Sterol Oxidation Mediates Stress-Responsive Vms1 Translocation to Mitochondria.

Vms1 translocates to damaged mitochondria in response to stress, whereupon its binding partner, Cdc48, contributes to mitochondrial protein homeostasis. Mitochondrial targeting of Vms1 is mediated by its conserved mitochondrial targeting domain (MTD), which, in unstressed conditions, is inhibited by intramolecular binding to the Vms1 leucine-rich sequence (LRS). Here, we report a 2.7 Å crystal structure of Vms1 that reveals that the LRS lies in a hydrophobic groove in the autoinhibited MTD. We also demonstrate that the oxidized sterol, ergosterol peroxide, is necessary and sufficient for Vms1 localization to mitochondria, through binding the MTD in an interaction that is competitive with binding to the LRS. These data support a model in which stressed mitochondria generate an oxidized sterol receptor that recruits Vms1 to support mitochondrial protein homeostasis.

IRE1α regulates macrophage polarization, PD-L1 expression, and tumor survival.

In the tumor microenvironment, local immune dysregulation is driven in part by macrophages and dendritic cells that are polarized to a mixed proinflammatory/immune-suppressive phenotype. The unfolded protein response (UPR) is emerging as the possible origin of these events. Here we report that the inositol-requiring enzyme 1 (IRE1α) branch of the UPR is directly involved in the polarization of macrophages in vitro and in vivo, including the up-regulation of interleukin 6 (IL-6), IL-23, Arginase1, as well as surface expression of CD86 and programmed death ligand 1 (PD-L1). Macrophages in which the IRE1α/X-box binding protein 1 (Xbp1) axis is blocked pharmacologically or deleted genetically have significantly reduced polarization and CD86 and PD-L1 expression, which was induced independent of IFNγ signaling, suggesting a novel mechanism in PD-L1 regulation in macrophages. Mice with IRE1α- but not Xbp1-deficient macrophages showed greater survival than controls when implanted with B16.F10 melanoma cells. Remarkably, we found a significant association between the IRE1α gene signature and CD274 gene expression in tumor-infiltrating macrophages in humans. RNA sequencing (RNASeq) analysis showed that bone marrow-derived macrophages with IRE1α deletion lose the integrity of the gene connectivity characteristic of regulated IRE1α-dependent decay (RIDD) and the ability to activate CD274 gene expression. Thus, the IRE1α/Xbp1 axis drives the polarization of macrophages in the tumor microenvironment initiating a complex immune dysregulation leading to failure of local immune surveillance.

The unfolded protein response links tumor aneuploidy to local immune dysregulation.

Aneuploidy is a chromosomal abnormality associated with poor prognosis in many cancer types. Here, we tested the hypothesis that the unfolded protein response (UPR) mechanistically links aneuploidy and local immune dysregulation. Using a single somatic copy number alteration (SCNA) score inclusive of whole-chromosome, chromosome arm, and focal alterations in a pan-cancer analysis of 9,375 samples in The Cancer Genome Atlas (TCGA) database, we found an inverse correlation with a cytotoxicity (CYT) score across disease stages. Co-expression patterns of UPR genes changed substantially between SCNAlow and SCNAhigh groups. Pathway activity scores showed increased activity of multiple branches of the UPR in response to aneuploidy. The PERK branch showed the strongest association with a reduced CYT score. The conditioned medium of aneuploid cells transmitted XBP1 splicing and caused IL-6 and arginase 1 transcription in receiver bone marrow-derived macrophages and markedly diminished the production of IFN-γ and granzyme B in activated human T cells. We propose the UPR as a mechanistic link between aneuploidy and immune dysregulation in the tumor microenvironment.

Associations of sex-related steroid hormones and proteins with alcohol dependence: A United Kingdom Biobank study.

BACKGROUND: Sex-related steroid hormones and proteins may contribute to the sex differences in the characteristics and health consequences of alcohol use disorder. This study aimed to examine the associations between alcohol dependence (AD) and sex-related hormones/proteins using a population-based dataset. METHODS: We retrieved serum total testosterone (TT) and estradiol (TE2), sex hormone binding globulin (SHBG), and albumin levels along with clinical data from the UK Biobank. Hormone/protein levels were compared between AD (lifetime AD and/or related diagnoses; 2218 males; 682 females) and control (no aforementioned diagnoses and AUDIT<8; 198,058 males; 250,830 females) groups with sex-dependent linear regression models adjusting for age and body mass index. Moderation and mediation analyses were performed to test whether SHBG was a moderator and/or mediator between hormones and AD or current drinking. RESULTS: AD males had higher TT, TE2, and SHBG levels but lower bioavailable testosterone, bioavailable estradiol, and albumin levels than controls (padjusted<0.001). After adjusting for menopause, AD females had higher TT and lower albumin levels than controls (padjusted<0.001). These differences remained after accounting for current drinking frequency (p < 0.001). SHBG moderated TT's effect on AD in males (pinteraction<0.001). SHBG was a positive mediator between TT and AD in both sexes and between TE2 and AD in males (p < 0.001), but a negative mediator between TT and current drinking in controls (both sexes) and AD males (p < 0.001). CONCLUSIONS: Testosterone and estradiol levels are altered in males and females with AD distinctly regardless of current drinking frequency. SHBG may play a critical role in these associations.

Metaboverse enables automated discovery and visualization of diverse metabolic regulatory patterns.

Metabolism is intertwined with various cellular processes, including controlling cell fate, influencing tumorigenesis, participating in stress responses and more. Metabolism is a complex, interdependent network, and local perturbations can have indirect effects that are pervasive across the metabolic network. Current analytical and technical limitations have long created a bottleneck in metabolic data interpretation. To address these shortcomings, we developed Metaboverse, a user-friendly tool to facilitate data exploration and hypothesis generation. Here we introduce algorithms that leverage the metabolic network to extract complex reaction patterns from data. To minimize the impact of missing measurements within the network, we introduce methods that enable pattern recognition across multiple reactions. Using Metaboverse, we identify a previously undescribed metabolite signature that correlated with survival outcomes in early stage lung adenocarcinoma patients. Using a yeast model, we identify metabolic responses suggesting an adaptive role of citrate homeostasis during mitochondrial dysfunction facilitated by the citrate transporter, Ctp1. We demonstrate that Metaboverse augments the user's ability to extract meaningful patterns from multi-omics datasets to develop actionable hypotheses.

Compartment and hub definitions tune metabolic networks for metabolomic interpretations.

BACKGROUND: Metabolic networks represent all chemical reactions that occur between molecular metabolites in an organism's cells. They offer biological context in which to integrate, analyze, and interpret omic measurements, but their large scale and extensive connectivity present unique challenges. While it is practical to simplify these networks by placing constraints on compartments and hubs, it is unclear how these simplifications alter the structure of metabolic networks and the interpretation of metabolomic experiments. RESULTS: We curated and adapted the latest systemic model of human metabolism and developed customizable tools to define metabolic networks with and without compartmentalization in subcellular organelles and with or without inclusion of prolific metabolite hubs. Compartmentalization made networks larger, less dense, and more modular, whereas hubs made networks larger, more dense, and less modular. When present, these hubs also dominated shortest paths in the network, yet their exclusion exposed the subtler prominence of other metabolites that are typically more relevant to metabolomic experiments. We applied the non-compartmental network without metabolite hubs in a retrospective, exploratory analysis of metabolomic measurements from 5 studies on human tissues. Network clusters identified individual reactions that might experience differential regulation between experimental conditions, several of which were not apparent in the original publications. CONCLUSIONS: Exclusion of specific metabolite hubs exposes modularity in both compartmental and non-compartmental metabolic networks, improving detection of relevant clusters in omic measurements. Better computational detection of metabolic network clusters in large data sets has potential to identify differential regulation of individual genes, transcripts, and proteins.

Control of intestinal stem cell function and proliferation by mitochondrial pyruvate metabolism.

Most differentiated cells convert glucose to pyruvate in the cytosol through glycolysis, followed by pyruvate oxidation in the mitochondria. These processes are linked by the mitochondrial pyruvate carrier (MPC), which is required for efficient mitochondrial pyruvate uptake. In contrast, proliferative cells, including many cancer and stem cells, perform glycolysis robustly but limit fractional mitochondrial pyruvate oxidation. We sought to understand the role this transition from glycolysis to pyruvate oxidation plays in stem cell maintenance and differentiation. Loss of the MPC in Lgr5-EGFP-positive stem cells, or treatment of intestinal organoids with an MPC inhibitor, increases proliferation and expands the stem cell compartment. Similarly, genetic deletion of the MPC in Drosophila intestinal stem cells also increases proliferation, whereas MPC overexpression suppresses stem cell proliferation. These data demonstrate that limiting mitochondrial pyruvate metabolism is necessary and sufficient to maintain the proliferation of intestinal stem cells.