The lysosome is an acidic multi-functional organelle with roles in macromolecular digestion, nutrient sensing, and signaling. However, why cells require acidic lysosomes to proliferate and which ...nutrients become limiting under lysosomal dysfunction are unclear. To address this, we performed CRISPR-Cas9-based genetic screens and identified cholesterol biosynthesis and iron uptake as essential metabolic pathways when lysosomal pH is altered. While cholesterol synthesis is only necessary, iron is both necessary and sufficient for cell proliferation under lysosomal dysfunction. Remarkably, iron supplementation restores cell proliferation under both pharmacologic and genetic-mediated lysosomal dysfunction. The rescue was independent of metabolic or signaling changes classically associated with increased lysosomal pH, uncoupling lysosomal function from cell proliferation. Finally, our experiments revealed that lysosomal dysfunction dramatically alters mitochondrial metabolism and hypoxia inducible factor (HIF) signaling due to iron depletion. Altogether, these findings identify iron homeostasis as the key function of lysosomal acidity for cell proliferation.
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•Cells starve for cholesterol and iron under lysosomal dysfunction•Upon increased lysosomal pH, only iron addition enables cell proliferation•Iron does not restore lysosomal pH-related catabolic and signaling functions•Iron reverses other cellular processes related to depleted cellular iron
The lysosome is a multi-functional organelle whose low pH is required for cell viability. Weber et al. identified iron as necessary and sufficient for cell proliferation under lysosomal dysfunction. While iron addition uncouples lysosomal acidity from cell viability, iron chelation combined with lysosome-targeting compounds represents a potential cancer therapeutic strategy.
Glutathione (GSH) is a small-molecule thiol that is abundant in all eukaryotes and has key roles in oxidative metabolism
. Mitochondria, as the major site of oxidative reactions, must maintain ...sufficient levels of GSH to perform protective and biosynthetic functions
. GSH is synthesized exclusively in the cytosol, yet the molecular machinery involved in mitochondrial GSH import remains unknown. Here, using organellar proteomics and metabolomics approaches, we identify SLC25A39, a mitochondrial membrane carrier of unknown function, as a regulator of GSH transport into mitochondria. Loss of SLC25A39 reduces mitochondrial GSH import and abundance without affecting cellular GSH levels. Cells lacking both SLC25A39 and its paralogue SLC25A40 exhibit defects in the activity and stability of proteins containing iron-sulfur clusters. We find that mitochondrial GSH import is necessary for cell proliferation in vitro and red blood cell development in mice. Heterologous expression of an engineered bifunctional bacterial GSH biosynthetic enzyme (GshF) in mitochondria enables mitochondrial GSH production and ameliorates the metabolic and proliferative defects caused by its depletion. Finally, GSH availability negatively regulates SLC25A39 protein abundance, coupling redox homeostasis to mitochondrial GSH import in mammalian cells. Our work identifies SLC25A39 as an essential and regulated component of the mitochondrial GSH-import machinery.
Mitochondria must maintain adequate amounts of metabolites for protective and biosynthetic functions. However, how mitochondria sense the abundance of metabolites and regulate metabolic homeostasis ...is not well understood. In this work, we focused on glutathione (GSH), a critical redox metabolite in mitochondria, and identified a feedback mechanism that controls its abundance through the mitochondrial GSH transporter, SLC25A39. Under physiological conditions, SLC25A39 is rapidly degraded by mitochondrial protease AFG3L2. Depletion of GSH dissociates AFG3L2 from SLC25A39, causing a compensatory increase in mitochondrial GSH uptake. Genetic and proteomic analyses identified a putative iron-sulfur cluster in the matrix-facing loop of SLC25A39 as essential for this regulation, coupling mitochondrial iron homeostasis to GSH import. Altogether, our work revealed a paradigm for the autoregulatory control of metabolic homeostasis in organelles.
Summary Background Cancer is a major cause of death in children worldwide, and the recorded incidence tends to increase with time. Internationally comparable data on childhood cancer incidence in the ...past two decades are scarce. This study aimed to provide internationally comparable local data on the incidence of childhood cancer to promote research of causes and implementation of childhood cancer control. Methods This population-based registry study, devised by the International Agency for Research on Cancer in collaboration with the International Association of Cancer Registries, collected data on all malignancies and non-malignant neoplasms of the CNS diagnosed before age 20 years in populations covered by high-quality cancer registries with complete data for 2001–10. Incidence rates per million person-years for the 0–14 years and 0–19 years age groups were age-adjusted using the world standard population to provide age-standardised incidence rates (WSRs), using the age-specific incidence rates (ASR) for individual age groups (0–4 years, 5–9 years, 10–14 years, and 15–19 years). All rates were reported for 19 geographical areas or ethnicities by sex, age group, and cancer type. The regional WSRs for children aged 0–14 years were compared with comparable data obtained in the 1980s. Findings Of 532 invited cancer registries, 153 registries from 62 countries, departments, and territories met quality standards, and contributed data for the entire decade of 2001–10. 385 509 incident cases in children aged 0–19 years occurring in 2·64 billion person-years were included. The overall WSR was 140·6 per million person-years in children aged 0–14 years (based on 284 649 cases), and the most common cancers were leukaemia (WSR 46·4), followed by CNS tumours (WSR 28·2), and lymphomas (WSR 15·2). In children aged 15–19 years (based on 100 860 cases), the ASR was 185·3 per million person-years, the most common being lymphomas (ASR 41·8) and the group of epithelial tumours and melanoma (ASR 39·5). Incidence varied considerably between and within the described regions, and by cancer type, sex, age, and racial and ethnic group. Since the 1980s, the global WSR of registered cancers in children aged 0–14 years has increased from 124·0 (95% CI 123·3–124·7) to 140·6 (140·1–141·1) per million person-years. Interpretation This unique global source of childhood cancer incidence will be used for aetiological research and to inform public health policy, potentially contributing towards attaining several targets of the Sustainable Development Goals. The observed geographical, racial and ethnic, age, sex, and temporal variations require constant monitoring and research. Funding International Agency for Research on Cancer and the Union for International Cancer Control.
Abstract Objectives Delusional parasitosis is a syndrome with which most infectious diseases physicians and microbiologists are familiar. However, little is known about the epidemiology of this ...disorder, and most reports consist of case reports or small series. We conducted a case series with long-term questionnaire follow-up of delusional parasitosis patients presenting to our academic medical center. Methods From 1994 through 1999, 23 patients with primary delusional parasitosis (as defined by DSM IV criteria) were identified through the Infectious Diseases Clinic or Emergency Room at the University of Washington and cases were analyzed for prospectively established demographic, clinical, and social variables of interest. Results Of 23 patients, 15 were women and eight were men. Mean duration (± standard deviation) of symptoms was 2.6 ± 2.8 years (median 1.5), with shorter duration related to improved prognosis. Mean symptom duration in women was 3.1 ± 3.0 years versus 1.5 ± 1.5 years in men. Patients saw an average of six physicians before presenting to our center. Reduced social interactions were common, but employment affect was not significant. Treatment with pimozide or gabapentin combined with antidepressants appeared to be effective in some cases. Conclusions This is the largest study of primary delusional parasitosis originating from an academic medical center, and highlights the burden of disease borne by patients and the healthcare system.
With the Reliability Pricing Model (RPM), PJM capacity market provides locational capacity pricing to recognize and quantify the locational value of resource capacity to meet the reliability ...requirements. In September 2012 for the 2014/2015 PJM first Incremental Auction (IA), in addition to the existing Limited Demand Resource (DR) product, two new DR products with fewer limitations need to be modeled: Annual (ANL) DR and Extended Summer (ES) DR. Meanwhile, the buy bids also have options to pay by the demand quality. The RPM IA clearing needs to recognize and model the offers and bids with various quality types, and ensure the clearing results to comply with the market rules. The DR integrated RPM engine has been used to clear PJM IA in September 2012. The integration of DR model into RPM IA model is presented in this paper. Impacts of the capacity requirement enforcement on the market clearing solution and prices are analyzed in details.
A structure-based model describing the interaction of the two-domain PI- Sce I endonuclease with its 31-base pair DNA substrate suggests that the endonuclease domain (domain II) contacts the cleavage
...site region of the substrate, while the protein splicing domain (domain I) interacts with a distal region that is sufficient
for high affinity binding. To support this model, alanine-scanning mutagenesis was used to assemble a set of 49 PI- Sce I mutant proteins that were purified and assayed for their DNA binding and cleavage properties. Fourteen mutant proteins were
4- to >500-fold less active than wild-type PI- Sce I in cleavage assays, and one mutant (T225A) was 3-fold more active. Alanine substitution at two positions in domain I reduces
overall binding >60-fold by perturbing the interaction of PI- Sce I with the minimal binding region. Conversely, mutations in domain II have little effect on binding, reduce binding to the
cleavage site region only, or affect binding to both regions. Interestingly, substitutions at Lys 301 , which is part of the endonucleolytic active site, eliminate binding to the cleavage site region but permit contact with
the minimal binding region. This experimental evidence demonstrates that the protein splicing domain as well as the endonuclease
domain is involved in binding of a DNA substrate with the requisite length.
A hallmark of pancreatic ductal adenocarcinoma (PDAC) is an exuberant stroma comprised of diverse cell types that enable or suppress tumor progression. Here, we explored the role of oncogenic KRAS in ...protumorigenic signaling interactions between cancer cells and host cells. We show that KRAS mutation (KRAS*) drives cell-autonomous expression of type I cytokine receptor complexes (IL2rγ-IL4rα and IL2rγ-IL13rα1) in cancer cells that in turn are capable of receiving cytokine growth signals (IL4 or IL13) provided by invading Th2 cells in the microenvironment. Early neoplastic lesions show close proximity of cancer cells harboring KRAS* and Th2 cells producing IL4 and IL13. Activated IL2rγ-IL4rα and IL2rγ-IL13rα1 receptors signal primarily via JAK1-STAT6. Integrated transcriptomic, chromatin occupancy, and metabolomic studies identified MYC as a direct target of activated STAT6 and that MYC drives glycolysis. Thus, paracrine signaling in the tumor microenvironment plays a key role in the KRAS*-driven metabolic reprogramming of PDAC. SIGNIFICANCE: Type II cytokines, secreted by Th2 cells in the tumor microenvironment, can stimulate cancer cell-intrinsic MYC transcriptional upregulation to drive glycolysis. This KRAS*-driven heterotypic signaling circuit in the early and advanced tumor microenvironment enables cooperative protumorigenic interactions, providing candidate therapeutic targets in the KRAS* pathway for this intractable disease.
Cancer cells rewire their metabolism and rely on endogenous antioxidants to mitigate lethal oxidative damage to lipids. However, the metabolic processes that modulate the response to lipid ...peroxidation are poorly defined. Using genetic screens, we compared metabolic genes essential for proliferation upon inhibition of cystine uptake or glutathione peroxidase-4 (GPX4). Interestingly, very few genes were commonly required under both conditions, suggesting that cystine limitation and GPX4 inhibition may impair proliferation via distinct mechanisms. Our screens also identify tetrahydrobiopterin (BH4) biosynthesis as an essential metabolic pathway upon GPX4 inhibition. Mechanistically, BH4 is a potent radical-trapping antioxidant that protects lipid membranes from autoxidation, alone and in synergy with vitamin E. Dihydrofolate reductase catalyzes the regeneration of BH4, and its inhibition by methotrexate synergizes with GPX4 inhibition. Altogether, our work identifies the mechanism by which BH4 acts as an endogenous antioxidant and provides a compendium of metabolic modifiers of lipid peroxidation.
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