Prioritization of efforts to maintain biodiversity is an important component of conservation, but is more often applied to ecosystems or species than within species. We assessed distinctiveness among 27 populations of rainbow trout (Salmonidae: Oncorhynchus mykiss) from British Columbia, Canada, using microsatellite DNA variation (representing historical or contemporary demography) and morphology (representing adaptive variation). Standardized genetic scores, that is, the average deviation across individuals within populations from the overall genetic score generated by factorial correspondence analysis, ranged from 1.05 to 4.90 among populations. Similar standardized morphological scores, generated by principal components analysis, ranged from 1.19 to 5.35. There was little correlation between genetic and morphological distinctiveness across populations, although one population was genetically and morphologically the most distinctive. There was, however, a significant correlation (r = 0.26, P = 0.008) between microsatellite (F ST) and morphological (P ST) divergence. We combined measures of allelic richness, genetic variation within, and divergence among, populations and morphological variation to provide a conservation ranking of populations. Our approach can be combined with other measures of biodiversity value (habitat, rarity, human uses, threat status) to rationalize the prioritization of populations, especially for widespread species where geographic isolation across distinct environments promotes intraspecific variability.

Artificial breeding programs initiated to enhance the size of animal populations are often motivated by the desire to increase harvest opportunities. The introduction of non-native genotypes, however, can have negative evolutionary impacts. These may be direct, such as introgressive hybridization, or indirect via competition. Less is known about the effects of stocking with native genotypes. We assayed variation at nine microsatellite loci in 902 steelhead trout (Oncorhynchus mykiss) from five rivers in British Columbia, Canada. These samples were collected over 58 years, a time period that spanned the initiation of native steelhead trout broodstock hatchery supplementation in these rivers. We detected no changes in estimates of effective population size, genetic variation or temporal genetic structure within any population, nor of altered genetic structure among them. Genetic interactions with nonmigratory O. mykiss, the use of substantial numbers of primarily native broodstock with an approximate 1:1 male-to-female ratio, and/or poor survival and reproductive success of hatchery fish may have minimized potential genetic changes. Although no genetic changes were detected, ecological effects of hatchery programs still may influence wild population productivity and abundance. Their effects await the design and implementation of a more comprehensive evaluation program.

Gluconeogenesis is critical for maintenance of euglycemia during fasting. Elevated gluconeogenesis during type 2 diabetes (T2D) contributes to chronic hyperglycemia. Pyruvate is a major gluconeogenic substrate and requires import into the mitochondrial matrix for channeling into gluconeogenesis. Here, we demonstrate that the mitochondrial pyruvate carrier (MPC) comprising the Mpc1 and Mpc2 proteins is required for efficient regulation of hepatic gluconeogenesis. Liver-specific deletion of Mpc1 abolished hepatic MPC activity and markedly decreased pyruvate-driven gluconeogenesis and TCA cycle flux. Loss of MPC activity induced adaptive utilization of glutamine and increased urea cycle activity. Diet-induced obesity increased hepatic MPC expression and activity. Constitutive Mpc1 deletion attenuated the development of hyperglycemia induced by a high-fat diet. Acute, virally mediated Mpc1 deletion after diet-induced obesity decreased hyperglycemia and improved glucose tolerance. We conclude that the MPC is required for efficient regulation of gluconeogenesis and that the MPC contributes to the elevated gluconeogenesis and hyperglycemia in T2D.

In diverse organisms, adaptation to low oxygen (hypoxia) is mediated through complex gene expression changes that can, in part, be mimicked by exposure to metals such as cobalt. Although much is known about the transcriptional response to hypoxia and cobalt, little is known about the all-important cell metabolism effects that trigger these responses.

Macrophages are susceptible to human immunodeficiency virus type 1 (HIV-1) infection despite abundant expression of antiviral proteins. Perhaps the most important antiviral protein is the restriction factor sterile alpha motif domain and histidine/aspartic acid domain-containing protein 1 (SAMHD1). We investigated the role of SAMHD1 and its phospho-dependent regulation in the context of HIV-1 infection in primary human monocyte-derived macrophages and the ability of various interferons (IFNs) and pharmacologic agents to modulate SAMHD1. Here we show that stimulation by type I, type II, and to a lesser degree, type III interferons share activation of SAMHD1 via dephosphorylation at threonine-592 as a consequence of signaling. Cyclin-dependent kinase 1 (CDK1), a known effector kinase for SAMHD1, was downregulated at the protein level by all IFN types tested. Pharmacologic inhibition or small interfering RNA (siRNA)-mediated knockdown of CDK1 phenocopied the effects of IFN on SAMHD1. A panel of FDA-approved tyrosine kinase inhibitors potently induced activation of SAMHD1 and subsequent HIV-1 inhibition. The viral restriction imposed via IFNs or dasatinib could be overcome through depletion of SAMHD1, indicating that their effects are exerted primarily through this pathway. Our results demonstrate that SAMHD1 activation, but not transcriptional upregulation or protein induction, is the predominant mechanism of HIV-1 restriction induced by type I, type II, and type III IFN signaling in macrophages. Furthermore, SAMHD1 activation presents a pharmacologically actionable target through which HIV-1 infection can be subverted.IMPORTANCE Our experimental results demonstrate that SAMHD1 dephosphorylation at threonine-592 represents a central mechanism of HIV-1 restriction that is common to the three known families of IFNs. While IFN types I and II were potent inhibitors of HIV-1, type III IFN showed modest to undetectable activity. Regulation of SAMHD1 by IFNs involved changes in phosphorylation status but not in protein levels. Phosphorylation of SAMHD1 in macrophages occurred at least in part via CDK1. Tyrosine kinase inhibitors similarly induced SAMHD1 dephosphorylation, which protects macrophages from HIV-1 in a SAMHD1-dependent manner. SAMHD1 is a critical restriction factor regulating HIV-1 infection of macrophages.

Cells harbor two systems for fatty acid synthesis, one in the cytoplasm (catalyzed by fatty acid synthase, FASN) and one in the mitochondria (mtFAS). In contrast to FASN, mtFAS is poorly characterized, especially in higher eukaryotes, with the major product(s), metabolic roles, and cellular function(s) being essentially unknown. Here we show that hypomorphic mtFAS mutant mouse skeletal myoblast cell lines display a severe loss of electron transport chain (ETC) complexes and exhibit compensatory metabolic activities including reductive carboxylation. This effect on ETC complexes appears to be independent of protein lipoylation, the best characterized function of mtFAS, as mutants lacking lipoylation have an intact ETC. Finally, mtFAS impairment blocks the differentiation of skeletal myoblasts in vitro. Together, these data suggest that ETC activity in mammals is profoundly controlled by mtFAS function, thereby connecting anabolic fatty acid synthesis with the oxidation of carbon fuels.

Macrophages have a defined role in the pathogenesis of metabolic disease and cholesterol metabolism where alternative activation of macrophages is thought to be beneficial to both glucose and cholesterol metabolism during high fat diet induced disease. It is well established that helminth infection protects from metabolic disease, but the mechanisms underlying protection are not well understood. Here, we investigated the effects of Schistosoma mansoni infection and cytokine activation in the metabolic signatures of bone marrow derived macrophages using an approach that integrated transcriptomics, metabolomics, and lipidomics in a metabolic disease prone mouse model. We demonstrate that bone marrow derived macrophages (BMDM) from S. mansoni infected male ApoE-/- mice have dramatically increased mitochondrial respiration compared to those from uninfected mice. This change is associated with increased glucose and palmitate shuttling into TCA cycle intermediates, increased accumulation of free fatty acids, and decreased accumulation of cellular cholesterol esters, tri and diglycerides, and is dependent on mgll activity. Systemic injection of IL-4 complexes is unable to recapitulate either reductions in systemic glucose AUC or the re-programing of BMDM mitochondrial respiration seen in infected males. Importantly, the metabolic reprogramming of male myeloid cells is transferrable via bone marrow transplantation to an uninfected host, indicating maintenance of reprogramming in the absence of sustained antigen exposure. Finally, schistosome induced metabolic and bone marrow modulation is sex-dependent, with infection protecting male, but not female mice from glucose intolerance and obesity. Our findings identify a transferable, long-lasting sex-dependent reprograming of the metabolic signature of macrophages by helminth infection, providing key mechanistic insight into the factors regulating the beneficial roles of helminth infection in metabolic disease.

In addition to fatty acids, glucose and lactate are important myocardial substrates under physiologic and stress conditions. They are metabolized to pyruvate, which enters mitochondria via the mitochondrial pyruvate carrier (MPC) for citric acid cycle metabolism. In the present study, we show that MPC-mediated mitochondrial pyruvate utilization is essential for the partitioning of glucose-derived cytosolic metabolic intermediates, which modulate myocardial stress adaptation. Mice with cardiomyocyte-restricted deletion of subunit 1 of MPC (cMPC1-/-) developed age-dependent pathologic cardiac hypertrophy, transitioning to a dilated cardiomyopathy and premature death. Hypertrophied hearts accumulated lactate, pyruvate and glycogen, and displayed increased protein O-linked N-acetylglucosamine, which was prevented by increasing availability of non-glucose substrates in vivo by a ketogenic diet (KD) or a high-fat diet, which reversed the structural, metabolic and functional remodelling of non-stressed cMPC1-/- hearts. Although concurrent short-term KDs did not rescue cMPC1-/- hearts from rapid decompensation and early mortality after pressure overload, 3 weeks of a KD before transverse aortic constriction was sufficient to rescue this phenotype. Together, our results highlight the centrality of pyruvate metabolism to myocardial metabolism and function.

Mitochondrial Ca2+ transport is essential for regulating cell bioenergetics, Ca2+ signaling and cell death. Mitochondria accumulate Ca2+ via the mitochondrial Ca2+ uniporter (MCU), whereas Ca2+ is extruded by the mitochondrial Na+/Ca2+ (mtNCX) and H+/Ca2+ exchangers. The balance between these processes is essential for preventing toxic mitochondrial Ca2+ overload. Recent work demonstrated that MCU activity varies significantly among tissues, likely reflecting tissue-specific Ca2+ signaling and energy needs. It is less clear whether this diversity in MCU activity is matched by tissue-specific diversity in mitochondrial Ca2+ extrusion. Here we compared properties of mitochondrial Ca2+ extrusion in three tissues with prominent mitochondria function: brain, heart and liver. At the transcript level, expression of the Na+/Ca2+/Li+ exchanger (NCLX), which has been proposed to mediate mtNCX transport, was significantly greater in liver than in brain or heart. At the functional level, Na+ robustly activated Ca2+ efflux from brain and heart mitochondria, but not from liver mitochondria. The mtNCX inhibitor CGP37157 blocked Ca2+ efflux from brain and heart mitochondria but had no effect in liver mitochondria. Replacement of Na+ with Li+ to test the involvement of NCLX, resulted in a slowing of mitochondrial Ca2+ efflux by ∼70 %. Collectively, our findings suggest that mtNCX is responsible for Ca2+ extrusion from the mitochondria of the brain and heart, but plays only a small, if any, role in mitochondria of the liver. They also reveal that Li+ is significantly less effective than Na+ in driving mitochondrial Ca2+ efflux.

We discovered that the survival and growth of many primary acute myeloid leukemia (AML) samples and cell lines, but not normal CD34+ cells, are dependent on SIRT5, a lysine deacylase implicated in regulating multiple metabolic pathways. Dependence on SIRT5 is genotype-agnostic and extends to RAS- and p53-mutated AML. Results were comparable between SIRT5 knockdown and SIRT5 inhibition using NRD167, a potent and selective SIRT5 inhibitor. Apoptosis induced by SIRT5 disruption is preceded by reductions in oxidative phosphorylation and glutamine utilization, and an increase in mitochondrial superoxide that is attenuated by ectopic superoxide dismutase 2. These data indicate that SIRT5 controls and coordinates several key metabolic pathways in AML and implicate SIRT5 as a vulnerability in AML.

Multiple myeloma (MM) is a hematological malignancy that emerges from antibody-producing plasma B cells. Proteasome inhibitors, including the US Food and Drug Administration-approved bortezomib (BTZ) and carfilzomib (CFZ), are frequently used for the treatment of patients with MM. Nevertheless, a significant proportion of patients with MM are refractory or develop resistance to this class of inhibitors, which represents a significant challenge in the clinic. Thus, identifying factors that determine the potency of proteasome inhibitors in MM is of paramount importance to bolster their efficacy in the clinic. Using genome-wide CRISPR-based screening, we identified a subunit of the mitochondrial pyruvate carrier (MPC) complex, MPC1, as a common modulator of BTZ response in 2 distinct human MM cell lines in vitro. We noticed that CRISPR-mediated deletion or pharmacological inhibition of the MPC complex enhanced BTZ/CFZ-induced MM cell death with minimal impact on cell cycle progression. In fact, targeting the MPC complex compromised the bioenergetic capacity of MM cells, which is accompanied by reduced proteasomal activity, thereby exacerbating BTZ-induced cytotoxicity in vitro. Importantly, we observed that the RNA expression levels of several regulators of pyruvate metabolism were altered in advanced stages of MM for which they correlated with poor patient prognosis. Collectively, this study highlights the importance of the MPC complex for the survival of MM cells and their responses to proteasome inhibitors. These findings establish mitochondrial pyruvate metabolism as a potential target for the treatment of MM and an unappreciated strategy to increase the efficacy of proteasome inhibitors in the clinic.

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.

Organelle interactions play a significant role in compartmentalizing metabolism and signaling. Lipid droplets (LDs) interact with numerous organelles, including mitochondria, which is largely assumed to facilitate lipid transfer and catabolism. However, quantitative proteomics of hepatic peridroplet mitochondria (PDM) and cytosolic mitochondria (CM) reveals that CM are enriched in proteins comprising various oxidative metabolism pathways, whereas PDM are enriched in proteins involved in lipid anabolism. Isotope tracing and super-resolution imaging confirms that fatty acids (FAs) are selectively trafficked to and oxidized in CM during fasting. In contrast, PDM facilitate FA esterification and LD expansion in nutrient-replete medium. Additionally, mitochondrion-associated membranes (MAM) around PDM and CM differ in their proteomes and ability to support distinct lipid metabolic pathways. We conclude that CM and CM-MAM support lipid catabolic pathways, whereas PDM and PDM-MAM allow hepatocytes to efficiently store excess lipids in LDs to prevent lipotoxicity.

Lipid hydroperoxides (LOOH) have been implicated in skeletal muscle atrophy with age and disuse. Lysophosphatidylcholine acyltransferase 3 (LPCAT3), an enzyme of Lands cycle, conjugates a polyunsaturated fatty acyl chain to a lysophospholipid (PUFA-PL) molecule, providing substrates for LOOH propagation. Previous studies suggest that inhibition of Lands cycle is an effective strategy to suppress LOOH. Mice with skeletal muscle-specific tamoxifen-inducible knockout of LPCAT3 (LPCAT3-MKO) were utilized to determine if muscle-specific attenuation of LOOH may alleviate muscle atrophy and weakness with disuse.

Understanding the extent of interspecific hybridization and how ecological segregation may influence hybridization requires comprehensively sampling different habitats over a range of life history stages. Arctic char (Salvelinus alpinus) and Dolly Varden (S. malma) are recently diverged salmonid fishes that come into contact in several areas of the North Pacific where they occasionally hybridize. To better quantify the degree of hybridization and ecological segregation between these taxa, we sampled over 700 fish from multiple lake (littoral and profundal) and stream sites in two large, interconnected southwestern Alaskan lakes. Individuals were genotyped at 12 microsatellite markers, and genetic admixture (Q) values generated through Bayesian-based clustering revealed hybridization levels generally lower than reported in a previous study (<0.6% to 5% of samples classified as late-generation hybrids). Dolly Varden and Arctic char tended to make different use of stream habitats with the latter apparently abandoning streams for lake habitats after 2-3 years of age. Our results support the distinct biological species status of Dolly Varden and Arctic char and suggest that ecological segregation may be an important factor limiting opportunities for hybridization and/or the ecological performance of hybrid char.

Resolving the genetic population structure of species inhabiting pristine, high latitude ecosystems can provide novel insights into the post-glacial, evolutionary processes shaping the distribution of contemporary genetic variation. In this study, we assayed genetic variation in lake trout (Salvelinus namaycush) from Great Bear Lake (GBL), NT and one population outside of this lake (Sandy Lake, NT) at 11 microsatellite loci and the mtDNA control region (d-loop). Overall, population subdivision was low, but significant (global F(ST) θ = 0.025), and pairwise comparisons indicated that significance was heavily influenced by comparisons between GBL localities and Sandy Lake. Our data indicate that there is no obvious genetic structure among the various basins within GBL (global F(ST) = 0.002) despite the large geographic distances between sampling areas. We found evidence of low levels of contemporary gene flow among arms within GBL, but not between Sandy Lake and GBL. Coalescent analyses suggested that some historical gene flow occurred among arms within GBL and between GBL and Sandy Lake. It appears, therefore, that contemporary (ongoing dispersal and gene flow) and historical (historical gene flow and large founding and present-day effective population sizes) factors contribute to the lack of neutral genetic structure in GBL. Overall, our results illustrate the importance of history (e.g., post-glacial colonization) and contemporary dispersal ecology in shaping genetic population structure of Arctic faunas and provide a better understanding of the evolutionary ecology of long-lived salmonids in pristine, interconnected habitats.

Hepatocellular carcinoma (HCC) is a devastating cancer increasingly caused by non-alcoholic fatty liver disease (NAFLD). Disrupting the liver Mitochondrial Pyruvate Carrier (MPC) in mice attenuates NAFLD. Thus, we considered whether liver MPC disruption also prevents HCC. Here, we use the N-nitrosodiethylamine plus carbon tetrachloride model of HCC development to test how liver-specific MPC knock out affects hepatocellular tumorigenesis. Our data show that liver MPC ablation markedly decreases tumorigenesis and that MPC-deficient tumors transcriptomically downregulate glutathione metabolism. We observe that MPC disruption and glutathione depletion in cultured hepatomas are synthetically lethal. Stable isotope tracing shows that hepatocyte MPC disruption reroutes glutamine from glutathione synthesis into the tricarboxylic acid (TCA) cycle. These results support a model where inducing metabolic competition for glutamine by MPC disruption impairs hepatocellular tumorigenesis by limiting glutathione synthesis. These findings raise the possibility that combining MPC disruption and glutathione stress may be therapeutically useful in HCC and additional cancers.

Host defenses against pathogens are energetically expensive, leading ecological immunologists to postulate that they might participate in energetic trade-offs with other maintenance programs. However, the metabolic costs of immunity and the nature of physiologic trade-offs it engages are largely unknown. We report here that activation of immunity causes an energetic trade-off with the homeothermy (the stable maintenance of core temperature), resulting in hypometabolism and hypothermia. This immunity-induced physiologic trade-off was independent of sickness behaviors but required hematopoietic sensing of lipopolysaccharide (LPS) via the toll-like receptor 4 (TLR4). Metabolomics and genome-wide expression profiling revealed that distinct metabolic programs supported entry and recovery from the energy-conserving hypometabolic state. During bacterial infections, hypometabolic states, which could be elicited by competition for energy between maintenance programs or energy restriction, promoted disease tolerance. Together, our findings suggest that energy-conserving hypometabolic states, such as dormancy, might have evolved as a mechanism of tissue tolerance.

Lipid droplets (LDs) are frequently increased when excessive lipid accumulation leads to cellular dysfunction. Distinct from mouse β-cells, LDs are prominent in human β-cells. However, the regulation of LD mobilization (lipolysis) in human β-cells remains unclear. We found that glucose increases lipolysis in nondiabetic human islets but not in islets in patients with type 2 diabetes (T2D), indicating dysregulation of lipolysis in T2D islets. Silencing adipose triglyceride lipase (ATGL) in human pseudoislets with shRNA targeting ATGL (shATGL) increased triglycerides (TGs) and the number and size of LDs, indicating that ATGL is the principal lipase in human β-cells. In shATGL pseudoislets, biphasic glucose-stimulated insulin secretion (GSIS), and insulin secretion to 3-isobutyl-1-methylxanthine and KCl were all reduced without altering oxygen consumption rate compared with scramble control. Like human islets, INS1 cells showed visible LDs, glucose-responsive lipolysis, and impairment of GSIS after ATGL silencing. ATGL-deficient INS1 cells and human pseudoislets showed reduced SNARE protein syntaxin 1a (STX1A), a key SNARE component. Proteasomal degradation of Stx1a was accelerated likely through reduced palmitoylation in ATGL-deficient INS1 cells. Therefore, ATGL is responsible for LD mobilization in human β-cells and supports insulin secretion by stabilizing STX1A. The dysregulated lipolysis may contribute to LD accumulation and β-cell dysfunction in T2D islets.

Tumor associated macrophage responses are regulated by distinct metabolic states that affect their function. However, the ability of specific signals in the local tumor microenvironment to program macrophage metabolism remains under investigation. Here, we identify NAMPT, the rate limiting enzyme in NAD salvage synthesis, as a target of STAT1 during cellular activation by interferon gamma, an important driver of macrophage polarization and antitumor responses. We demonstrate that STAT1 occupies a conserved element within the first intron of Nampt, termed Nampt-Regulatory Element-1 (NRE1). Through disruption of NRE1 or pharmacological inhibition, a subset of M1 genes is sensitive to NAMPT activity through its impact on glycolytic processes. scRNAseq is used to profile in vivo responses by NRE1-deficient, tumor-associated leukocytes in melanoma tumors through the creation of a unique mouse strain. Reduced Nampt and inflammatory gene expression are present in specific myeloid and APC populations; moreover, targeted ablation of NRE1 in macrophage lineages results in greater tumor burden. Finally, elevated NAMPT expression correlates with IFNγ responses and melanoma patient survival. This study identifies IFN and STAT1-inducible Nampt as an important factor that shapes the metabolic program and function of tumor associated macrophages.