus naive mice (n = 3). (D) Significantly regulated metabolic genes in the livers of pair-fed versus naive animals super-imposed on the previously identified significantly regulated metabolic transcripts and proteins at the indicated time points after LCMV infection (based on Figure 1G). For (A)–(C), transcriptomic data are derived from one experiment. See also Figure S2.
Figure 3
Figure 3
Hepatocyte-Intrinsic IFNAR1 Signaling Is a Transcriptional Regulator of Liver Metabolism and Shapes Systemic Metabolism (A and B) Oxygen consumption rate (OCR) (A) and extracellular acidification rate (ECAR) (B) of wild-type and Ifnar1−/− primary hepatocytes treated for 4 h with IFN-β (n = 11). (C) Clustering by expression profile (FPKM; k-means; Pearson’s correlation) of transcripts significantly regulated (limma interaction model) by hepatocyte-intrinsic IFNAR1 signaling (n = 3). (D) Enriched GO terms and pathways (ClueGO) of transcripts identified in the groups clusters in (C). (E) Metabolism-associated transcripts significantly regulated by hepatocyte-intrinsic IFNAR1 signaling super-imposed on the previously identified significantly regulated metabolic transcripts and proteins at the indicated time points after LCMV infection (based on Figure 1G). (F) Serum metabolites significantly regulated (limma interaction model) by hepatocyte-intrinsic IFNAR1 signaling in naive and LCMV-infected animals (n = 3). For (A) and (B), data for one out of two representative experiments are shown. For (C), (D), and (F), transcriptomic and metabolomic data are derived from one experiment. Symbols represent the arithmetic mean ± SEM; p < 0.05; ∗∗p < 0.01 (Student’s t test). See also Figure S3.
Figure 4
Figure 4
IFNAR1 Contributes to Infection-Induced Metabolic Reprogramming of Hepatocytes (A) Depiction of the urea cycle, expression, and concentrations of the associated genes and serum metabolites in naive and LCMV-clone-13-infected Alb-Cre ERT2 Ifnar1fl/fl (Ifnar1Δ/Δ) and Ifnar1+/+ mice (n = 3). (B) Uniform Manifold Approximation and Projection (UMAP) clustering of single-cell RNA-seq data of primary hepatocytes sorted from naive or infected wild-type animals 2 days after LCMV infection. (C and D) Expression levels and correlation with Ifit1 levels (IFN-I-stimulated gene) of (C) Otc and (D) Ass1 in hepatocytes isolated from naive versus infected animals (n = 2; pooled for each condition). Each dot represents a single hepatocyte. (E) Representative immunohistochemical staining of OTC, ASS1, and STAT1 in liver sections of naive or LCMV-infected (2 and 8 days after infection) wild-type mice. (F) Quantification of immunohistochemical staining of OTC, ASS1, and STAT1 using HistoQuest software (n = 4 and 8 pictures per mouse were quantified). (G) Expression of Otc and Ass1 in sorted primary hepatocytes isolated from naive and LCMV-infected (2 and 8 days after infection) mice (n = 3). For metabolite data in (A), one of two representative experiments is shown. For (B)–(G), single-cell transcriptomic and histological data are derived from one experiment. Symbols represent the arithmetic mean ± SEM; p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001 (Student’s t test). Log 2 fold changes and adjusted p values for single-cell RNA-seq data were computed as described in the STAR Methods. See also Figure S4.
Figure 5
Figure 5
Viral Infection Reprograms the Hepatic Urea Cycle to Shape Systemic Metabolism (A) Blood ammonia upon LCMV-clone-13-infected wild-type mice (n = 5–6). (B) Schematic depiction of the carbon flow in urea cycle starting from 13C6 arginine illustrating 13C-labeled (full circles) and unlabeled 12C atoms of the respective metabolites. (C) Concentrations of 13C6-labeled arginine, 13C5-labeled ornithine, and 13C5-labeled citrulline in liver tissue of naive and LCMV-infected (2 and 8 days after infection) mice (n = 3–6). (D) Concentrations of 13C6-labeled arginine, 13C5-labeled ornithine, and 13C5-labeled citrulline in serum of naive and LCMV-infected (2 and 8 days after infection) mice (n = 3–6). (E) Systemic arginine-to-ornithine ratio of LCMV-infected (n = 8) wild-type animals. For (A), one of two representative experiments is shown. For (E), data are pooled from two independent experiments. For (C) and (D), metabolite tracing data are derived from one experiment. ND, not detected. Symbols represent the arithmetic mean ± SEM; p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001 (Student’s t test). See also Figure S5.
Figure 6
Figure 6
Arginase 1 Treatment Reduces Antiviral CD8 T Cell Responses and Ameliorates Virus-Induced Hepatitis during LCMV Cl13 Infection (A) IFN-γ and TNF-α production of primary murine splenic CD8 T cells cultured in medium containing 1,150 μM (standard RPMI 1640 concentration) or 11.5 μM L-arginine with or without 1,150 μM L-ornithine upon CD3/28 activation for 3 days (n = 4). (B) GP33- and NP396-specific splenic tetramer+ CD8 T cells (n = 5). (C–E) (C) GP33, (D) NP396, (E) GP276 virus-specific IFN-γ and TNF-α producing virus-specific splenic CD8 T cells upon recArg1 treatment 8 days after LCMV clone 13 infection (n = 5). (F) IFN-γ and TNF-α producing splenic GP33-specific CD8 T cells in mice treated with recArg1 twice per week up to 50 days after infection (n = 5). (G) Viral load in blood upon recArg1 treatment (n = 5). (H and I) Serum ALT (H) and AST (I) levels upon recArg1 treatment (n = 5). Data in (A)–(I) are representative for one of at least two experiments. Symbols represent the arithmetic mean ± SEM. Dotted line implicates limit of detection. p < 0.05; ∗∗p < 0.01 (Student’s t test, A–F, H, and I, or two-way ANOVA, G). See also Figure S6.

Comment in

  • Virus-Induced Interferon Regulates the Urea Cycle
    A Nishio et al. Immunity 51 (6), 975-977. PMID 31951542.
    Integrating transcriptomic, proteomic, and metabolomic data, Lercher et al. show in a mouse model of LCMV infection that type I interferon alters the expression and funct …

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