bims-cagime Biomed News
on Cancer, aging and metabolism
Issue of 2020‒07‒12
fifty-four papers selected by
Kıvanç Görgülü
Technical University of Munich

  1. J Exp Med. 2020 Sep 07. pii: e20192389. [Epub ahead of print]217(9):
    Oni TE, Biffi G, Baker LA, Hao Y, Tonelli C, Somerville TDD, Deschênes A, Belleau P, Hwang CI, Sánchez-Rivera FJ, Cox H, Brosnan E, Doshi A, Lumia RP, Khaledi K, Park Y, Trotman LC, Lowe SW, Krasnitz A, Vakoc CR, Tuveson DA.
      Pancreatic ductal adenocarcinoma (PDAC) has a dismal prognosis, and new therapies are needed. Altered metabolism is a cancer vulnerability, and several metabolic pathways have been shown to promote PDAC. However, the changes in cholesterol metabolism and their role during PDAC progression remain largely unknown. Here we used organoid and mouse models to determine the drivers of altered cholesterol metabolism in PDAC and the consequences of its disruption on tumor progression. We identified sterol O-acyltransferase 1 (SOAT1) as a key player in sustaining the mevalonate pathway by converting cholesterol to inert cholesterol esters, thereby preventing the negative feedback elicited by unesterified cholesterol. Genetic targeting of Soat1 impairs cell proliferation in vitro and tumor progression in vivo and reveals a mevalonate pathway dependency in p53 mutant PDAC cells that have undergone p53 loss of heterozygosity (LOH). In contrast, pancreatic organoids lacking p53 mutation and p53 LOH are insensitive to SOAT1 loss, indicating a potential therapeutic window for inhibiting SOAT1 in PDAC.
  2. Elife. 2020 Jul 10. pii: e56782. [Epub ahead of print]9
    Lau AN, Li Z, Danai LV, Westermark AM, Darnell AM, Ferreira R, Gocheva V, Sivanand S, Lien EC, Sapp KM, Mayers JR, Biffi G, Chin CR, Davidson SM, Tuveson DA, Jacks T, Matheson NJ, Yilmaz O, Vander Heiden MG.
      Tumors are composed of many different cell types including cancer cells, fibroblasts, and immune cells. Dissecting functional metabolic differences between cell types within a mixed population can be challenging due to the rapid turnover of metabolites relative to the time needed to isolate cells. To overcome this challenge, we traced isotope-labeled nutrients into macromolecules that turn over more slowly than metabolites. This approach was used to assess differences between cancer cell and fibroblast metabolism in murine pancreatic cancer organoid-fibroblast co-cultures and tumors. Pancreatic cancer cells exhibited increased pyruvate carboxylation relative to fibroblasts, and this flux depended on both pyruvate carboxylase and malic enzyme 1 activity. Consequently, expression of both enzymes in cancer cells was necessary for organoid and tumor growth, demonstrating that dissecting the metabolism of specific cell populations within heterogeneous systems can identify dependencies that may not be evident from studying isolated cells in culture or bulk tissue.
    Keywords:  cancer biology; mouse
  3. Trends Biochem Sci. 2020 Jul 02. pii: S0968-0004(20)30151-1. [Epub ahead of print]
    Zhao Q, Gao SM, Wang MC.
      Lysosomes transcend the role of degradation stations, acting as key nodes for interorganelle crosstalk and signal transduction. Lysosomes communicate with the nucleus through physical proximity and functional interaction. In response to external and internal stimuli, lysosomes actively adjust their distribution between peripheral and perinuclear regions and modulate lysosome-nucleus signaling pathways; in turn, the nucleus fine-tunes lysosomal biogenesis and functions through transcriptional controls. Changes in coordination between these two essential organelles are associated with metabolic disorders, neurodegenerative diseases, and aging. In this review, we address recent advances in lysosome-nucleus communication by multi-tiered regulatory mechanisms and discuss how these regulations couple metabolic inputs with organellar motility, cellular signaling, and transcriptional network.
    Keywords:  lysosomal adaptation; lysosomal metabolites; lysosome positioning; lysosome-to-nucleus signaling; transcription factors
  4. Cancers (Basel). 2020 Jul 04. pii: E1790. [Epub ahead of print]12(7):
    Cash TP, Alcalá S, Rico-Ferreira MDR, Hernández-Encinas E, García J, Albarrán MI, Valle S, Muñoz J, Martínez-González S, Blanco-Aparicio C, Pastor J, Serrano M, Sainz B.
      Despite significant efforts to improve pancreatic ductal adenocarcinoma (PDAC) clinical outcomes, overall survival remains dismal. The poor response to current therapies is partly due to the existence of pancreatic cancer stem cells (PaCSCs), which are efficient drivers of PDAC tumorigenesis, metastasis and relapse. To find new therapeutic agents that could efficiently kill PaCSCs, we screened a chemical library of 680 compounds for candidate small molecules with anti-CSC activity, and identified two compounds of a specific chemical series with potent activity in vitro and in vivo against patient-derived xenograft (PDX) cultures. The anti-CSC mechanism of action of this specific chemical series was found to rely on induction of lysosomal membrane permeabilization (LMP), which is likely associated with the increased lysosomal mass observed in PaCSCs. Using the well characterized LMP-inducer siramesine as a tool molecule, we show elimination of the PaCSC population in mice implanted with tumors from two PDX models. Collectively, our approach identified lysosomal disruption as a promising anti-CSC therapeutic strategy for PDAC.
    Keywords:  cancer stem cells; compound library; lysosomal membrane permeabilization; pancreatic ductal adenocarcinoma; patient-derived xenografts
  5. Mol Cell Proteomics. 2020 Jul 10. pii: mcp.RA120.002046. [Epub ahead of print]
    Wiechmann S, Saupp E, Schilling D, Heinzlmeir S, Schneider G, Schmid RM, Combs SE, Kuster B, Dobiasch S.
      Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive cancers and known for its extensive genetic heterogeneity, high therapeutic resistance and strong variation in intrinsic radiosensitivity. In order to understand the molecular mechanisms underlying radioresistance, we screened the phenotypic response of 38 PDAC cell lines to ionizing radiation. Subsequent phosphoproteomic analysis of two representative sensitive and resistant lines led to the reproducible identification of 7,800 proteins and 13,000 phosphorylation sites (p-sites). Approximately 700 p-sites on 400 proteins showed abundance changes after radiation in all cell lines regardless of their phenotypic sensitivity. Apart from recapitulating known radiation response phosphorylation markers such as on proteins involved in DNA damage repair, the analysis uncovered many novel members of a radiation-responsive signaling network that was apparent only at the level of protein phosphorylation. These regulated p-sites were enriched in potential ATM substrates and in-vitro kinase assays corroborated 10 of these. Comparing the proteomes and phosphoproteomes of radiosensitive and -resistant cells pointed to additional tractable radioresistance mechanisms involving apoptotic proteins. For instance, elevated NADPH quinine oxidoreductase 1 (NQO1) expression in radioresistant cells may aid in clearing harmful reactive oxygen species. Resistant cells also showed elevated phosphorylation levels of proteins involved in cytoskeleton organization including actin dynamics and focal adhesion kinase (FAK) activity and one resistant cell line showed a strong migration phenotype. Pharmacological inhibition of the kinases FAK by Defactinib and of CHEK1 by Rabusertib showed a statistically significant sensitization to radiation in radioresistant PDAC cells. Together, the presented data map a comprehensive molecular network of radiation-induced signaling, improves the understanding of radioresistance and provides avenues for developing radiotherapeutic strategies.
    Keywords:  Cancer therapeutics; Enzyme Inhibition*; Kinases*; Pancreatic cancer; Phosphoproteome; kinase inhibitors; kinase substrates; radioresistance
  6. Sci Adv. 2020 Jun;6(26): eaaz9805
    Higuchi-Sanabria R, Shen K, Kelet N, Frankino PA, Durieux J, Bar-Ziv R, Sing CN, Garcia EJ, Homentcovschi S, Sanchez M, Wu R, Tronnes SU, Joe L, Webster B, Ahilon-Jeronimo A, Monshietehadi S, Dallarda S, Pender C, Pon LA, Zoncu R, Dillin A.
      Recent work has highlighted the fact that lysosomes are a critical signaling hub of metabolic processes, providing fundamental building blocks crucial for anabolic functions. How lysosomal functions affect other cellular compartments is not fully understood. Here, we find that lysosomal recycling of the amino acids lysine and arginine is essential for proper ER quality control through the UPRER. Specifically, loss of the lysine and arginine amino acid transporter LAAT-1 results in increased sensitivity to proteotoxic stress in the ER and decreased animal physiology. We find that these LAAT-1-dependent effects are linked to glycine metabolism and transport and that the loss of function of the glycine transporter SKAT-1 also increases sensitivity to ER stress. Direct lysine and arginine supplementation, or glycine supplementation alone, can ameliorate increased ER stress sensitivity found in laat-1 mutants. These data implicate a crucial role in recycling lysine, arginine, and glycine in communication between the lysosome and ER.
  7. Proc Natl Acad Sci U S A. 2020 Jul 09. pii: 202006238. [Epub ahead of print]
    Peters CH, Myers ME, Juchno J, Haimbaugh C, Bichraoui H, Du Y, Bankston JR, Walker LA, Proenza C.
      Ion channels in excitable cells function in macromolecular complexes in which auxiliary proteins modulate the biophysical properties of the pore-forming subunits. Hyperpolarization-activated, cyclic nucleotide-sensitive HCN4 channels are critical determinants of membrane excitability in cells throughout the body, including thalamocortical neurons and cardiac pacemaker cells. We previously showed that the properties of HCN4 channels differ dramatically in different cell types, possibly due to the endogenous expression of auxiliary proteins. Here, we report the discovery of a family of endoplasmic reticulum (ER) transmembrane proteins that associate with and modulate HCN4. Lymphoid-restricted membrane protein (LRMP, Jaw1) and inositol trisphosphate receptor-associated guanylate kinase substrate (IRAG, Mrvi1, and Jaw1L) are homologous proteins with small ER luminal domains and large cytoplasmic domains. Despite their homology, LRMP and IRAG have distinct effects on HCN4. LRMP is a loss-of-function modulator that inhibits the canonical depolarizing shift in the voltage dependence of HCN4 in response to the binding of cAMP. In contrast, IRAG causes a gain of HCN4 function by depolarizing the basal voltage dependence in the absence of cAMP. The mechanisms of action of LRMP and IRAG are independent of trafficking and cAMP binding, and they are specific to the HCN4 isoform. We also found that IRAG is highly expressed in the mouse sinoatrial node where computer modeling predicts that its presence increases HCN4 current. Our results suggest important roles for LRMP and IRAG in the regulation of cellular excitability, as tools for advancing mechanistic understanding of HCN4 channel function, and as possible scaffolds for coordination of signaling pathways.
    Keywords:  HCN channel; IRAG; LRMP; ion channel; sinoatrial node
  8. Science. 2020 Jul 10. 369(6500): 167-173
    Horowitz AM, Fan X, Bieri G, Smith LK, Sanchez-Diaz CI, Schroer AB, Gontier G, Casaletto KB, Kramer JH, Williams KE, Villeda SA.
      Reversing brain aging may be possible through systemic interventions such as exercise. We found that administration of circulating blood factors in plasma from exercised aged mice transferred the effects of exercise on adult neurogenesis and cognition to sedentary aged mice. Plasma concentrations of glycosylphosphatidylinositol (GPI)-specific phospholipase D1 (Gpld1), a GPI-degrading enzyme derived from liver, were found to increase after exercise and to correlate with improved cognitive function in aged mice, and concentrations of Gpld1 in blood were increased in active, healthy elderly humans. Increasing systemic concentrations of Gpld1 in aged mice ameliorated age-related regenerative and cognitive impairments by altering signaling cascades downstream of GPI-anchored substrate cleavage. We thus identify a liver-to-brain axis by which blood factors can transfer the benefits of exercise in old age.
  9. Cancer Cell. 2020 Jun 23. pii: S1535-6108(20)30274-9. [Epub ahead of print]
    Hayes JD, Dinkova-Kostova AT, Tew KD.
      Contingent upon concentration, reactive oxygen species (ROS) influence cancer evolution in apparently contradictory ways, either initiating/stimulating tumorigenesis and supporting transformation/proliferation of cancer cells or causing cell death. To accommodate high ROS levels, tumor cells modify sulfur-based metabolism, NADPH generation, and the activity of antioxidant transcription factors. During initiation, genetic changes enable cell survival under high ROS levels by activating antioxidant transcription factors or increasing NADPH via the pentose phosphate pathway (PPP). During progression and metastasis, tumor cells adapt to oxidative stress by increasing NADPH in various ways, including activation of AMPK, the PPP, and reductive glutamine and folate metabolism.
    Keywords:  AP-1; BACH1; FOXO; HIF-1alpha; HSF1; NADPH generation; NF-κB; NRF2; PGC-1alpha; TP53; adaptation; antioxidant; dormant cancer cell; folate metabolism; glutathione; initiation; metastasis; oxidative stress; pentose phosphate pathway; progression; reactive oxygen species; recurrent disease; redox signaling; reductive glutamine metabolism; thioredoxin; tumorigenesis
  10. Nat Rev Cancer. 2020 Jul 06.
    Leone RD, Powell JD.
      Through the successes of checkpoint blockade and adoptive cellular therapy, immunotherapy has become an established treatment modality for cancer. Cellular metabolism has emerged as a critical determinant of the viability and function of both cancer cells and immune cells. In order to sustain prodigious anabolic needs, tumours employ a specialized metabolism that differs from untransformed somatic cells. This metabolism leads to a tumour microenvironment that is commonly acidic, hypoxic and/or depleted of critical nutrients required by immune cells. In this context, tumour metabolism itself is a checkpoint that can limit immune-mediated tumour destruction. Because our understanding of immune cell metabolism and cancer metabolism has grown significantly in the past decade, we are on the cusp of being able to unravel the interaction of cancer cell metabolism and immune metabolism in therapeutically meaningful ways. Although there are metabolic processes that are seemingly fundamental to both cancer and responding immune cells, metabolic heterogeneity and plasticity may serve to distinguish the two. As such, understanding the differential metabolic requirements of the diverse cells that comprise an immune response to cancer offers an opportunity to selectively regulate immune cell function. Such a nuanced evaluation of cancer and immune metabolism can uncover metabolic vulnerabilities and therapeutic windows upon which to intervene for enhanced immunotherapy.
  11. Aging Cell. 2020 Jul 06. e13187
    Liang W, Moyzis AG, Lampert MA, Diao RY, Najor RH, Gustafsson ÅB.
      Advancing age is a major risk factor for developing heart disease, and the biological processes contributing to aging are currently under intense investigation. Autophagy is an important cellular quality control mechanism that is reduced in tissues with age but the molecular mechanisms underlying the age-associated defects in autophagy remain poorly characterized. Here, we have investigated how the autophagic process is altered in aged mouse hearts. We report that autophagic activity is reduced in aged hearts due to a reduction in autophagosome formation. Gene expression profile analysis to evaluate changes in autophagy regulators uncovered a reduction in Atg9b transcript and protein levels. Atg9 proteins are critical in delivering membrane to the growing autophagosome, and siRNA knockdown of Atg9b in cells confirmed a reduction in autophagosome formation. Autophagy is also the main pathway involved in eliminating dysfunctional mitochondria via a process known as mitophagy. The E3 ubiquitin ligase Parkin plays a key role in labeling mitochondria for mitophagy. We also found increased levels of Parkin-positive mitochondria in the aged hearts, an indication that they have been labeled for mitophagy. In contrast, Nrf1, a major transcriptional regulator of mitochondrial biogenesis, was significantly reduced in aged hearts. Additionally, our data showed reduced Drp1-mediated mitochondrial fission and formation of enlarged mitochondria in the aged heart. Overall, our findings suggest that cardiac aging is associated with reduced autophagosome number, decreased mitochondrial turnover, and formation of megamitochondria.
    Keywords:  Atg9; Parkin; aging; autophagy; heart; mitochondria; mitophagy
  12. Proc Natl Acad Sci U S A. 2020 Jul 06. pii: 202008030. [Epub ahead of print]
    Bhargava HK, Tabata K, Byck JM, Hamasaki M, Farrell DP, Anishchenko I, DiMaio F, Im YJ, Yoshimori T, Hurley JH.
      Rubicon is a potent negative regulator of autophagy and a potential target for autophagy-inducing therapeutics. Rubicon-mediated inhibition of autophagy requires the interaction of the C-terminal Rubicon homology (RH) domain of Rubicon with Rab7-GTP. Here we report the 2.8-Å crystal structure of the Rubicon RH domain in complex with Rab7-GTP. Our structure reveals a fold for the RH domain built around four zinc clusters. The switch regions of Rab7 insert into pockets on the surface of the RH domain in a mode that is distinct from those of other Rab-effector complexes. Rubicon residues at the dimer interface are required for Rubicon and Rab7 to colocalize in living cells. Mutation of Rubicon RH residues in the Rab7-binding site restores efficient autophagic flux in the presence of overexpressed Rubicon, validating the Rubicon RH domain as a promising therapeutic target.
    Keywords:  Rab GTPase; autophagy; crystal structure
  13. JCI Insight. 2020 Jul 07. pii: 138290. [Epub ahead of print]
    Le Large TY, Mantini G, Meijer LL, Pham TV, Funel N, van Grieken NC, Kok B, Knol JC, van Laarhoven HWM, Piersma SR, Jimenez CR, Kazemier G, Giovannetti E, Bijlsma MF.
      Pancreatic ductal adenocarcinoma (PDAC) is characterized by a relative paucity of cancer cells that are surrounded by an abundance of non-tumor cells and extracellular matrix, known as stroma. The interaction between stroma and cancer cells contributes to poor outcome, but how proteins from these individual compartments drive aggressive tumor behavior is not known. Here, we report the proteomic analysis of laser-capture microdissected (LCM) PDAC samples. We isolated stroma, tumor, and bulk samples from a cohort with long- and short-term survivors. Compartment-specific proteins were measured by mass spectrometry, yielding the largest PDAC proteome landscape to date. These analyses revealed that in bulk analysis, tumor-derived proteins were typically masked and that LCM was required to reveal biology and novel prognostic markers. We validated tumor CALB2 and stromal COL11A1 expression as compartment-specific prognostic markers. We identified and functionally addressed the contributions of the tumor cell receptor EPHA2 to tumor cell viability and motility, underscoring the value of compartment-specific protein analysis in PDAC.
    Keywords:  Cancer; Cell Biology; Molecular biology; Oncology; Proteomics
  14. Nat Rev Clin Oncol. 2020 Jul 06.
    Dasari A, Morris VK, Allegra CJ, Atreya C, Benson AB, Boland P, Chung K, Copur MS, Corcoran RB, Deming DA, Dwyer A, Diehn M, Eng C, George TJ, Gollub MJ, Goodwin RA, Hamilton SR, Hechtman JF, Hochster H, Hong TS, Innocenti F, Iqbal A, Jacobs SA, Kennecke HF, Lee JJ, Lieu CH, Lenz HJ, Lindwasser OW, Montagut C, Odisio B, Ou FS, Porter L, Raghav K, Schrag D, Scott AJ, Shi Q, Strickler JH, Venook A, Yaeger R, Yothers G, You YN, Zell JA, Kopetz S.
      An increasing number of studies are describing potential uses of circulating tumour DNA (ctDNA) in the care of patients with colorectal cancer. Owing to this rapidly developing area of research, the Colon and Rectal-Anal Task Forces of the United States National Cancer Institute convened a panel of multidisciplinary experts to summarize current data on the utility of ctDNA in the management of colorectal cancer and to provide guidance in promoting the efficient development and integration of this technology into clinical care. The panel focused on four key areas in which ctDNA has the potential to change clinical practice, including the detection of minimal residual disease, the management of patients with rectal cancer, monitoring responses to therapy, and tracking clonal dynamics in response to targeted therapies and other systemic treatments. The panel also provides general guidelines with relevance for ctDNA-related research efforts, irrespective of indication.
  15. Proc Natl Acad Sci U S A. 2020 Jul 10. pii: 201920988. [Epub ahead of print]
    Lidsky PV, Andino R.
      Species-specific limits to lifespan (lifespan setpoint) determine the life expectancy of any given organism. Whether limiting lifespan provides an evolutionary benefit or is the result of an inevitable decline in fitness remains controversial. The identification of mutations extending lifespan suggests that aging is under genetic control, but the evolutionary driving forces limiting lifespan have not been defined. By examining the impact of lifespan on pathogen spread in a population, we propose that epidemics drive lifespan setpoints' evolution. Shorter lifespan limits infection spread and accelerates pathogen clearance when compared to populations with longer-lived individuals. Limiting longevity is particularly beneficial in the context of zoonotic transmissions, where pathogens must undergo adaptation to a new host. Strikingly, in populations exposed to pathogens, shorter-living variants outcompete individuals with longer lifespans. We submit that infection outbreaks can contribute to control the evolution of species' lifespan setpoints.
    Keywords:  aging; epidemics; evolution; lifespan
  16. Nat Rev Cancer. 2020 Jul 07.
    Weber J, Braun CJ, Saur D, Rad R.
      With the genetic portraits of all major human malignancies now available, we next face the challenge of characterizing the function of mutated genes, their downstream targets, interactions and molecular networks. Moreover, poorly understood at the functional level are also non-mutated but dysregulated genomes, epigenomes or transcriptomes. Breakthroughs in manipulative mouse genetics offer new opportunities to probe the interplay of molecules, cells and systemic signals underlying disease pathogenesis in higher organisms. Herein, we review functional screening strategies in mice using genetic perturbation and chemical mutagenesis. We outline the spectrum of genetic tools that exist, such as transposons, CRISPR and RNAi and describe discoveries emerging from their use. Genome-wide or targeted screens are being used to uncover genomic and regulatory landscapes in oncogenesis, metastasis or drug resistance. Versatile screening systems support experimentation in diverse genetic and spatio-temporal settings to integrate molecular, cellular or environmental context-dependencies. We also review the combination of in vivo screening and barcoding strategies to study genetic interactions and quantitative cancer dynamics during tumour evolution. These scalable functional genomics approaches are transforming our ability to interrogate complex biological systems.
  17. Curr Biol. 2020 Jul 06. pii: S0960-9822(20)30666-7. [Epub ahead of print]30(13): R770-R773
    Roberts MA, Segura-Roman A, Olzmann JA.
      Lipid droplets (LDs) are neutral lipid storage organelles assembled at the endoplasmic reticulum (ER). A new study reveals that the high membrane curvature of ER tubules catalyzes the nucleation of a neutral lipid lens, an early step in LD biogenesis.
  18. Nat Methods. 2020 Jul 06.
    Hu KH, Eichorst JP, McGinnis CS, Patterson DM, Chow ED, Kersten K, Jameson SC, Gartner ZJ, Rao AA, Krummel MF.
      Spatial transcriptomics seeks to integrate single cell transcriptomic data within the three-dimensional space of multicellular biology. Current methods to correlate a cell's position with its transcriptome in living tissues have various limitations. We developed an approach, called 'ZipSeq', that uses patterned illumination and photocaged oligonucleotides to serially print barcodes ('zipcodes') onto live cells in intact tissues, in real time and with an on-the-fly selection of patterns. Using ZipSeq, we mapped gene expression in three settings: in vitro wound healing, live lymph node sections and a live tumor microenvironment. In all cases, we discovered new gene expression patterns associated with histological structures. In the tumor microenvironment, this demonstrated a trajectory of myeloid and T cell differentiation from the periphery inward. A combinatorial variation of ZipSeq efficiently scales in the number of regions defined, providing a pathway for complete mapping of live tissues, subsequent to real-time imaging or perturbation.
  19. Cell Metab. 2020 Jul 02. pii: S1550-4131(20)30311-9. [Epub ahead of print]
    Kelly B, Pearce EL.
      Amino acids are fundamental building blocks supporting life. Their role in protein synthesis is well defined, but they contribute to a host of other intracellular metabolic pathways, including ATP generation, nucleotide synthesis, and redox balance, to support cellular and organismal function. Immune cells critically depend on such pathways to acquire energy and biomass and to reprogram their metabolism upon activation to support growth, proliferation, and effector functions. Amino acid metabolism plays a key role in this metabolic rewiring, and it supports various immune cell functions beyond increased protein synthesis. Here, we review the mechanisms by which amino acid metabolism promotes immune cell function, and how these processes could be targeted to improve immunity in pathological conditions.
    Keywords:  ▪▪▪
  20. Nature. 2020 Jul 08.
    Martínez-Reyes I, Cardona LR, Kong H, Vasan K, McElroy GS, Werner M, Kihshen H, Reczek CR, Weinberg SE, Gao P, Steinert EM, Piseaux R, Budinger GRS, Chandel NS.
      The mitochondrial electron transport chain (ETC) is necessary for tumour growth1-6 and its inhibition has demonstrated anti-tumour efficacy in combination with targeted therapies7-9. Furthermore, human brain and lung tumours display robust glucose oxidation by mitochondria10,11. However, it is unclear why a functional ETC is necessary for tumour growth in vivo. ETC function is coupled to the generation of ATP-that is, oxidative phosphorylation and the production of metabolites by the tricarboxylic acid (TCA) cycle. Mitochondrial complexes I and II donate electrons to ubiquinone, resulting in the generation of ubiquinol and the regeneration of the NAD+ and FAD cofactors, and complex III oxidizes ubiquinol back to ubiquinone, which also serves as an electron acceptor for dihydroorotate dehydrogenase (DHODH)-an enzyme necessary for de novo pyrimidine synthesis. Here we show impaired tumour growth in cancer cells that lack mitochondrial complex III. This phenotype was rescued by ectopic expression of Ciona intestinalis alternative oxidase (AOX)12, which also oxidizes ubiquinol to ubiquinone. Loss of mitochondrial complex I, II or DHODH diminished the tumour growth of AOX-expressing cancer cells deficient in mitochondrial complex III, which highlights the necessity of ubiquinone as an electron acceptor for tumour growth. Cancer cells that lack mitochondrial complex III but can regenerate NAD+ by expression of the NADH oxidase from Lactobacillus brevis (LbNOX)13 targeted to the mitochondria or cytosol were still unable to grow tumours. This suggests that regeneration of NAD+ is not sufficient to drive tumour growth in vivo. Collectively, our findings indicate that tumour growth requires the ETC to oxidize ubiquinol, which is essential to drive the oxidative TCA cycle and DHODH activity.
  21. EMBO J. 2020 Jul 09. e104730
    Dong R, Libby KA, Blaeschke F, Fuchs W, Marson A, Vale RD, Su X.
      The chimeric antigen receptor (CAR) directs T cells to target and kill specific cancer cells. Despite the success of CAR T therapy in clinics, the intracellular signaling pathways that lead to CAR T cell activation remain unclear. Using CD19 CAR as a model, we report that, similar to the endogenous T cell receptor (TCR), antigen engagement triggers the formation of CAR microclusters that transduce downstream signaling. However, CAR microclusters do not coalesce into a stable central supramolecular activation cluster (cSMAC). Moreover, LAT, an essential scaffold protein for TCR signaling, is not required for microcluster formation, immunological synapse formation, nor actin remodeling following CAR activation. However, CAR T cells still require LAT for an optimal production of the cytokine IL-2. Together, these data show that CAR T cells can bypass LAT for a subset of downstream signaling outputs, thus revealing a rewired signaling pathway as compared to native T cells.
    Keywords:   CAR ; LAT ; T cell signaling; actin; immunological synapse
  22. Dev Cell. 2020 Jun 30. pii: S1534-5807(20)30497-4. [Epub ahead of print]
    Li F, Simon MC.
      Solid tumors reside in harsh tumor microenvironments (TMEs) together with various stromal cell types. During tumor progression and metastasis, both tumor and stromal cells undergo rapid metabolic adaptations. Tumor cells metabolically coordinate or compete with their "neighbors" to maintain biosynthetic and bioenergetic demands while escaping immunosurveillance or therapeutic interventions. Here, we provide an update on metabolic communication between tumor cells and heterogeneous stromal components in primary and metastatic TMEs and discuss emerging strategies to target metabolic communications for improved cancer treatments.
    Keywords:  antitumor immunity; combination therapy; immunomodulation; metabolic communication; metabolic symbiosis; metabolism; metastasis; nutrient competition; signaling molecule; stromal cells; tumor microenvironment
  23. Cell. 2020 Jul 09. pii: S0092-8674(20)30756-X. [Epub ahead of print]182(1): 12-23
    Vijg J, Dong X.
      Age-related accumulation of postzygotic DNA mutations results in tissue genetic heterogeneity known as somatic mosaicism. Although implicated in aging as early as the 1950s, somatic mutations in normal tissue have been difficult to study because of their low allele fractions. With the recent emergence of cost-effective high-throughput sequencing down to the single-cell level, enormous progress has been made in our capability to quantitatively analyze somatic mutations in human tissue in relation to aging and disease. Here we first review how recent technological progress has opened up this field, providing the first broad sets of quantitative information on somatic mutations in vivo necessary to gain insight into their possible causal role in human aging and disease. We then propose three major mechanisms that can lead from accumulated de novo mutations across tissues to cell functional loss and human disease.
    Keywords:  age-related disease; aging; genome mosaicism; pathogenic effects; somatic DNA mutation; transcriptional noise
  24. Sci Adv. 2020 Jun;6(26): eaaz4764
    Sze CC, Ozark PA, Cao K, Ugarenko M, Das S, Wang L, Marshall SA, Rendleman EJ, Ryan CA, Zha D, Douillet D, Chen FX, Shilatifard A.
      Set1A and Set1B, two members of the COMPASS family of methyltransferases that methylate the histone H3 lysine 4 (H3K4) residue, have been accredited as primary depositors of global H3K4 trimethylation (H3K4me3) in mammalian cells. Our previous studies in mouse embryonic stem cells (ESCs) demonstrated that deleting the enzymatic SET domain of Set1A does not perturb bulk H3K4me3, indicating possible compensatory roles played by other COMPASS methyltransferases. Here, we generated a series of ESC lines harboring compounding mutations of COMPASS methyltransferases. We find that Set1B is functionally redundant to Set1A in implementing H3K4me3 at highly expressed genes, while Mll2 deposits H3K4me3 at less transcriptionally active promoters. While Set1A-B/COMPASS is responsible for broad H3K4me3 peaks, Mll2/COMPASS establishes H3K4me3 with narrow breadth. Additionally, Mll2 helps preserve global H3K4me3 levels and peak breadth in the absence of Set1A-B activity. Our results illustrate the biological flexibility of such enzymes in regulating transcription in a context-dependent manner to maintain stem cell identity.
  25. J Biol Chem. 2020 Jul 07. pii: jbc.REV119.007624. [Epub ahead of print]
    Wang YP, Li JT, Qu J, Yin M, Lei QY.
      Metabolites are not only substrates in metabolic reactions, but also signaling molecules controlling a wide range of cellular processes. Discovery of the oncometabolite 2-hydroxyglutarate (2HG) provides an important link between metabolic dysfunction and cancer, unveiling the signaling function of metabolites in regulating epigenetic and epitranscriptomic modifications, genome integrity, and signal transduction. It is now known that cancer cells sense and remodel their metabolic network to support biogenesis, caused by or resulting in the dysregulation of various metabolites. Cancer cells can sense alterations in metabolic intermediates to better coordinate multiple biological processes and enhance cell metabolism. Recent studies have demonstrated that metabolite signaling is involved in the regulation of malignant transformation, cell proliferation, epithelial-to-mesenchymal transition (EMT), differentiation blockade, and cancer stemness. Additionally, intercellular metabolite signaling modulates inflammatory response and immunosurveillance in the tumor microenvironment. Here, we review recent advances in cancer-associated metabolite signaling. An in-depth understanding of metabolite signaling will provide new opportunities for the development of therapeutic interventions that target cancer.
    Keywords:  Metabolite; Sensing; Signaling; cancer; metabolic disease; metabolic regulation; metabolomics; oncometabolite; signaling
  26. Elife. 2020 Jul 08. pii: e56862. [Epub ahead of print]9
    Kishikawa JI, Nakanishi A, Furuta A, Kato T, Namba K, Tamakoshi M, Mitsuoka K, Yokoyama K.
      V-ATPase is an energy converting enzyme, coupling ATP hydrolysis/synthesis in the hydrophilic V1 domain, with proton flow through the Vo membrane domain, via rotation of the central rotor complex relative to the surrounding stator apparatus. Upon dissociation from the V1 domain, the Vo domain of the eukaryotic V-ATPase can adopt a physiologically relevant auto-inhibited form in which proton conductance through the Vo domain is prevented, however the molecular mechanism of this inhibition is not fully understood. Using cryo-electron microscopy, we determined the structure of both the holo V/A-ATPase and isolated Vo at near-atomic resolution, respectively. These structures clarify how the isolated Vo domain adopts the auto-inhibited form and how the holo complex prevents formation of the inhibited Vo form.
    Keywords:  molecular biophysics; structural biology
  27. J Cell Sci. 2020 Jul 10. pii: jcs227900. [Epub ahead of print]133(13):
    Pakshir P, Noskovicova N, Lodyga M, Son DO, Schuster R, Goodwin A, Karvonen H, Hinz B.
      In 1971, Gabbiani and co-workers discovered and characterized the "modification of fibroblasts into cells which are capable of an active spasm" (contraction) in rat wound granulation tissue and, accordingly, named these cells 'myofibroblasts'. Now, myofibroblasts are not only recognized for their physiological role in tissue repair but also as cells that are key in promoting the development of fibrosis in all organs. In this Cell Science at a Glance and the accompanying poster, we provide an overview of the current understanding of central aspects of myofibroblast biology, such as their definition, activation from different precursors, the involved signaling pathways and most widely used models to study their function. Myofibroblasts will be placed into context with their extracellular matrix and with other cell types communicating in the fibrotic environment. Furthermore, the challenges and strategies to target myofibroblasts in anti-fibrotic therapies are summarized to emphasize their crucial role in disease progression.
    Keywords:  Fibrosis; Growth factor activation; Tissue repair; Wound healing
  28. Trends Cell Biol. 2020 Jul 02. pii: S0962-8924(20)30121-5. [Epub ahead of print]
    St Paul M, Ohashi PS.
      Effector CD8+ T cells are typically thought to be a homogenous group of cytotoxic cells that produce interferon-(IFN) γ. However, recent findings have challenged this notion because multiple subsets of CD8+ T cells have been described, each with distinct effector functions and cytotoxic potential. These subsets, referred to as the Tc subsets, have also been detected in tumor microenvironments (TMEs), where they potentially influence the antitumor response and patient outcomes. In this review, we highlight the prevalence and roles of Tc subsets in the TME. We also discuss their therapeutic applications in the context of adoptive immunotherapy to treat cancer.
    Keywords:  CD8(+) T cell; Tc subset; cytokines; immunotherapy
  29. Immunity. 2020 Jul 01. pii: S1074-7613(20)30271-5. [Epub ahead of print]
    Kumagai S, Togashi Y, Sakai C, Kawazoe A, Kawazu M, Ueno T, Sato E, Kuwata T, Kinoshita T, Yamamoto M, Nomura S, Tsukamoto T, Mano H, Shitara K, Nishikawa H.
      Only a small percentage of patients afflicted with gastric cancer (GC) respond to immune checkpoint blockade (ICB). To study the mechanisms underlying this resistance, we examined the immune landscape of GC. A subset of these tumors was characterized by high frequencies of regulatory T (Treg) cells and low numbers of effector T cells. Genomic analyses revealed that these tumors bore mutations in RHOA that are known to drive tumor progression. RHOA mutations in cancer cells activated the PI3K-AKT-mTOR signaling pathway, increasing production of free fatty acids that are more effectively consumed by Treg cells than effector T cells. RHOA mutant tumors were resistant to PD-1 blockade but responded to combination of PD-1 blockade with inhibitors of the PI3K pathway or therapies targeting Treg cells. We propose that the metabolic advantage conferred by RHOA mutations enables Treg cell accumulation within GC tumors, generating an immunosuppressive TME that underlies resistance to ICB.
    Keywords:  PI3K inhibitor; RHOA mutation; fatty acid metabolism; gastric cancer; non-inflamed tumor; regulatory T cell
  30. Mol Cell. 2020 Jul 03. pii: S1097-2765(20)30421-4. [Epub ahead of print]
    Wang C, Duan Y, Duan G, Wang Q, Zhang K, Deng X, Qian B, Gu J, Ma Z, Zhang S, Guo L, Liu C, Fang Y.
      Despite the prominent role of TDP-43 in neurodegeneration, its physiological and pathological functions are not fully understood. Here, we report an unexpected role of TDP-43 in the formation of dynamic, reversible, liquid droplet-like nuclear bodies (NBs) in response to stress. Formation of NBs alleviates TDP-43-mediated cytotoxicity in mammalian cells and fly neurons. Super-resolution microscopy reveals distinct functions of the two RRMs in TDP-43 NB formation. TDP-43 NBs are partially colocalized with nuclear paraspeckles, whose scaffolding lncRNA NEAT1 is dramatically upregulated in stressed neurons. Moreover, increase of NEAT1 promotes TDP-43 liquid-liquid phase separation (LLPS) in vitro. Finally, we discover that the ALS-associated mutation D169G impairs the NEAT1-mediated TDP-43 LLPS and NB assembly, causing excessive cytoplasmic translocation of TDP-43 to form stress granules, which become phosphorylated TDP-43 cytoplasmic foci upon prolonged stress. Together, our findings suggest a stress-mitigating role and mechanism of TDP-43 NBs, whose dysfunction may be involved in ALS pathogenesis.
    Keywords:  ALS; TDP-43; lncRNA NEAT1; nuclear body; paraspeckle; phase separation; stress granules
  31. Cell Metab. 2020 Jun 30. pii: S1550-4131(20)30309-0. [Epub ahead of print]
    Jensen-Cody SO, Flippo KH, Claflin KE, Yavuz Y, Sapouckey SA, Walters GC, Usachev YM, Atasoy D, Gillum MP, Potthoff MJ.
      Fibroblast growth factor 21 (FGF21) is an endocrine hormone produced by the liver that regulates nutrient and metabolic homeostasis. FGF21 production is increased in response to macronutrient imbalance and signals to the brain to suppress sugar intake and sweet-taste preference. However, the central targets mediating these effects have been unclear. Here, we identify FGF21 target cells in the hypothalamus and reveal that FGF21 signaling to glutamatergic neurons is both necessary and sufficient to mediate FGF21-induced sugar suppression and sweet-taste preference. Moreover, we show that FGF21 acts directly in the ventromedial hypothalamus (VMH) to specifically regulate sucrose intake, but not non-nutritive sweet-taste preference, body weight, or energy expenditure. Finally, our data demonstrate that FGF21 affects neuronal activity by increasing activation and excitability of neurons in the VMH. Thus, FGF21 signaling to glutamatergic neurons in the VMH is an important component of the neurocircuitry that functions to regulate sucrose intake.
    Keywords:  FGF21; PVN; VMH; brain; glucose sensing; hepatokine; hypothalamus; liver; macronutrient; sugar intake; sweet-taste preference
  32. EMBO Rep. 2020 Jul 06. e201949801
    Han S, Jeong YY, Sheshadri P, Su X, Cai Q.
      Synaptic mitochondria are particularly vulnerable to physiological insults, and defects in synaptic mitochondria are linked to early pathophysiology of Alzheimer's disease (AD). Mitophagy, a cargo-specific autophagy for elimination of dysfunctional mitochondria, constitutes a key quality control mechanism. However, how mitophagy ensures synaptic mitochondrial integrity remains largely unknown. Here, we reveal Rheb and Snapin as key players regulating mitochondrial homeostasis at synapses. Rheb initiates mitophagy to target damaged mitochondria for autophagy, whereas dynein-Snapin-mediated retrograde transport promotes clearance of mitophagosomes from synaptic terminals. We demonstrate that synaptic accumulation of mitophagosomes is a feature in AD-related mutant hAPP mouse brains, which is attributed to increased mitophagy initiation coupled with impaired removal of mitophagosomes from AD synapses due to defective retrograde transport. Furthermore, while deficiency in dynein-Snapin-mediated retrograde transport recapitulates synaptic mitophagy stress and induces synaptic degeneration, elevated Snapin expression attenuates mitochondrial defects and ameliorates synapse loss in AD mouse brains. Taken together, our study provides new insights into mitophagy regulation of synaptic mitochondrial integrity, establishing a foundation for mitigating AD-associated mitochondria deficits and synaptic damage through mitophagy enhancement.
    Keywords:  Alzheimer's; mitophagosome; retrograde transport; synaptic mitochondrial deficits; synaptic mitophagy
  33. Elife. 2020 Jul 10. pii: e57544. [Epub ahead of print]9
    Kitajima N, Takikawa K, Sekiya H, Satoh K, Asanuma D, Sakamoto H, Takahashi S, Hanaoka K, Urano Y, Namiki S, Iino M, Hirose K.
      Adenosine 5' triphosphate (ATP) is a ubiquitous extracellular signaling messenger. Here, we describe a method for in-vivo imaging of extracellular ATP with high spatiotemporal resolution. We prepared a comprehensive set of cysteine-substitution mutants of ATP-binding protein, Bacillus FoF1-ATP synthase e subunit, labeled with small-molecule fluorophores at the introduced cysteine residue. Screening revealed that the Cy3-labeled glutamine-105 mutant (Q105C-Cy3; designated ATPOS) shows a large fluorescence change in the presence of ATP, with submicromolar affinity, pH-independence, and high selectivity for ATP over ATP metabolites and other nucleotides. To enable in-vivo validation, we introduced BoNT/C-Hc for binding to neuronal plasma membrane and Alexa Fluor 488 for ratiometric measurement. The resulting ATPOS complex binds to neurons in cerebral cortex of living mice, and clearly visualized a concentrically propagating wave of extracellular ATP release in response to electrical stimulation. ATPOS should be useful to probe the extracellular ATP dynamics of diverse biological processes in vivo.
    Keywords:  biochemistry; chemical biology; mouse; neuroscience
  34. Mol Cell. 2020 Jul 01. pii: S1097-2765(20)30429-9. [Epub ahead of print]
    Huang J, Zhou Q, Gao M, Nowsheen S, Zhao F, Kim W, Zhu Q, Kojima Y, Yin P, Zhang Y, Guo G, Tu X, Deng M, Luo K, Qin B, Machida Y, Lou Z.
      DNA-protein crosslinks (DPCs) are highly toxic DNA lesions that threaten genomic integrity. Recent findings highlight that SPRTN, a specialized DNA-dependent metalloprotease, is a central player in proteolytic cleavage of DPCs. Previous studies suggest that SPRTN deubiquitination is important for its chromatin association and activation. However, the regulation and consequences of SPRTN deubiquitination remain unclear. Here we report that, in response to DPC induction, the deubiquitinase VCPIP1/VCIP135 is phosphorylated and activated by ATM/ATR. VCPIP1, in turn, deubiquitinates SPRTN and promotes its chromatin relocalization. Deubiquitination of SPRTN is required for its subsequent acetylation, which promotes SPRTN relocation to the site of chromatin damage. Furthermore, Vcpip1 knockout mice are prone to genomic instability and premature aging. We propose a model where two sequential post-translational modifications (PTMs) regulate SPRTN chromatin accessibility to repair DPCs and maintain genomic stability and a healthy lifespan.
    Keywords:  DNA repair; DNA-protein crosslink; SPRTN; Top1cc; VCPIP1/VCIP135; acetylation; aging; genomic instability; metalloprotease; ubiquitination
  35. Cancer Res. 2020 Jul 07. pii: canres.2888.2019. [Epub ahead of print]
    Hayashi M, Kuga A, Suzuki M, Panda H, Kitamura H, Motohashi H, Yamamoto M.
      The transcription factor Nrf2 activates transcription of cytoprotective genes during oxidative and electrophilic insults. Nrf2 activity is regulated by Keap1 in a stress-dependent manner in normal cells, and somatic loss-of-function mutations of Keap1 are known to induce constitutive Nrf2 activation, especially in lung adenocarcinomas, conferring survival and proliferative benefits to tumors. Therefore, several therapeutic strategies that aim to inhibit Nrf2 in tumors have been developed for the treatment of Nrf2-activated cancers. Here we addressed whether targeting Nrf2 activation in the microenvironment can suppress the progression of Nrf2-activated tumors. We combined two types of Keap1-flox mice expressing variable levels of Keap1 with a Kras-driven adenocarcinoma model to generate Keap1-deficient lung tumors surrounded by normal or Keap1-knockdown host cells. In this model system, activation of Nrf2 in the microenvironment prolonged the survival of Nrf2-activated tumor-bearing mice. The Nrf2-activated microenvironment suppressed tumor burden; in particular, preinvasive lesion formation was significantly suppressed. Notably, loss of Nrf2 in bone marrow-derived cells in Nrf2-activated host cells appeared to counteract the suppression of Nrf2-activated cancer progression. Thus, these results demonstrate that microenvironmental Nrf2 activation suppresses the progression of malignant Nrf2-activated tumors and that Nrf2 activation in immune cells at least partially contributes to these suppressive effects.
  36. Cancer Discov. 2020 Jul 10. pii: CD-20-0036. [Epub ahead of print]
    Marigo I, Trovato R, Hofer F, Ingangi V, DE Sanctis F, Ugel S, Cane S, Simonelli A, Lamolinara A, Iezzi M, Fassan M, Rugge M, Boschi F, Borile G, Eisenhaure T, Sarkizova S, Lieb D, Hacohen N, Azzolin L, Piccolo S, Lawlor R, Scarpa A, Carbognin L, Bria E, Bicciato S, Murray PJ, Bronte V.
      Tumor-associated macrophages (TAMs) are regulators of extracellular matrix (ECM) remodeling and metastatic progression, the main cause of cancer-associated death. We found that disabled 2 mitogen-responsive phosphoprotein (DAB2) is highly expressed in tumor-infiltrating TAMs and its genetic ablation significantly impairs lung metastasis formation. DAB2-expressing TAMs, mainly localized along the tumor invasive front, participate in integrin recycling, ECM remodeling and directional migration in a tridimensional matrix. DAB2+ macrophages escort the invasive dissemination of cancer cells by a mechanosensing pathway requiring the transcription factor Yes-Associated Protein. In human lobular breast and gastric carcinomas, DAB2+ TAMs correlated with a poor clinical outcome, identifying DAB2 as potential prognostic biomarker for cancer patient stratification. DAB2 is therefore central for the pro-metastatic activity of TAMs.
  37. Nature. 2020 Jul 08.
    Jimenez-Blasco D, Busquets-Garcia A, Hebert-Chatelain E, Serrat R, Vicente-Gutierrez C, Ioannidou C, Gómez-Sotres P, Lopez-Fabuel I, Resch-Beusher M, Resel E, Arnouil D, Saraswat D, Varilh M, Cannich A, Julio-Kalajzic F, Bonilla-Del Río I, Almeida A, Puente N, Achicallende S, Lopez-Rodriguez ML, Jollé C, Déglon N, Pellerin L, Josephine C, Bonvento G, Panatier A, Lutz B, Piazza PV, Guzmán M, Bellocchio L, Bouzier-Sore AK, Grandes P, Bolaños JP, Marsicano G.
      Astrocytes take up glucose from the bloodstream to provide energy to the brain, thereby allowing neuronal activity and behavioural responses1-5. By contrast, astrocytes are under neuronal control through specific neurotransmitter receptors5-7. However, whether the activation of astroglial receptors can directly regulate cellular glucose metabolism to eventually modulate behavioural responses is unclear. Here we show that activation of mouse astroglial type-1 cannabinoid receptors associated with mitochondrial membranes (mtCB1) hampers the metabolism of glucose and the production of lactate in the brain, resulting in altered neuronal functions and, in turn, impaired behavioural responses in social interaction assays. Specifically, activation of astroglial mtCB1 receptors reduces the phosphorylation of the mitochondrial complex I subunit NDUFS4, which decreases the stability and activity of complex I. This leads to a reduction in the generation of reactive oxygen species by astrocytes and affects the glycolytic production of lactate through the hypoxia-inducible factor 1 pathway, eventually resulting in neuronal redox stress and impairment of behavioural responses in social interaction assays. Genetic and pharmacological correction of each of these effects abolishes the effect of cannabinoid treatment on the observed behaviour. These findings suggest that mtCB1 receptor signalling can directly regulate astroglial glucose metabolism to fine-tune neuronal activity and behaviour in mice.
  38. Autophagy. 2020 Jul 05. 1-21
    Sanchez-Garrido J, Shenoy AR.
      Nutrients not only act as building blocks but also as signaling molecules. Nutrient-availability promotes cell growth and proliferation and suppresses catabolic processes, such as macroautophagy/autophagy. These effects are mediated by checkpoint kinases such as MTOR (mechanistic target of rapamycin kinase), which is activated by amino acids and growth factors, and AMP-activated protein kinase (AMPK), which is activated by low levels of glucose or ATP. These kinases have wide-ranging activities that can be co-opted by immune cells upon exposure to danger signals, cytokines or pathogens. Here, we discuss recent insight into the regulation and repurposing of nutrient-sensing responses by the innate immune system during infection. Moreover, we examine how natural mutations and pathogen-mediated interventions can alter the balance between anabolic and autophagic pathways leading to a breakdown in tissue homeostasis and/or host defense.ABBREVIATIONS: AKT1/PKB: AKT serine/threonine kinase 1; ATG: autophagy related; BECN1: beclin 1; CGAS: cyclic GMP-AMP synthase; EIF2AK4/GCN2: eukaryotic translation initiation factor 2 alpha kinase 4; ER: endoplasmic reticulum; FFAR: free fatty acid receptor; GABARAP: GABA type A receptor-associated protein; IFN: interferon; IL: interleukin; LAP: LC3-associated phagocytosis; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MAP3K7/TAK1: mitogen-activated protein kinase kinase kinase 7; MAPK: mitogen-activated protein kinase; MTOR: mechanistic target of rapamycin kinase; NLR: NOD (nucleotide-binding oligomerization domain) and leucine-rich repeat containing proteins; PI3K, phosphoinositide 3-kinase; PRR: pattern-recognition receptor; PtdIns3K: phosphatidylinositol 3-kinase; RALB: RAS like proto-oncogene B; RHEB: Ras homolog, MTORC1 binding; RIPK1: receptor interacting serine/threonine kinase 1; RRAG: Ras related GTP binding; SQSTM1/p62: sequestosome 1; STING1/TMEM173: stimulator of interferon response cGAMP interactor 1; STK11/LKB1: serine/threonine kinase 11; TBK1: TANK binding kinase 1; TLR: toll like receptor; TNF: tumor necrosis factor; TRAF6: TNF receptor associated factor 6; TRIM: tripartite motif protein; ULK1: unc-51 like autophagy activating kinase 1; V-ATPase: vacuolar-type H+-proton-translocating ATPase.
    Keywords:  AMPK; LC3-associated phagocytosis; MTOR; immunity; microbial pathogenesis; unconventional secretion
  39. Cell Death Dis. 2020 Jul 08. 11(7): 517
    Noguchi M, Hirata N, Tanaka T, Suizu F, Nakajima H, Chiorini JA.
      The balance between cell death and survival is a critical parameter in the regulation of cells and the maintenance of homeostasis in vivo. Three major mechanisms for cell death have been identified in mammalian cells: apoptosis (type I), autophagic cell death (type II), and necrosis (type III). These three mechanisms have been suggested to engage in cross talk with each other. Among them, autophagy was originally characterized as a cell survival mechanism for amino acid recycling during starvation. Whether autophagy functions primarily in cell survival or cell death is a critical question yet to be answered. Here, we present a comprehensive review of the cell death-related events that take place during autophagy and their underlying mechanisms in cancer and autoimmune disease development.
  40. Sci Signal. 2020 Jul 07. pii: eaax6313. [Epub ahead of print]13(639):
    Shrestha S, Katiyar S, Sanz-Rodriguez CE, Kemppinen NR, Kim HW, Kadirvelraj R, Panagos C, Keyhaninejad N, Colonna M, Chopra P, Byrne DP, Boons GJ, van der Knaap E, Eyers PA, Edison AS, Wood ZA, Kannan N.
      Aberrant regulation of metabolic kinases by altered redox homeostasis substantially contributes to aging and various diseases, such as diabetes. We found that the catalytic activity of a conserved family of fructosamine-3-kinases (FN3Ks), which are evolutionarily related to eukaryotic protein kinases, is regulated by redox-sensitive cysteine residues in the kinase domain. The crystal structure of the FN3K homolog from Arabidopsis thaliana revealed that it forms an unexpected strand-exchange dimer in which the ATP-binding P-loop and adjoining β strands are swapped between two chains in the dimer. This dimeric configuration is characterized by strained interchain disulfide bonds that stabilize the P-loop in an extended conformation. Mutational analysis and solution studies confirmed that the strained disulfides function as redox "switches" to reversibly regulate the activity and dimerization of FN3K. Human FN3K, which contains an equivalent P-loop Cys, was also redox sensitive, whereas ancestral bacterial FN3K homologs, which lack a P-loop Cys, were not. Furthermore, CRISPR-mediated knockout of FN3K in human liver cancer cells altered the abundance of redox metabolites, including an increase in glutathione. We propose that redox regulation evolved in FN3K homologs in response to changing cellular redox conditions. Our findings provide insights into the origin and evolution of redox regulation in the protein kinase superfamily and may open new avenues for targeting human FN3K in diabetic complications.
  41. Nat Commun. 2020 Jul 08. 11(1): 3409
    Beel S, Kolloch L, Apken LH, Jürgens L, Bolle A, Sudhof N, Ghosh S, Wardelmann E, Meisterernst M, Steinestel K, Oeckinghaus A.
      Pancreatic ductal adenocarcinoma (PDAC) is associated with high mortality and therapy resistance. Here, we show that low expression of κB-Ras GTPases is frequently detected in PDAC and correlates with higher histologic grade. In a model of KRasG12D-driven PDAC, loss of κB-Ras accelerates tumour development and shortens median survival. κB-Ras deficiency promotes acinar-to-ductal metaplasia (ADM) during tumour initiation as well as tumour progression through intrinsic effects on proliferation and invasion. κB-Ras proteins are also required for acinar regeneration after pancreatitis, demonstrating a general role in control of plasticity. Molecularly, upregulation of Ral GTPase activity and Sox9 expression underlies the observed phenotypes, identifying a previously unrecognized function of Ral signalling in ADM. Our results provide evidence for a tumour suppressive role of κB-Ras proteins and highlight low κB-Ras levels and consequent loss of Ral control as risk factors, thus emphasizing the necessity for therapeutic options that allow interference with Ral-driven signalling.
  42. EMBO Rep. 2020 Jul 09. e49583
    Du G, Qiao Y, Zhuo Z, Zhou J, Li X, Liu Z, Li Y, Chen H.
      The age-associated decline of adult stem cell function is closely related to the decline in tissue function and age-related diseases. However, the underlying mechanisms that ultimately lead to the observed functional decline of stem cells still remain largely unexplored. This study investigated Drosophila midguts and found a continuous downregulation of lipoic acid synthase, which encodes the key enzyme for the endogenous synthesis of alpha-lipoic acid (ALA), upon aging. Importantly, orally administration of ALA significantly reversed the age-associated hyperproliferation of intestinal stem cells (ISCs) and the observed decline of intestinal function, thus extending the lifespan of Drosophila. This study reports that ALA reverses age-associated ISC dysfunction by promoting the activation of the endocytosis-autophagy network, which decreases in aged ISCs. Moreover, this study suggests that ALA may be used as a safe and effective anti-aging compound for the treatment of ISC-dysfunction-related diseases and for the promotion of healthy aging in humans.
    Keywords:  aging; alpha-lipoic acid; endocytosis; intestinal stem cell; longevity
  43. Proc Natl Acad Sci U S A. 2020 Jul 10. pii: 202006965. [Epub ahead of print]
    Hiratsuka T, Bordeu I, Pruessner G, Watt FM.
      Fluctuation in signal transduction pathways is frequently observed during mammalian development. However, its role in regulating stem cells has not been explored. Here we tracked spatiotemporal ERK MAPK dynamics in human epidermal stem cells. While stem cells and differentiated cells were distinguished by high and low stable basal ERK activity, respectively, we also found cells with pulsatile ERK activity. Transitions from Basalhi-Pulselo (stem) to Basalhi-Pulsehi, Basalmid-Pulsehi, and Basallo-Pulselo (differentiated) cells occurred in expanding keratinocyte colonies and in response to differentiation stimuli. Pharmacological inhibition of ERK induced differentiation only when cells were in the Basalmid-Pulsehi state. Basal ERK activity and pulses were differentially regulated by DUSP10 and DUSP6, leading us to speculate that DUSP6-mediated ERK pulse down-regulation promotes initiation of differentiation, whereas DUSP10-mediated down-regulation of mean ERK activity promotes and stabilizes postcommitment differentiation. Levels of MAPK1/MAPK3 transcripts correlated with DUSP6 and DUSP10 transcripts in individual cells, suggesting that ERK activity is negatively regulated by transcriptional and posttranslational mechanisms. When cells were cultured on a topography that mimics the epidermal-dermal interface, spatial segregation of mean ERK activity and pulses was observed. In vivo imaging of mouse epidermis revealed a patterned distribution of basal cells with pulsatile ERK activity, and down-regulation was linked to the onset of differentiation. Our findings demonstrate that ERK MAPK signal fluctuations link kinase activity to stem cell dynamics.
    Keywords:  ERK; cell signaling; keratinocytes; live imaging; stem cells
  44. Elife. 2020 Jul 09. pii: e55388. [Epub ahead of print]9
    Tsang AH, Koch CE, Kiehn JT, Schmidt CX, Oster H.
      Endogenous circadian clocks have evolved to anticipate 24-hour rhythms in environmental demands. Recent studies suggest that circadian rhythm disruption is a major risk factor for the development of metabolic disorders in humans. Conversely, alterations in energy state can disrupt circadian rhythms of behavior and physiology, creating a vicious circle of metabolic dysfunction. How peripheral energy state affects diurnal food intake, however, is still poorly understood. We here show that the adipokine adiponectin (ADIPOQ) regulates diurnal feeding rhythms through clocks in energy regulatory centers of the mediobasal hypothalamus (MBH). Adipoq-deficient mice show increased rest phase food intake associated with disrupted transcript rhythms of clock and appetite-regulating genes in the MBH. ADIPOQ regulates MBH clocks via AdipoR1-mediated upregulation of the core clock gene Bmal1. BMAL1, in turn, controls expression of orexigenic neuropeptide expression in the MBH. Together, these data reveal a systemic metabolic circuit to regulate central circadian clocks and energy intake.
    Keywords:  genetics; genomics; mouse; neuroscience
  45. Nature. 2020 Jul 08.
    Muller PA, Schneeberger M, Matheis F, Wang P, Kerner Z, Ilanges A, Pellegrino K, Del Mármol J, Castro TBR, Furuichi M, Perkins M, Han W, Rao A, Picard AJ, Cross JR, Honda K, de Araujo I, Mucida D.
      Connections between the gut and brain monitor the intestinal tissue and its microbial and dietary content1, regulating both physiological intestinal functions such as nutrient absorption and motility2,3, and brain-wired feeding behaviour2. It is therefore plausible that circuits exist to detect gut microorganisms and relay this information to areas of the central nervous system that, in turn, regulate gut physiology4. Here we characterize the influence of the microbiota on enteric-associated neurons by combining gnotobiotic mouse models with transcriptomics, circuit-tracing methods and functional manipulations. We find that the gut microbiome modulates gut-extrinsic sympathetic neurons: microbiota depletion leads to increased expression of the neuronal transcription factor cFos, and colonization of germ-free mice with bacteria that produce short-chain fatty acids suppresses cFos expression in the gut sympathetic ganglia. Chemogenetic manipulations, translational profiling and anterograde tracing identify a subset of distal intestine-projecting vagal neurons that are positioned to have an afferent role in microbiota-mediated modulation of gut sympathetic neurons. Retrograde polysynaptic neuronal tracing from the intestinal wall identifies brainstem sensory nuclei that are activated during microbial depletion, as well as efferent sympathetic premotor glutamatergic neurons that regulate gastrointestinal transit. These results reveal microbiota-dependent control of gut-extrinsic sympathetic activation through a gut-brain circuit.
  46. Elife. 2020 Jul 10. pii: e55513. [Epub ahead of print]9
    Chiao YA, Zhang H, Sweetwyne M, Whitson J, Ting YS, Basisty N, Pino LK, Quarles E, Nguyen NH, Campbell MD, Zhang T, Gaffrey MJ, Merrihew G, Wang L, Yue Y, Duan D, Granzier HL, Szeto HH, Qian WJ, Marcinek D, MacCoss MJ, Rabinovitch P.
      Diastolic dysfunction is a prominent feature of cardiac aging in both mice and humans. We show here that 8-week treatment of old mice with the mitochondrial targeted peptide SS-31 (elamipretide) can substantially reverse this deficit. SS-31 normalized the increase in proton leak and reduced mitochondrial ROS in cardiomyocytes from old mice, accompanied by reduced protein oxidation and a shift towards a more reduced protein thiol redox state in old hearts. Improved diastolic function was concordant with increased phosphorylation of cMyBP-C Ser282 but was independent of titin isoform shift. Late-life viral expression of mitochondrial-targeted catalase (mCAT) produced similar functional benefits in old mice and SS-31 did not improve cardiac function of old mCAT mice, implicating normalizing mitochondrial oxidative stress as an overlapping mechanism. These results demonstrate that pre-existing cardiac aging phenotypes can be reversed by targeting mitochondrial dysfunction and implicate mitochondrial energetics and redox signaling as therapeutic targets for cardiac aging.
    Keywords:  human biology; medicine; mouse
  47. Gut. 2020 Jul 08. pii: gutjnl-2020-320946. [Epub ahead of print]
    Reyes-Uribe L, Wu W, Gelincik O, Bommi PV, Francisco-Cruz A, Solis LM, Lynch PM, Lim R, Stoffel EM, Kanth P, Samadder NJ, Mork ME, Taggart MW, Milne GL, Marnett LJ, Vornik L, Liu DD, Revuelta M, Chang K, You YN, Kopelovich L, Wistuba II, Lee JJ, Sei S, Shoemaker RH, Szabo E, Richmond E, Umar A, Perloff M, Brown PH, Lipkin SM, Vilar E.
      OBJECTIVE: Patients with Lynch syndrome (LS) are at markedly increased risk for colorectal cancer. It is being increasingly recognised that the immune system plays an essential role in LS tumour development, thus making an ideal target for cancer prevention. Our objective was to evaluate the safety, assess the activity and discover novel molecular pathways involved in the activity of naproxen as primary and secondary chemoprevention in patients with LS.DESIGN: We conducted a Phase Ib, placebo-controlled, randomised clinical trial of two dose levels of naproxen sodium (440 and 220 mg) administered daily for 6 months to 80 participants with LS, and a co-clinical trial using a genetically engineered mouse model of LS and patient-derived organoids (PDOs).
    RESULTS: Overall, the total number of adverse events was not different across treatment arms with excellent tolerance of the intervention. The level of prostaglandin E2 in the colorectal mucosa was significantly decreased after treatment with naproxen when compared with placebo. Naproxen activated different resident immune cell types without any increase in lymphoid cellularity, and changed the expression patterns of the intestinal crypt towards epithelial differentiation and stem cell regulation. Naproxen demonstrated robust chemopreventive activity in a mouse co-clinical trial and gene expression profiles induced by naproxen in humans showed perfect discrimination of mice specimens with LS and PDOs treated with naproxen and control.
    CONCLUSIONS: Naproxen is a promising strategy for immune interception in LS. We have discovered naproxen-induced gene expression profiles for their potential use as predictive biomarkers of drug activity.
    TRIAL REGISTRATION NUMBER: gov Identifier: NCT02052908.
    Keywords:  HNPCC syndrome; cancer syndromes; chemoprevention; gene expression; non-steroidal anti-inflammatory drugs
  48. J Cachexia Sarcopenia Muscle. 2020 Jul 08.
    Cho DS, Schmitt RE, Dasgupta A, Ducharme AM, Doles JD.
      BACKGROUND: Persistent loss of skeletal muscle mass and function as well as altered fat metabolism are frequently observed in severe sepsis survivors. Studies examining sepsis-associated tissue dysfunction from the perspective of the tissue microenvironment are scarce. In this study, we comprehensively assessed transcriptional changes in muscle and fat at single-cell resolution following experimental sepsis induction.METHODS: Skeletal muscle and visceral white adipose tissue from control mice or mice 1 day or 1 month following faecal slurry-induced sepsis were used. Single cells were mechanically and enzymatically prepared from whole tissue, and viable cells were further isolated by fluorescence activated cell sorting. Droplet-based single-cell RNA-sequencing (scRNA-seq; 10× Genomics) was used to generate single-cell gene expression profiles of thousands of muscle and fat-resident cells. Bioinformatics analyses were performed to identify and compare individual cell populations in both tissues.
    RESULTS: In skeletal muscle, scRNA-seq analysis classified 1438 single cells into myocytes, endothelial cells, fibroblasts, mesenchymal stem cells, macrophages, neutrophils, T-cells, B-cells, and dendritic cells. In adipose tissue, scRNA-seq analysis classified 2281 single cells into adipose stem cells, preadipocytes, endothelial cells, fibroblasts, macrophages, dendritic cells, B-cells, T-cells, NK cells, and gamma delta T-cells. One day post-sepsis, the proportion of most non-immune cell populations was decreased, while immune cell populations, particularly neutrophils and macrophages, were highly enriched. Proportional changes of endothelial cells, neutrophils, and macrophages were validated using faecal slurry and cecal ligation and puncture models. At 1 month post-sepsis, we observed persistent enrichment/depletion of cell populations and further uncovered a cell-type and tissue-specific ability to return to a baseline transcriptomic state. Differential gene expression analyses revealed key genes and pathways altered in post-sepsis muscle and fat and highlighted the engagement of infection/inflammation and tissue damage signalling. Finally, regulator analysis identified gonadotropin-releasing hormone and Bay 11-7082 as targets/compounds that we show can reduce sepsis-associated loss of lean or fat mass.
    CONCLUSIONS: These data demonstrate persistent post-sepsis muscle and adipose tissue disruption at the single-cell level and highlight opportunities to combat long-term post-sepsis tissue wasting using bioinformatics-guided therapeutic interventions.
    Keywords:  Fat; Sepsis; Single-cell RNA-sequencing; Skeletal muscle
  49. Cancers (Basel). 2020 Jul 04. pii: E1787. [Epub ahead of print]12(7):
    Sahinbegovic H, Jelinek T, Hrdinka M, Bago JR, Turi M, Sevcikova T, Kurtovic-Kozaric A, Hajek R, Simicek M.
      Cell-to-cell communication is a fundamental process in every multicellular organism. In addition to membrane-bound and released factors, the sharing of cytosolic components represents a new, poorly explored signaling route. An extraordinary example of this communication channel is the direct transport of mitochondria between cells. In this review, we discuss how intercellular mitochondrial transfer can be used by cancer cells to sustain their high metabolic requirements and promote drug resistance and describe relevant molecular players in the context of current and future cancer therapy.
    Keywords:  cancer; mitochondria; mitochondrial transfer; tumor microenvironment; tunneling nanotubes
  50. Nature. 2020 Jul 08.
    Gallotta I, Sandhu A, Peters M, Haslbeck M, Jung R, Agilkaya S, Blersch JL, Rödelsperger C, Röseler W, Huang C, Sommer RJ, David DC.
      In metazoans, the secreted proteome participates in intercellular signalling and innate immunity, and builds the extracellular matrix scaffold around cells. Compared with the relatively constant intracellular environment, conditions for proteins in the extracellular space are harsher, and low concentrations of ATP prevent the activity of intracellular components of the protein quality-control machinery. Until now, only a few bona fide extracellular chaperones and proteases have been shown to limit the aggregation of extracellular proteins1-5. Here we performed a systematic analysis of the extracellular proteostasis network in Caenorhabditis elegans with an RNA interference screen that targets genes that encode the secreted proteome. We discovered 57 regulators of extracellular protein aggregation, including several proteins related to innate immunity. Because intracellular proteostasis is upregulated in response to pathogens6-9, we investigated whether pathogens also stimulate extracellular proteostasis. Using a pore-forming toxin to mimic a pathogenic attack, we found that C. elegans responded by increasing the expression of components of extracellular proteostasis and by limiting aggregation of extracellular proteins. The activation of extracellular proteostasis was dependent on stress-activated MAP kinase signalling. Notably, the overexpression of components of extracellular proteostasis delayed ageing and rendered worms resistant to intoxication. We propose that enhanced extracellular proteostasis contributes to systemic host defence by maintaining a functional secreted proteome and avoiding proteotoxicity.
  51. Nat Immunol. 2020 Jul 06.
    Wolf T, Jin W, Zoppi G, Vogel IA, Akhmedov M, Bleck CKE, Beltraminelli T, Rieckmann JC, Ramirez NJ, Benevento M, Notarbartolo S, Bumann D, Meissner F, Grimbacher B, Mann M, Lanzavecchia A, Sallusto F, Kwee I, Geiger R.
      In response to pathogenic threats, naive T cells rapidly transition from a quiescent to an activated state, yet the underlying mechanisms are incompletely understood. Using a pulsed SILAC approach, we investigated the dynamics of mRNA translation kinetics and protein turnover in human naive and activated T cells. Our datasets uncovered that transcription factors maintaining T cell quiescence had constitutively high turnover, which facilitated their depletion following activation. Furthermore, naive T cells maintained a surprisingly large number of idling ribosomes as well as 242 repressed mRNA species and a reservoir of glycolytic enzymes. These components were rapidly engaged following stimulation, promoting an immediate translational and glycolytic switch to ramp up the T cell activation program. Our data elucidate new insights into how T cells maintain a prepared state to mount a rapid immune response, and provide a resource of protein turnover, absolute translation kinetics and protein synthesis rates in T cells (
  52. Cancers (Basel). 2020 Jun 30. pii: E1745. [Epub ahead of print]12(7):
    Perrigue PM, Rakoczy M, Pawlicka KP, Belter A, Giel-Pietraszuk M, Naskręt-Barciszewska M, Barciszewski J, Figlerowicz M.
      Cellular senescence is a tumor-suppressive mechanism blocking cell proliferation in response to stress. However, recent evidence suggests that senescent tumor cells can re-enter the cell cycle to become cancer stem cells, leading to relapse after cancer chemotherapy treatment. Understanding how the senescence reprogramming process is a precursor to cancer stem cell formation is of great medical importance. To study the interplay between senescence, stemness, and cancer, we applied a stem cell medium (SCM) to human embryonic fibroblasts (MRC5 and WI-38) and cancer cell lines (A549 and 293T). MRC5 and WI-38 cells treated with SCM showed symptoms of oxidative stress and became senescent. Transcriptome analysis over a time course of SCM-induced senescence, revealed a developmental process overlapping with the upregulation of genes for growth arrest and the senescence-associated secretory phenotype (SASP). We demonstrate that histone demethylases jumonji domain-containing protein D3 (Jmjd3) and ubiquitously transcribed tetratricopeptide repeat, X chromosome (Utx), which operate by remodeling chromatin structure, are implicated in the senescence reprogramming process to block stem cell formation in fibroblasts. In contrast, A549 and 293T cells cultured in SCM were converted to cancer stem cells that displayed the phenotype of senescence uncoupled from growth arrest. The direct overexpression of DNA methyltransferases (Dnmt1 and Dnmt3A), ten-eleven translocation methylcytosine dioxygenases (Tet1 and Tet3), Jmjd3, and Utx proteins could activate senescence-associated beta-galactosidase (SA-β-gal) activity in 293T cells, suggesting that epigenetic alteration and chromatin remodeling factors trigger the senescence response. Overall, our study suggests that chromatin machinery controlling senescence reprogramming is significant in cancer stem cell formation.
    Keywords:  cancer stem cells; cellular senescence; chromatin; epigenetics; reprogramming