bims-cagime Biomed News
on Cancer, aging and metabolism
Issue of 2020‒08‒23
77 papers selected by
Kıvanç Görgülü
Technical University of Munich

  1. Nature. 2020 Aug 19.
    Gomes AP, Ilter D, Low V, Endress JE, Fernández-García J, Rosenzweig A, Schild T, Broekaert D, Ahmed A, Planque M, Elia I, Han J, Kinzig C, Mullarky E, Mutvei AP, Asara J, de Cabo R, Cantley LC, Dephoure N, Fendt SM, Blenis J.
      The risk of cancer and associated mortality increases substantially in humans from the age of 65 years onwards1-6. Nonetheless, our understanding of the complex relationship between age and cancer is still in its infancy2,3,7,8. For decades, this link has largely been attributed to increased exposure time to mutagens in older individuals. However, this view does not account for the established role of diet, exercise and small molecules that target the pace of metabolic ageing9-12. Here we show that metabolic alterations that occur with age can produce a systemic environment that favours the progression and aggressiveness of tumours. Specifically, we show that methylmalonic acid (MMA), a by-product of propionate metabolism, is upregulated in the serum of older people and functions as a mediator of tumour progression. We traced this to the ability of MMA to induce SOX4 expression and consequently to elicit transcriptional reprogramming that can endow cancer cells with aggressive properties. Thus, the accumulation of MMA represents a link between ageing and cancer progression, suggesting that MMA is a promising therapeutic target for advanced carcinomas.
  2. Clin Sci (Lond). 2020 Aug 28. 134(16): 2091-2115
    van Duijneveldt G, Griffin MDW, Putoczki TL.
      Pancreatic cancer has one of the poorest prognoses of all malignancies, with little improvement in clinical outcome over the past 40 years. Pancreatic ductal adenocarcinoma is responsible for the vast majority of pancreatic cancer cases, and is characterised by the presence of a dense stroma that impacts therapeutic efficacy and drives pro-tumorigenic programs. More specifically, the inflammatory nature of the tumour microenvironment is thought to underlie the loss of anti-tumour immunity and development of resistance to current treatments. Inflammatory pathways are largely mediated by the expression of, and signalling through, cytokines, chemokines, and other cellular messengers. In recent years, there has been much attention focused on dual targeting of cancer cells and the tumour microenvironment. Here we review our current understanding of the role of IL-6, and the broader IL-6 cytokine family, in pancreatic cancer, including their contribution to pancreatic inflammation and various roles in pancreatic cancer pathogenesis. We also summarise potential opportunities for therapeutic targeting of these pathways as an avenue towards combating poor patient outcomes.
    Keywords:  Cytokine; Interleukin; Pancreatic
  3. Cell. 2020 Aug 13. pii: S0092-8674(20)30929-6. [Epub ahead of print]
    Pfitzner AK, Mercier V, Jiang X, Moser von Filseck J, Baum B, Šarić A, Roux A.
      The endosomal sorting complex required for transport-III (ESCRT-III) catalyzes membrane fission from within membrane necks, a process that is essential for many cellular functions, from cell division to lysosome degradation and autophagy. How it breaks membranes, though, remains unknown. Here, we characterize a sequential polymerization of ESCRT-III subunits that, driven by a recruitment cascade and by continuous subunit-turnover powered by the ATPase Vps4, induces membrane deformation and fission. During this process, the exchange of Vps24 for Did2 induces a tilt in the polymer-membrane interface, which triggers transition from flat spiral polymers to helical filament to drive the formation of membrane protrusions, and ends with the formation of a highly constricted Did2-Ist1 co-polymer that we show is competent to promote fission when bound on the inside of membrane necks. Overall, our results suggest a mechanism of stepwise changes in ESCRT-III filament structure and mechanical properties via exchange of the filament subunits to catalyze ESCRT-III activity.
    Keywords:  CHMP1; CHMP4; Did2; ESCRT; ESCRT-III; Ist1; Snf7; Vps2; in vitro reconstitution; membrane fission; membrane remodeling
  4. Cancer Res. 2020 Aug 19. pii: canres.1255.2020. [Epub ahead of print]
    Rozeveld CN, Johnson KM, Zhang L, Razidlo GL.
      Oncogene-induced metabolic reprogramming is a hallmark of pancreatic cancer (PDAC), yet the metabolic drivers of metastasis are unclear. In PDAC, obesity and excess fatty acids accelerate tumor growth and increase metastasis. Here, we report that excess lipids, stored in organelles called lipid droplets (LD), are a key resource to fuel the energy-intensive process of metastasis. The oncogene KRAS controlled the storage and utilization of LD through regulation of hormone sensitive lipase (HSL), which was downregulated in human PDAC. Disruption of the KRAS-HSL axis reduced lipid storage, reprogrammed tumor cell metabolism, and inhibited invasive migration in vitro and metastasis in vivo. Finally, microscopy-based metabolic analysis revealed that migratory cells selectively utilize oxidative metabolism during the process of migration to metabolize stored lipids and fuel invasive migration. Taken together, these results reveal a mechanism that can be targeted to attenuate PDAC metastasis.
  5. Proc Natl Acad Sci U S A. 2020 Aug 14. pii: 202005976. [Epub ahead of print]
    Vais H, Payne R, Paudel U, Li C, Foskett JK.
      Ca2+ uptake by mitochondria regulates bioenergetics, apoptosis, and Ca2+ signaling. The primary pathway for mitochondrial Ca2+ uptake is the mitochondrial calcium uniporter (MCU), a Ca2+-selective ion channel in the inner mitochondrial membrane. MCU-mediated Ca2+ uptake is driven by the sizable inner-membrane potential generated by the electron-transport chain. Despite the large thermodynamic driving force, mitochondrial Ca2+ uptake is tightly regulated to maintain low matrix [Ca2+] and prevent opening of the permeability transition pore and cell death, while meeting dynamic cellular energy demands. How this is accomplished is controversial. Here we define a regulatory mechanism of MCU-channel activity in which cytoplasmic Ca2+ regulation of intermembrane space-localized MICU1/2 is controlled by Ca2+-regulatory mechanisms localized across the membrane in the mitochondrial matrix. Ca2+ that permeates through the channel pore regulates Ca2+ affinities of coupled inhibitory and activating sensors in the matrix. Ca2+ binding to the inhibitory sensor within the MCU amino terminus closes the channel despite Ca2+ binding to MICU1/2. Conversely, disruption of the interaction of MICU1/2 with the MCU complex disables matrix Ca2+ regulation of channel activity. Our results demonstrate how Ca2+ influx into mitochondria is tuned by coupled Ca2+-regulatory mechanisms on both sides of the inner mitochondrial membrane.
    Keywords:  EMRE; MICU1; calcium; electrophysiology; mitochondria
  6. Sci Immunol. 2020 Aug 21. pii: eabb4432. [Epub ahead of print]5(50):
    Boland BS, He Z, Tsai MS, Olvera JG, Omilusik KD, Duong HG, Kim ES, Limary AE, Jin W, Milner JJ, Yu B, Patel SA, Louis TL, Tysl T, Kurd NS, Bortnick A, Quezada LK, Kanbar JN, Miralles A, Huylebroeck D, Valasek MA, Dulai PS, Singh S, Lu LF, Bui JD, Murre C, Sandborn WJ, Goldrath AW, Yeo GW, Chang JT.
      Inflammatory bowel disease (IBD) encompasses a spectrum of gastrointestinal disorders driven by dysregulated immune responses against gut microbiota. We integrated single-cell RNA and antigen receptor sequencing to elucidate key components, cellular states, and clonal relationships of the peripheral and gastrointestinal mucosal immune systems in health and ulcerative colitis (UC). UC was associated with an increase in IgG1+ plasma cells in colonic tissue, increased colonic regulatory T cells characterized by elevated expression of the transcription factor ZEB2, and an enrichment of a γδ T cell subset in the peripheral blood. Moreover, we observed heterogeneity in CD8+ tissue-resident memory T (TRM) cells in colonic tissue, with four transcriptionally distinct states of differentiation observed across health and disease. In the setting of UC, there was a marked shift of clonally related CD8+ TRM cells toward an inflammatory state, mediated, in part, by increased expression of the T-box transcription factor Eomesodermin. Together, these results provide a detailed atlas of transcriptional changes occurring in adaptive immune cells in the context of UC and suggest a role for CD8+ TRM cells in IBD.
  7. Autophagy. 2020 Aug 18. 1-2
    Tsang YH, Mills GB.
      The melanoma-associated antigen family A (MAGEA) antigens are expressed in a wide variety of malignant tumors but not in adult somatic cells, rendering them attractive targets for cancer immunotherapy. Recent studies uncovered a role for MAGEA6 in suppression of macroautophagy/autophagy implicating MAGEA6 in tumorigenesis. The impact of cancer-associated MAGEA6 mutations on tumor pathophysiology are less well explored. In pancreatic cancer cell models, MAGEA6 inhibits autophagy, facilitating pancreatic cancer initiation. However, autophagy places a brake on cancer progression and is released upon MAGEA6 degradation, which can be induced by nutrient deficiency or by acquisition of cancer-associated mutations that reinstitute autophagy. Further cancer-associated mutations of the broader MAGEA genes frequently result in degradation of the corresponding protein products by proteasome-dependent machinery, potentially jeopardizing the utility of MAGEA genes as immunotherapeutic targets. Altogether, our findings provide mechanistic insight into the divergent roles of MAGEA6 during pancreatic cancer initiation and progression, and could inform cancer immunotherapeutic strategies for targeting MAGEA antigens.
    Keywords:   MAGEA6 ; Autophagy; cancer; degradation; mutation; oncogene; pancreatic ductal adenocarcinoma; proteasome and tumor suppressor
  8. Autophagy. 2020 Aug 19.
    Han S, Jeong YY, Sheshadri P, Cai Q.
      Mitochondria sustain various essential functions at synaptic terminals. Synaptic mitochondria deficits have been implicated in early Alzheimer disease (AD) pathophysiology. Mitophagy, a selective autophagy for removal of damaged mitochondria, plays a key role in mitochondrial quality control in neurons. However, fundamental questions remain unanswered as to whether mitophagy regulates synaptic mitochondrial integrity and whether AD-associated early deficits in synaptic mitochondria are attributed to mitophagy failure. We have recently revealed that the integrity of synaptic mitochondria is maintained by a coordination of RHEB-mediated mitophagy with dynein- and SNAPIN-driven retrograde transport. We demonstrate that increased mitophagy initiation, coupled with defective retrograde transport, triggers mitophagy stress at AD synapses. Excitingly, SNAPIN-enhanced retrograde transport reduces synaptic mitophagy stress and ameliorates mitochondrial deficits, thereby counteracting synaptic damage in AD mouse brains. Therefore, our study provides new mechanistic insights into how mitophagy facilitates synaptic mitochondrial maintenance and how mitophagy failure exacerbates AD-linked mitochondrial defects and synaptic degeneration.
    Keywords:  Alzheimer; Nix; PRKN; RHEB; SNAPIN; mitophagosome; retrograde transport; synaptic degeneration; synaptic mitochondrial deficits; synaptic mitophagy
  9. J Biomol NMR. 2020 Aug 17.
    Marshall CB, KleinJan F, Gebregiworgis T, Lee KY, Fang Z, Eves BJ, Liu NF, Gasmi-Seabrook GMC, Enomoto M, Ikura M.
      Mutations in RAS oncogenes occur in ~ 30% of human cancers, with KRAS being the most frequently altered isoform. RAS proteins comprise a conserved GTPase domain and a C-terminal lipid-modified tail that is unique to each isoform. The GTPase domain is a 'switch' that regulates multiple signaling cascades that drive cell growth and proliferation when activated by binding GTP, and the signal is terminated by GTP hydrolysis. Oncogenic RAS mutations disrupt the GTPase cycle, leading to accumulation of the activated GTP-bound state and promoting proliferation. RAS is a key target in oncology, however it lacks classic druggable pockets and has been extremely challenging to target. RAS signaling has thus been targeted indirectly, by harnessing key downstream effectors as well as upstream regulators, or disrupting the proper membrane localization required for signaling, by inhibiting either lipid modification or 'carrier' proteins. As a small (20 kDa) protein with multiple conformers in dynamic equilibrium, RAS is an excellent candidate for NMR-driven characterization and screening for direct inhibitors. Several molecules have been discovered that bind RAS and stabilize shallow pockets through conformational selection, and recent compounds have achieved substantial improvements in affinity. NMR-derived insight into targeting the RAS-membrane interface has revealed a new strategy to enhance the potency of small molecules, while another approach has been development of peptidyl inhibitors that bind through large interfaces rather than deep pockets. Remarkable progress has been made with mutation-specific covalent inhibitors that target the thiol of a G12C mutant, and these are now in clinical trials. Here we review the history of RAS inhibitor development and highlight the utility of NMR and integrated biophysical approaches in RAS drug discovery.
    Keywords:  Conformational selection; Drug discovery; KRAS; Membrane-associated protein; NMR; Oncogene; Prenylation
  10. Ageing Res Rev. 2020 Aug 16. pii: S1568-1637(20)30277-4. [Epub ahead of print] 101142
    Santoro A, Martucci M, Conte M, Capri M, Franceschi C, Salvioli S.
      We propose in this review that hormesis, a concept profoundly and systematically addressed by Mark Mattson, has to be considered a sort of comprehensive "contact point" capable of unifying several conceptualizations of the aging process, including those focused on the stress response, oxidative stress and chronic inflammation/inflammaging. A major strength of hormesis and inflammaging is that both have a strong evolutionary basis. Moreover, both hormesis and inflammaging frame the aging process within a lifelong perspective of adaptation to different types of stresses. Such adaptation perspective also suggests that the aging process is malleable, and predicts that effective anti-aging strategies should mimic what evolution did on the course of million years and that we have to learn how to exploit the great potential inherent in the hormetic/inflammatory responses. To this regard, new topics such as the production of mitokines to cope with mitochondrial dysfunction are emerging as possible anti-aging target. This approach opens theoretically the door to the possibility of modulate the individual aging rate and trajectory by adopting the most effective scientifically-based lifestyle regarding fundamentally nutrition and physical activity. In this scenario Mark Mattson's lesson and personal example will permanently enlighten the aging field and the quest for a healthy aging and longevity.
    Keywords:  Inflammaging; adaptation; aging; mitokines; stress
  11. Nat Cancer. 2020 May;1(5): 546-561
    Ghorani E, Reading JL, Henry JY, de Massy MR, Rosenthal R, Turati V, Joshi K, Furness AJS, Aissa AB, Saini SK, Ramskov S, Georgiou A, Sunderland MW, Wong YNS, De Mucha MV, Day W, Galvez-Cancino F, Becker PD, Uddin I, Ismail M, Ronel T, Woolston A, Jamal-Hanjani M, Veeriah S, Birkbak NJ, Wilson GA, Litchfield K, Conde L, Guerra-Assunção JA, Blighe K, Biswas D, Salgado R, Lund T, Al Bakir M, Moore DA, Hiley CT, Loi S, Sun Y, Yuan Y, AbdulJabbar K, Turajilic S, Herrero J, Enver T, Hadrup SR, Hackshaw A, Peggs KS, McGranahan N, Chain B, Swanton C, Quezada SA.
      Tumour mutational burden (TMB) predicts immunotherapy outcome in non-small cell lung cancer (NSCLC), consistent with immune recognition of tumour neoantigens. However, persistent antigen exposure is detrimental for T cell function. How TMB affects CD4 and CD8 T cell differentiation in untreated tumours, and whether this affects patient outcomes is unknown. Here we paired high-dimensional flow cytometry, exome, single-cell and bulk RNA sequencing from patients with resected, untreated NSCLC to examine these relationships. TMB was associated with compartment-wide T cell differentiation skewing, characterized by loss of TCF7-expressing progenitor-like CD4 T cells, and an increased abundance of dysfunctional CD8 and CD4 T cell subsets, with significant phenotypic and transcriptional similarity to neoantigen-reactive CD8 T cells. A gene signature of redistribution from progenitor-like to dysfunctional states associated with poor survival in lung and other cancer cohorts. Single-cell characterization of these populations informs potential strategies for therapeutic manipulation in NSCLC.
  12. Pharmacol Res. 2020 Aug 15. pii: S1043-6618(20)31439-0. [Epub ahead of print]161 105131
    Ma Y, Zhang S, Jin Z, Shi M.
      Besides acting as principle cellular building blocks and energy reservoirs, lipids also carry important signals associated with many fundamental cell biological processes, such as proliferation, differentiation, migration, stress responses and cell demise. Hyperactive lipid metabolism is closely associated with cancer progression and unfavorable outcomes. The underlying mechanisms are being gradually deciphered. In this review, we aim to summarize recent advances on how reprogrammed lipid metabolism and accompanying signaling cascades directly modulate cancer cells, as well as influencing stromal cells and immune cells within the tumor microenvironment. For future studies, special attention should be paid to lipid-mediated crosstalk among cancer cells, their neighboring stromal cells, and immune cells, plus how these multi-level communications determine anti-tumor immunity and bring novel immunotherapeutic opportunities.
    Keywords:  Cancer therapy; Immune regulation; Lipid metabolism; Stress response; Tumor microenvironment
  13. Oncotarget. 2020 Aug 04. 11(31): 3013-3024
    Turcios L, Marti F, Watt DS, Kril LM, Khurana A, Chapelin F, Liu C, Zwischenberger JB, Evers BM, Gedaly R.
      BACKGROUND: Hepatocellular Carcinoma (HCC) is the third most common cause of cancer related death worldwide. Adequate treatment options for patients with advanced HCC are currently limited.MATERIALS AND METHODS: We studied the anti-HCC effect of FH535 and a novel derivative Y3, on proliferation, mitochondrial function and cellular metabolism focusing on the three key substrates, glutamine, glucose, and fatty acids.
    RESULTS: FH535 and Y3 disrupted mitochondrial redox control in HCC cells that resulted from uncoupling mechanisms that increased proton leakage and decreased ATP production leading to apoptosis. The uncoupling effects of the sulfonamides in HCC cells were supported by the loss of activity of the methylated analogs. The accumulation of ROS significantly contributed to cell damage after the impaired autophagic machinery. These sulfonamides, FH535 and Y3, targeted glutamine and fatty acid metabolism and caused HCC cell reprograming towards the preferential use of glucose and the glycolytic pathway.
    CONCLUSIONS: FH535, and Y3, demonstrated potent anti-HCC activity by targeting OXPHOS, increasing dangerous levels of ROS and reducing ATP production. These sulfonamides target glutamine and FA metabolic pathways significantly increasing the cellular dependency on glycolysis.
    Keywords:  Wnt/β-catenin pathway; glutamine metabolism; hepatocellular carcinoma; mitochondria; proton uncouplers
  14. Cancer Discov. 2020 Aug 19. pii: CD-20-0142. [Epub ahead of print]
    Hofmann MH, Gmachl M, Ramharter J, Savarese F, Gerlach D, Marszalek JR, Sanderson MP, Kessler D, Trapani F, Arnhof H, Rumpel K, Botesteanu DA, Ettmayer P, Gerstberger T, Kofink C, Wunberg T, Zoephel A, Fu SC, Teh JL, Bottcher J, Pototschnig N, Schachinger F, Schipany K, Lieb S, Vellano CP, O'Connell JC, Mendes RL, Moll J, Petronczki M, Heffernan TP, Pearson M, McConnell DB, Kraut N.
      KRAS is the most frequently mutated driver of pancreatic, colorectal, and non-small cell lung cancers. Direct KRAS blockade has proven challenging and inhibition of a key downstream effector pathway, the RAF-MEK-ERK cascade, has shown limited success due to activation of feedback networks that keep the pathway in check. We hypothesized that inhibiting SOS1, a KRAS activator and important feedback node, represents an effective approach to treat KRAS-driven cancers. We report the discovery of a highly potent, selective and orally bioavailable small-molecule SOS1 inhibitor, BI-3406, that binds to the catalytic domain of SOS1 thereby preventing the interaction with KRAS. BI-3406 reduces formation of GTP-loaded RAS and limits cellular proliferation of a broad range of KRAS-driven cancers. Importantly, BI-3406 attenuates feedback reactivation induced by MEK inhibitors and thereby enhances sensitivity of KRAS-dependent cancers to MEK inhibition. Combined SOS1 and MEK inhibition represents a novel and effective therapeutic concept to address KRAS-driven tumors.
  15. Cell. 2020 Aug 17. pii: S0092-8674(20)30946-6. [Epub ahead of print]
    Sadik A, Somarribas Patterson LF, Öztürk S, Mohapatra SR, Panitz V, Secker PF, Pfänder P, Loth S, Salem H, Prentzell MT, Berdel B, Iskar M, Faessler E, Reuter F, Kirst I, Kalter V, Foerster KI, Jäger E, Guevara CR, Sobeh M, Hielscher T, Poschet G, Reinhardt A, Hassel JC, Zapatka M, Hahn U, von Deimling A, Hopf C, Schlichting R, Escher BI, Burhenne J, Haefeli WE, Ishaque N, Böhme A, Schäuble S, Thedieck K, Trump S, Seiffert M, Opitz CA.
      Aryl hydrocarbon receptor (AHR) activation by tryptophan (Trp) catabolites enhances tumor malignancy and suppresses anti-tumor immunity. The context specificity of AHR target genes has so far impeded systematic investigation of AHR activity and its upstream enzymes across human cancers. A pan-tissue AHR signature, derived by natural language processing, revealed that across 32 tumor entities, interleukin-4-induced-1 (IL4I1) associates more frequently with AHR activity than IDO1 or TDO2, hitherto recognized as the main Trp-catabolic enzymes. IL4I1 activates the AHR through the generation of indole metabolites and kynurenic acid. It associates with reduced survival in glioma patients, promotes cancer cell motility, and suppresses adaptive immunity, thereby enhancing the progression of chronic lymphocytic leukemia (CLL) in mice. Immune checkpoint blockade (ICB) induces IDO1 and IL4I1. As IDO1 inhibitors do not block IL4I1, IL4I1 may explain the failure of clinical studies combining ICB with IDO1 inhibition. Taken together, IL4I1 blockade opens new avenues for cancer therapy.
    Keywords:  AHR; CLL; IL4I1; T cell exhaustion; adaptive immunity; aryl hydrocarbon receptor; interleukin 4 induced 1; kynurenic acid; tryptophan metabolism; tumor micro-environment
  16. Elife. 2020 Aug 21. pii: e56889. [Epub ahead of print]9
    McGilvray PT, Anghel SA, Sundaram A, Zhong F, Trnka MJ, Fuller JR, Hu H, Burlingame AL, Keenan RJ.
      Membrane proteins with multiple transmembrane domains play critical roles in cell physiology, but little is known about the machinery coordinating their biogenesis at the endoplasmic reticulum. Here we describe a ~360 kDa ribosome-associated complex comprising the core Sec61 channel and five accessory factors: TMCO1, CCDC47 and the Nicalin-TMEM147-NOMO complex. Cryo-electron microscopy reveals a large assembly at the ribosome exit tunnel organized around a central membrane cavity. Similar to protein-conducting channels that facilitate movement of transmembrane segments, cytosolic and luminal funnels in TMCO1 and TMEM147, respectively, suggest routes into the central membrane cavity. High-throughput mRNA sequencing shows selective translocon engagement with hundreds of different multi-pass membrane proteins. Consistent with a role in multi-pass membrane protein biogenesis, cells lacking different accessory components show reduced levels of one such client, the glutamate transporter EAAT1. These results identify a new human translocon and provide a molecular framework for understanding its role in multi-pass membrane protein biogenesis.
    Keywords:  biochemistry; cell biology; chemical biology; human
  17. Nat Commun. 2020 Aug 17. 11(1): 4128
    Castro A, Pyke RM, Zhang X, Thompson WK, Day CP, Alexandrov LB, Zanetti M, Carter H.
      Individual MHC genotype constrains the mutational landscape during tumorigenesis. Immune checkpoint inhibition reactivates immunity against tumors that escaped immune surveillance in approximately 30% of cases. Recent studies demonstrated poorer response rates in female and younger patients. Although immune responses differ with sex and age, the role of MHC-based immune selection in this context is unknown. We find that tumors in younger and female individuals accumulate more poorly presented driver mutations than those in older and male patients, despite no differences in MHC genotype. Younger patients show the strongest effects of MHC-based driver mutation selection, with younger females showing compounded effects and nearly twice as much MHC-II based selection. This study presents evidence that strength of immune selection during tumor development varies with sex and age, and may influence the availability of mutant peptides capable of driving effective response to immune checkpoint inhibitor therapy.
  18. Nature. 2020 Aug 19.
    Chitwood PJ, Hegde RS.
      Integral membrane proteins are encoded by approximately 25% of all protein-coding genes1. In eukaryotes, the majority of membrane proteins are inserted, modified and folded at the endoplasmic reticulum (ER)2. Research over the past several decades has determined how membrane proteins are targeted to the ER and how individual transmembrane domains (TMDs) are inserted into the lipid bilayer3. By contrast, very little is known about how multi-spanning membrane proteins with several TMDs are assembled within the membrane. During the assembly of TMDs, interactions between polar or charged amino acids typically stabilize the final folded configuration4-8. TMDs with hydrophilic amino acids are likely to be chaperoned during the co-translational biogenesis of membrane proteins; however, ER-resident intramembrane chaperones are poorly defined. Here we identify the PAT complex, an abundant obligate heterodimer of the widely conserved ER-resident membrane proteins CCDC47 and Asterix. The PAT complex engages nascent TMDs that contain unshielded hydrophilic side chains within the lipid bilayer, and it disengages concomitant with substrate folding. Cells that lack either subunit of the PAT complex show reduced biogenesis of numerous multi-spanning membrane proteins. Thus, the PAT complex is an intramembrane chaperone that protects TMDs during assembly to minimize misfolding of multi-spanning membrane proteins and maintain cellular protein homeostasis.
  19. Nat Chem Biol. 2020 Aug 17.
    Kim KP, Choi J, Yoon J, Bruder JM, Shin B, Kim J, Arauzo-Bravo MJ, Han D, Wu G, Han DW, Kim J, Cramer P, Schöler HR.
      Identifying molecular and cellular processes that regulate reprogramming competence of transcription factors broadens our understanding of reprogramming mechanisms. In the present study, by a chemical screen targeting major epigenetic pathways in human reprogramming, we discovered that inhibiting specific epigenetic roadblocks including disruptor of telomeric silencing 1-like (DOT1L)-mediated H3K79/K27 methylation, but also other epigenetic pathways, catalyzed by lysine-specific histone demethylase 1A, DNA methyltransferases and histone deacetylases, allows induced pluripotent stem cell generation with almost all OCT factors. We found that simultaneous inhibition of these pathways not only dramatically enhances reprogramming competence of most OCT factors, but in fact enables dismantling of species-dependent reprogramming competence of OCT6, NR5A1, NR5A2, TET1 and GATA3. Harnessing these induced permissive epigenetic states, we performed an additional screen with 98 candidate genes. Thereby, we identified 25 transcriptional regulators (OTX2, SIX3, and so on) that can functionally replace OCT4 in inducing pluripotency. Our findings provide a conceptual framework for understanding how transcription factors elicit reprogramming in dependency of the donor cell epigenome that differs across species.
  20. J Biol Chem. 2020 Aug 19. pii: jbc.AC120.014993. [Epub ahead of print]
    Perez M, Bak DW, Bergholtz SE, Crooks DR, Arimilli BS, Yang Y, Weerapana E, Linehan WM, Meier JL.
      An important context in which metabolism influences tumorigenesis is the genetic cancer syndrome hereditary leiomyomatosis and renal cell carcinoma (HLRCC), a disease in which mutation of the TCA cycle enzyme fumarate hydratase (FH) causes hyperaccumulation of fumarate. This electrophilic oncometabolite can alter gene activity at the level of transcription, via reversible inhibition of epigenetic dioxygenases, as well as posttranslationally, via covalent modification of cysteine residues. To better understand the potential for metabolites to influence posttranslational modifications important to tumorigenesis and cancer cell growth, here we report a chemoproteomic analysis of a kidney-derived HLRCC cell line. Using a general reactivity probe, we generated a dataset of proteomic cysteine residues sensitive to the reduction in fumarate levels caused by genetic re-introduction of active FH into HLRCC cell lines. This revealed a broad upregulation of cysteine reactivity upon FH rescue, which evidence suggests is caused by an approximately equal proportion of transcriptional and posttranslational modification-mediated regulation. Gene ontology analysis highlighted several new targets and pathways potentially modulated by FH mutation. Comparison of the new dataset to prior studies highlights considerable heterogeneity in the adaptive response of cysteine-containing proteins in different models of HLRCC. This is consistent with emerging studies indicating the existence of cell and tissue-specific cysteine-omes, further emphasizing the need for characterization of diverse models. Our analysis provides a resource for understanding the proteomic adaptation to fumarate accumulation, and a foundation for future efforts to exploit this knowledge for cancer therapy.
    Keywords:  chemical biology; inborn error of metabolism; metabolism; mitochondria; post-translational modification (PTM); proteomics; redox signaling; tricarboxylic acid cycle (TCA cycle) (Krebs cycle)
  21. Nat Commun. 2020 Aug 18. 11(1): 4150
    Yamamuro T, Kawabata T, Fukuhara A, Saita S, Nakamura S, Takeshita H, Fujiwara M, Enokidani Y, Yoshida G, Tabata K, Hamasaki M, Kuma A, Yamamoto K, Shimomura I, Yoshimori T.
      The systemic decline in autophagic activity with age impairs homeostasis in several tissues, leading to age-related diseases. A mechanistic understanding of adipocyte dysfunction with age could help to prevent age-related metabolic disorders, but the role of autophagy in aged adipocytes remains unclear. Here we show that, in contrast to other tissues, aged adipocytes upregulate autophagy due to a decline in the levels of Rubicon, a negative regulator of autophagy. Rubicon knockout in adipocytes causes fat atrophy and hepatic lipid accumulation due to reductions in the expression of adipogenic genes, which can be recovered by activation of PPARγ. SRC-1 and TIF2, coactivators of PPARγ, are degraded by autophagy in a manner that depends on their binding to GABARAP family proteins, and are significantly downregulated in Rubicon-ablated or aged adipocytes. Hence, we propose that age-dependent decline in adipose Rubicon exacerbates metabolic disorders by promoting excess autophagic degradation of SRC-1 and TIF2.
  22. Cancer Res. 2020 Aug 14. pii: canres.0285.2020. [Epub ahead of print]
    Gonda TA, Fang J, Salas M, Do C, Hsu E, Zhukovskaya A, Siegel A, Takahashi R, Lopez-Bujanda ZA, Drake CG, Manji GA, Wang TC, Olive KP, Tycko B.
      Pancreatic ductal adenocarcinoma (PDAC) is a lethal cancer that has proven refractory to immunotherapy. Previously, treatment with the DNA hypomethylating drug decitabine (5aza-dC; DAC) extended survival in the KPC-Brca1 mouse model of PDAC. Here we investigated the effects of DAC in the original KPC model and tested combination therapy with DAC followed by immune checkpoint inhibitors (ICI). Four protocols were tested: PBS vehicle, DAC, ICI (anti-PD-1 or anti-VISTA), and DAC followed by ICI. For each single-agent and combination treatment, tumor growth was measured by serial ultrasound, tumor infiltrating lymphoid and myeloid cells were characterized, and overall survival was assessed. Single-agent DAC led to increased CD4+ and CD8+ tumor-infiltrating T cells (TILs), PD1 expression, and tumor necrosis while slowing tumor growth and modestly increasing mouse survival without systemic toxicity. RNA-seq of DAC-treated tumors revealed increased expression of Chi3l3 (Ym1), reflecting an increase in a subset of tumor-infiltrating M2-polarized macrophages. While ICI alone had modest effects, DAC followed by either of ICI therapy additively inhibited tumor growth and prolonged mouse survival. The best results were obtained using DAC followed by anti-PD-1, which extended mean survival from 26 to 54 days (p<0.0001). In summary, low-dose DAC inhibits tumor growth and increases both TIL and a subset of tumor-infiltrating M2-polarized macrophages in the KPC model of PDAC, and DAC followed by anti-PD-1 substantially prolongs survival. Since M2-polarized macrophages are predicted to antagonize anti-tumor effects, targeting these cells may be important to enhance the efficacy of combination therapy with DAC plus ICI.
  23. Nat Immunol. 2020 Aug 17.
    Leinwand J, Miller G.
      Pancreatic ductal adenocarcinoma carries a dismal prognosis, and outcomes have improved little with modern therapeutics. Checkpoint-based immunotherapy has failed to elicit responses in the vast majority of patients with pancreatic cancer. Alongside tumor cell-intrinsic mechanisms associated with oncogenic KRAS-induced inflammation, the tolerogenic myeloid cell infiltrate has emerged as a critical impediment to adaptive antitumor immune responses. Furthermore, the discovery of an intratumoral microbiome and the elucidation of host-microbe interactions that curtail antitumor immunity also present opportunities for intervention. Here we review the mechanisms of immunotherapy resistance in pancreatic ductal adenocarcinoma and discuss strategies to directly augment T cell responses in parallel with myeloid cell- and microbiome-targeted approaches that may enable immune-mediated control of this malignancy.
  24. Science. 2020 Aug 21. 369(6506): 993-999
    Chin EN, Yu C, Vartabedian VF, Jia Y, Kumar M, Gamo AM, Vernier W, Ali SH, Kissai M, Lazar DC, Nguyen N, Pereira LE, Benish B, Woods AK, Joseph SB, Chu A, Johnson KA, Sander PN, Martínez-Peña F, Hampton EN, Young TS, Wolan DW, Chatterjee AK, Schultz PG, Petrassi HM, Teijaro JR, Lairson LL.
      Stimulator of interferon genes (STING) links innate immunity to biological processes ranging from antitumor immunity to microbiome homeostasis. Mechanistic understanding of the anticancer potential for STING receptor activation is currently limited by metabolic instability of the natural cyclic dinucleotide (CDN) ligands. From a pathway-targeted cell-based screen, we identified a non-nucleotide, small-molecule STING agonist, termed SR-717, that demonstrates broad interspecies and interallelic specificity. A 1.8-angstrom cocrystal structure revealed that SR-717 functions as a direct cyclic guanosine monophosphate-adenosine monophosphate (cGAMP) mimetic that induces the same "closed" conformation of STING. SR-717 displayed antitumor activity; promoted the activation of CD8+ T, natural killer, and dendritic cells in relevant tissues; and facilitated antigen cross-priming. SR-717 also induced the expression of clinically relevant targets, including programmed cell death 1 ligand 1 (PD-L1), in a STING-dependent manner.
  25. Cancer Discov. 2020 Aug 14. pii: CD-20-0047. [Epub ahead of print]
    Zhang Q, Luo J, Wu S, Si H, Gao C, Xu W, Abdullah SE, Higgs BW, Dennis PA, van der Heijden MS, Segal NH, Chaft JE, Hembrough T, Barrett JC, Hellmann MD.
      The utility of circulating tumor DNA (ctDNA) as a biomarker in patients with advanced cancers receiving immunotherapy is uncertain. We therefore analyzed pretreatment (n=978) and on-treatment (n=171) ctDNA samples across 16 advanced stage tumor types from three phase I/II trials of durvalumab (± anti-CTLA-4 therapy tremelimumab). Higher pretreatment variant allele frequencies (VAF) were associated with poorer overall survival and other known prognostic factors, but not objective response, suggesting a prognostic role for patient outcomes. On-treatment reductions in VAF and lower on-treatment VAF were independently associated with longer PFS and OS, and increased ORR, but not prognostic variables, suggesting that on-treatment ctDNA dynamics are predictive of benefit from immune checkpoint blockade. Accordingly, we propose a concept of "molecular response" using ctDNA, incorporating both pretreatment and on-treatment VAF that predicted long-term survival similarly to initial radiological response, while also permitting early differentiation of responders among patients with initially radiologically stable disease.
  26. Trends Immunol. 2020 Aug 13. pii: S1471-4906(20)30175-7. [Epub ahead of print]
    Li J, Stanger BZ.
      Reciprocal interactions between tumor cells and immune cells shape the tumor microenvironment. Recent studies indicate that enhanced cell cycle activity in cancer cells suppresses antitumor immunity. Herein we discuss potential mechanisms by which cell cycle programs intrinsic to tumor cells are coupled to immune behavior, with consequences for immunotherapy.
    Keywords:  cell cycle; immune evasion; immunotherapy; tumor immunology
  27. Sci Adv. 2020 Aug;6(32): eabc7288
    Ruan L, McNamara JT, Zhang X, Chang AC, Zhu J, Dong Y, Sun G, Peterson A, Na CH, Li R.
      Proteostasis declines with age, characterized by the accumulation of unfolded or damaged proteins. Recent studies suggest that proteins constituting pathological inclusions in neurodegenerative diseases also enter and accumulate in mitochondria. How unfolded proteins are managed within mitochondria remains unclear. Here, we found that excessive unfolded proteins in the mitochondrial matrix of yeast cells are consolidated into solid-phase inclusions, which we term deposits of unfolded mitochondrial proteins (DUMP). Formation of DUMP occurs in mitochondria near endoplasmic reticulum-mitochondria contact sites and is regulated by mitochondrial proteins controlling the production of cytidine 5'-diphosphate-diacylglycerol. DUMP formation is age dependent but accelerated by exogenous unfolded proteins. Many enzymes of the tricarboxylic acid cycle were enriched in DUMP. During yeast cell division, DUMP formation is necessary for asymmetric inheritance of damaged mitochondrial proteins between mother and daughter cells. We provide evidence that DUMP-like structures may be induced by excessive unfolded proteins in human cells.
  28. Nat Cell Biol. 2020 Aug 17.
    Nowosad A, Jeannot P, Callot C, Creff J, Perchey RT, Joffre C, Codogno P, Manenti S, Besson A.
      Autophagy is a catabolic process whereby cytoplasmic components are degraded within lysosomes, allowing cells to maintain energy homeostasis during nutrient depletion. Several studies reported that the CDK inhibitor p27Kip1 promotes starvation-induced autophagy by an unknown mechanism. Here we find that p27 controls autophagy via an mTORC1-dependent mechanism in amino acid-deprived cells. During prolonged starvation, a fraction of p27 is recruited to lysosomes, where it interacts with LAMTOR1, a component of the Ragulator complex required for mTORC1 activation. Binding of p27 to LAMTOR1 prevents Ragulator assembly and mTORC1 activation, promoting autophagy. Conversely, p27-/- cells exhibit elevated mTORC1 signalling as well as impaired lysosomal activity and autophagy. This is associated with cytoplasmic sequestration of TFEB, preventing induction of the lysosomal genes required for lysosome function. LAMTOR1 silencing or mTOR inhibition restores autophagy and induces apoptosis in p27-/- cells. Together, these results reveal a direct coordinated regulation between the cell cycle and cell growth machineries.
  29. EBioMedicine. 2020 Aug 17. pii: S2352-3964(20)30319-4. [Epub ahead of print]59 102943
    Missiroli S, Perrone M, Genovese I, Pinton P, Giorgi C.
      Mitochondria are dynamic organelles that have essential metabolic activity and are regarded as signalling hubs with biosynthetic, bioenergetics and signalling functions that orchestrate key biological pathways. However, mitochondria can influence all processes linked to oncogenesis, starting from malignant transformation to metastatic dissemination. In this review, we describe how alterations in the mitochondrial metabolic status contribute to the acquisition of typical malignant traits, discussing the most recent discoveries and the many unanswered questions. We also highlight that expanding our understanding of mitochondrial regulation and function mechanisms in the context of cancer cell metabolism could be an important task in biomedical research, thus offering the possibility of targeting mitochondria for the treatment of cancer.
    Keywords:  Calcium; Cancer; Metabolism; Mitochondria; ROS
  30. Trends Cell Biol. 2020 Aug 13. pii: S0962-8924(20)30144-6. [Epub ahead of print]
    Bakir B, Chiarella AM, Pitarresi JR, Rustgi AK.
      Cancer cell identity and plasticity are required in transition states, such as epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET), in primary tumor initiation, progression, and metastasis. The functional roles of EMT, MET, and the partial state (referred to as pEMT) may vary based on the type of tumor, the state of dissemination, and the degree of metastatic colonization. Herein, we review EMT, MET, pEMT, and plasticity in the context of tumor metastasis.
    Keywords:  EMT; MET; cellular plasticity; colonization; metastasis; pEMT
  31. Nature. 2020 Aug 19.
    Ubellacker JM, Tasdogan A, Ramesh V, Shen B, Mitchell EC, Martin-Sandoval MS, Gu Z, McCormick ML, Durham AB, Spitz DR, Zhao Z, Mathews TP, Morrison SJ.
      Cancer cells, including melanoma cells, often metastasize regionally through the lymphatic system before metastasizing systemically through the blood1-4; however, the reason for this is unclear. Here we show that melanoma cells in lymph experience less oxidative stress and form more metastases than melanoma cells in blood. Immunocompromised mice with melanomas derived from patients, and immunocompetent mice with mouse melanomas, had more melanoma cells per microlitre in tumour-draining lymph than in tumour-draining blood. Cells that metastasized through blood, but not those that metastasized through lymph, became dependent on the ferroptosis inhibitor GPX4. Cells that were pretreated with chemical ferroptosis inhibitors formed more metastases than untreated cells after intravenous, but not intralymphatic, injection. We observed multiple differences between lymph fluid and blood plasma that may contribute to decreased oxidative stress and ferroptosis in lymph, including higher levels of glutathione and oleic acid and less free iron in lymph. Oleic acid protected melanoma cells from ferroptosis in an Acsl3-dependent manner and increased their capacity to form metastatic tumours. Melanoma cells from lymph nodes were more resistant to ferroptosis and formed more metastases after intravenous injection than did melanoma cells from subcutaneous tumours. Exposure to the lymphatic environment thus protects melanoma cells from ferroptosis and increases their ability to survive during subsequent metastasis through the blood.
  32. Sci Immunol. 2020 Aug 21. pii: eabb5168. [Epub ahead of print]5(50):
    Aykut B, Chen R, Kim JI, Wu D, Shadaloey SAA, Abengozar R, Preiss P, Saxena A, Pushalkar S, Leinwand J, Diskin B, Wang W, Werba G, Berman M, Lee SKB, Khodadadi-Jamayran A, Saxena D, Coetzee WA, Miller G.
      Piezo1 is a mechanosensitive ion channel that has gained recognition for its role in regulating diverse physiological processes. However, the influence of Piezo1 in inflammatory disease, including infection and tumor immunity, is not well studied. We postulated that Piezo1 links physical forces to immune regulation in myeloid cells. We found signal transduction via Piezo1 in myeloid cells and established this channel as the primary sensor of mechanical stress in these cells. Global inhibition of Piezo1 with a peptide inhibitor was protective against both cancer and septic shock and resulted in a diminution in suppressive myeloid cells. Moreover, deletion of Piezo1 in myeloid cells protected against cancer and increased survival in polymicrobial sepsis. Mechanistically, we show that mechanical stimulation promotes Piezo1-dependent myeloid cell expansion by suppressing the retinoblastoma gene Rb1 We further show that Piezo1-mediated silencing of Rb1 is regulated via up-regulation of histone deacetylase 2. Collectively, our work uncovers Piezo1 as a targetable immune checkpoint that drives immunosuppressive myelopoiesis in cancer and infectious disease.
  33. ESMO Open. 2020 Aug;pii: e000818. [Epub ahead of print]4(Suppl 2):
    Taieb J, Abdallah R.
      Pancreatic adenocarcinoma (PA) represents 90% of solid pancreatic malignant tumours. With one of the worst prognoses in oncology (all stages 5-year overall survival (OS) of 9%), PA was the seventh-leading cause of cancer-related deaths worldwide in 2018, and during the last 20 years, there have been unexplained increases in its incidence and mortality.This article summarises how to manage, to our opinion, PA in everyday practice according to tumour staging into resectable, unresectable or metastatic disease. Surgery followed by consensual adjuvant chemotherapy is the first-intention treatment for resectable patients. Unresectable but non-metastatic PA should be treated with induction chemotherapy and optionally with chemoradiotherapy to enable when possible secondary surgical resection. First-line and second-line chemotherapy does improve quality of life and OS in the metastatic setting, FOLFIRINOX and gemcitabine + nab-paclitaxel being the two current standard first-line options. Molecular profiling of metastatic patients is emerging, as some personalised therapies are possible for rare subtypes such as MSI high, BRCA1-2 mutated and NRG1/NTRK fusion gene PA.
    Keywords:  borderline resectable pancreatic cancer; locally advanced pancreatic cancer; medical education; metastatic pancreatic cancer; pancreatic adnocarcinoma
  34. Cell Res. 2020 Aug 17.
    Gao K, Cheng M, Zuo X, Lin J, Hoogewijs K, Murphy MP, Fu XD, Zhang X.
      RNA interference (RNAi) has been thought to be a gene-silencing pathway present in most eukaryotic cells to safeguard the genome against retrotransposition. Small interfering RNAs (siRNAs) have also become a powerful tool for studying gene functions. Given the endosymbiotic hypothesis that mitochondria originated from prokaryotes, mitochondria have been generally assumed to lack active RNAi; however, certain bacteria have Argonaute homologs and various reports suggest the presence of specific microRNAs and nuclear genome (nDNA)-encoded Ago2 in the mitochondria. Here we report that transfected siRNAs are not only able to enter the matrix of mitochondria, but also function there to specifically silence targeted mitochondrial transcripts. The mitoRNAi effect is readily detectable at the mRNA level, but only recordable on relatively unstable proteins, such as the mtDNA-encoded complex IV subunits. We also apply mitoRNAi to directly determine the postulated crosstalk between individual respiratory chain complexes, and our result suggests that the controversial observations previously made in patient-derived cells might result from differential adaptation in different cell lines. Our findings bring a new tool to study mitochondrial biology.
  35. F1000Res. 2020 ;9 626
    Cohen IR, Marron A.
      The evolution of multicellular eukaryotes expresses two sorts of adaptations: local adaptations like fur or feathers, which characterize species in particular environments, and universal adaptations like microbiomes or sexual reproduction, which characterize most multicellulars in any environment. We reason that the mechanisms driving the universal adaptations of multicellulars should themselves be universal, and propose a mechanism based on properties of matter and systems: energy, entropy, and interaction. Energy from the sun, earth and beyond creates new arrangements and interactions. Metabolic networks channel some of this energy to form cooperating, interactive arrangements. Entropy, used here as a term for all forces that dismantle ordered structures (rather than as a physical quantity), acts as a selective force. Entropy selects for arrangements that resist it long enough to replicate, and dismantles those that do not. Interactions, energy-charged and dynamic, restrain entropy and enable survival and propagation of integrated living systems. This fosters survival-of-the-fitted - those entities that resist entropic destruction - and not only of the fittest - the entities with the greatest reproductive success. The "unit" of evolution is not a discrete entity, such as a gene, individual, or species; what evolves are collections of related interactions at multiple scales. Survival-of-the-fitted explains universal adaptations, including resident microbiomes, sexual reproduction, continuous diversification, programmed turnover, seemingly wasteful phenotypes, altruism, co-evolving environmental niches, and advancing complexity. Indeed survival-of-the-fittest may be a particular case of the survival-of-the-fitted mechanism, promoting local adaptations that express reproductive advantages in addition to resisting entropy. Survival-of-the-fitted accounts for phenomena that have been attributed to neutral evolution: in the face of entropy, there is no neutrality; all variations are challenged by ubiquitous energy and entropy, retaining those that are "fit enough". We propose experiments to test predictions of the survival-of-the-fitted theory, and discuss implications for the wellbeing of humans and the biosphere.
    Keywords:  Cooperation; Energy; Entropy; Evolution; Fitness; Fittedness; Interaction; Niche
  36. Proc Natl Acad Sci U S A. 2020 Aug 17. pii: 202003037. [Epub ahead of print]
    Shivers JL, Feng J, van Oosten ASG, Levine H, Janmey PA, MacKintosh FC.
      Tissues commonly consist of cells embedded within a fibrous biopolymer network. Whereas cell-free reconstituted biopolymer networks typically soften under applied uniaxial compression, various tissues, including liver, brain, and fat, have been observed to instead stiffen when compressed. The mechanism for this compression-stiffening effect is not yet clear. Here, we demonstrate that when a material composed of stiff inclusions embedded in a fibrous network is compressed, heterogeneous rearrangement of the inclusions can induce tension within the interstitial network, leading to a macroscopic crossover from an initial bending-dominated softening regime to a stretching-dominated stiffening regime, which occurs before and independently of jamming of the inclusions. Using a coarse-grained particle-network model, we first establish a phase diagram for compression-driven, stretching-dominated stress propagation and jamming in uniaxially compressed two- and three-dimensional systems. Then, we demonstrate that a more detailed computational model of stiff inclusions in a subisostatic semiflexible fiber network exhibits quantitative agreement with the predictions of our coarse-grained model as well as qualitative agreement with experiments.
    Keywords:  biopolymer networks; colloidal particles; compression stiffening; nonaffinity; tissues
  37. Cell Death Dis. 2020 Aug 03. 11(8): 648
    Díaz MI, Díaz P, Bennett JC, Urra H, Ortiz R, Orellana PC, Hetz C, Quest AFG.
      Caveolin-1 (CAV1), is a broadly expressed, membrane-associated scaffolding protein that acts both, as a tumor suppressor and a promoter of metastasis, depending on the type of cancer and stage. CAV1 is downregulated in human tumors, tumor cell lines and oncogene-transformed cells. The tumor suppressor activity of CAV1 is generally associated with its presence at the plasma membrane, where it participates, together with cavins, in the formation of caveolae and also has been suggested to interact with and inhibit a wide variety of proteins through interactions mediated by the scaffolding domain. However, a pool of CAV1 is also located at the endoplasmic reticulum (ER), modulating the secretory pathway in a manner dependent on serine-80 (S80) phosphorylation. In melanoma cells, CAV1 expression suppresses tumor formation, but the protein is largely absent from the plasma membrane and does not form caveolae. Perturbations to the function of the ER are emerging as a central driver of cancer, highlighting the activation of the unfolded protein response (UPR), a central pathway involved in stress mitigation. Here we provide evidence indicating that the expression of CAV1 represses the activation of the UPR in vitro and in solid tumors, reflected in the attenuation of PERK and IRE1α signaling. These effects correlated with increased susceptibility of cells to ER stress and hypoxia. Interestingly, the tumor suppressor activity of CAV1 was abrogated by site-directed mutagenesis of S80, correlating with a reduced ability to repress the UPR. We conclude that the tumor suppression by CAV1 involves the attenuation of the UPR, and identified S80 as essential in this context. This suggests that intracellular CAV1 regulates cancer through alternative signaling outputs.
  38. Nat Neurosci. 2020 Aug 17.
    Babetto E, Wong KM, Beirowski B.
      Axon degeneration is a hallmark of many neurodegenerative disorders. The current assumption is that the decision of injured axons to degenerate is cell-autonomously regulated. Here we show that Schwann cells (SCs), the glia of the peripheral nervous system, protect injured axons by virtue of a dramatic glycolytic upregulation that arises in SCs as an inherent adaptation to axon injury. This glycolytic response, paired with enhanced axon-glia metabolic coupling, supports the survival of axons. The glycolytic shift in SCs is largely driven by the metabolic signaling hub, mammalian target of rapamycin complex 1, and the downstream transcription factors hypoxia-inducible factor 1-alpha and c-Myc, which together promote glycolytic gene expression. The manipulation of glial glycolytic activity through this pathway enabled us to accelerate or delay the degeneration of perturbed axons in acute and subacute rodent axon degeneration models. Thus, we demonstrate a non-cell-autonomous metabolic mechanism that controls the fate of injured axons.
  39. Trends Endocrinol Metab. 2020 Aug 14. pii: S1043-2760(20)30139-9. [Epub ahead of print]
    Wang X, Wang Y, Antony V, Sun H, Liang G.
      Metabolic diseases pose a tremendous health threat in both developed and developing countries. The pathophysiology of metabolic diseases is complex but has been shown to be closely associated with sterile inflammation, which is initiated by various danger molecules derived from metabolic overload, such as oxidized low-density lipoproteins (OxLDLs), free fatty acids (FFAs), glucose, advanced glycation end products (AGEs), and cholesterol. These danger signals are sensed by pattern recognition receptors (PRRs) to activate proinflammatory signaling pathways and promote the release of proinflammatory mediators, leading to chronic low-grade inflammation. Although these harmful metabolic stimuli are generally regarded as damage-associated molecular patterns (DAMPs), a more specific definition and accurate classification for these DAMPs is still missing. In this opinion, we classify the harmful metabolic stimuli that can incite inflammatory responses and tissue damage via instigating PRRs as metabolism-associated molecular patterns (MAMPs), and we summarize their roles in metaflammation-mediated metabolic diseases.
    Keywords:  innate immunity; metabolic disease; metabolism-associated molecular patterns; metaflammation; pattern recognition receptors
  40. Elife. 2020 Aug 19. pii: e59166. [Epub ahead of print]9
    Walls JF, Subleski JJ, Palmieri EM, Gonzalez Cotto M, Gardiner CM, McVicar DW, Finlay DK.
      Natural Killer (NK) cells have an important role in immune responses to viruses and tumours. Integrating changes in signal transduction pathways and cellular metabolism is essential for effective NK cells responses. The glycolytic enzyme Pyruvate Kinase Muscle 2 (PKM2) has described roles in regulating glycolytic flux and signal transduction, particularly gene transcription. While PKM2 expression is robustly induced in activated NK cells, mice lacking PKM2 in NK cells showed no defect in NK cell metabolism, transcription or anti-viral responses to MCMV infection. NK cell metabolism was maintained due to compensatory PKM1 expression in PKM2-null NK cells. To further investigate the role of PKM2 we used TEPP-46, which increases PKM2 catalytic activity while inhibiting any PKM2 signalling functions. NK cells activated with TEPP-46 had reduced effector function due to TEPP-46-induced increases in oxidative stress. Overall, PKM2-regulated glycolytic metabolism and redox status, not transcriptional control, facilitate optimal NK cells responses.
    Keywords:  cell biology; immunology; inflammation; mouse
  41. Nat Genet. 2020 Aug 17.
    Kim H, Nguyen NP, Turner K, Wu S, Gujar AD, Luebeck J, Liu J, Deshpande V, Rajkumar U, Namburi S, Amin SB, Yi E, Menghi F, Schulte JH, Henssen AG, Chang HY, Beck CR, Mischel PS, Bafna V, Verhaak RGW.
      Extrachromosomal DNA (ecDNA) amplification promotes intratumoral genetic heterogeneity and accelerated tumor evolution1-3; however, its frequency and clinical impact are unclear. Using computational analysis of whole-genome sequencing data from 3,212 cancer patients, we show that ecDNA amplification frequently occurs in most cancer types but not in blood or normal tissue. Oncogenes were highly enriched on amplified ecDNA, and the most common recurrent oncogene amplifications arose on ecDNA. EcDNA amplifications resulted in higher levels of oncogene transcription compared to copy number-matched linear DNA, coupled with enhanced chromatin accessibility, and more frequently resulted in transcript fusions. Patients whose cancers carried ecDNA had significantly shorter survival, even when controlled for tissue type, than patients whose cancers were not driven by ecDNA-based oncogene amplification. The results presented here demonstrate that ecDNA-based oncogene amplification is common in cancer, is different from chromosomal amplification and drives poor outcome for patients across many cancer types.
  42. Science. 2020 Aug 20. pii: eabb9818. [Epub ahead of print]
    Humphries F, Shmuel-Galia L, Ketelut-Carneiro N, Li S, Wang B, Nemmara VV, Wilson R, Jiang Z, Khalighinejad F, Muneeruddin K, Shaffer SA, Dutta R, Ionete C, Pesiridis S, Yang S, Thompson PR, Fitzgerald KA.
      Activated macrophages undergo a metabolic switch to aerobic glycolysis accumulating Krebs cycle intermediates that alter transcription of immune response genes. Here we extend these observations by defining fumarate as an inhibitor of pyroptotic cell death. We found that dimethyl fumarate (DMF) delivered to cells or endogenous fumarate reacts with gasdermin D (GSDMD) at critical cysteine residues to form S-(2-succinyl)-cysteine. GSDMD succination prevents its interaction with caspases, limiting its processing, oligomerization, and capacity to induce cell death. In mice, the administration of DMF protects against LPS shock and alleviates familial Mediterranean fever and experimental autoimmune encephalitis (EAE) by targeting GSDMD. Collectively, these findings identify GSDMD as a target of fumarate and reveal a mechanism of action for fumarate-based therapeutics including DMF used to treat multiple sclerosis.
  43. Sci Immunol. 2020 Aug 21. pii: eabd6832. [Epub ahead of print]5(50):
    Maucourant C, Filipovic I, Ponzetta A, Aleman S, Cornillet M, Hertwig L, Strunz B, Lentini A, Reinius B, Brownlie D, Gomez AC, Ask EH, Hull RM, Haroun-Izquierdo A, Schaffer M, Klingström J, Folkesson E, Buggert M, Sandberg JK, Eriksson LI, Rooyackers O, Ljunggren HG, Malmberg KJ, Michaëlsson J, Marquardt N, Hammer Q, Strålin K, Björkström NK, .
      Understanding innate immune responses in COVID-19 is important to decipher mechanisms of host responses and interpret disease pathogenesis. Natural killer (NK) cells are innate effector lymphocytes that respond to acute viral infections but might also contribute to immunopathology. Using 28-color flow cytometry, we here reveal strong NK cell activation across distinct subsets in peripheral blood of COVID-19 patients. This pattern was mirrored in scRNA-seq signatures of NK cells in bronchoalveolar lavage from COVID-19 patients. Unsupervised high-dimensional analysis of peripheral blood NK cells furthermore identified distinct NK cell immunotypes that were linked to disease severity. Hallmarks of these immunotypes were high expression of perforin, NKG2C, and Ksp37, reflecting increased presence of adaptive NK cells in circulation of patients with severe disease. Finally, arming of CD56bright NK cells was observed across COVID-19 disease states, driven by a defined protein-protein interaction network of inflammatory soluble factors. This study provides a detailed map of the NK cell activation landscape in COVID-19 disease.
  44. Oncogene. 2020 Aug 17.
    Cho C, Mukherjee R, Peck AR, Sun Y, McBrearty N, Katlinski KV, Gui J, Govindaraju PK, Puré E, Rui H, Fuchs SY.
      Activation of cancer-associated fibroblasts (CAFs) and ensuing desmoplasia play an important role in the growth and progression of solid tumors. Here we demonstrate that, within colon and pancreatic ductal adenocarcinoma tumors, efficient stromagenesis relies on downregulation of the IFNAR1 chain of the type I interferon (IFN1) receptor. Expression of the fibroblast activation protein (FAP) and accumulation of the extracellular matrix (ECM) was notably impaired in tumors grown in the Ifnar1S526A (SA) knock-in mice, which are deficient in IFNAR1 downregulation. Primary fibroblasts from these mice exhibited elevated levels of Smad7, a negative regulator of the transforming growth factor-β (TGFβ) pathway. Knockdown of Smad7 alleviated deficient ECM production in SA fibroblasts in response to TGFβ. Analysis of human colorectal cancers revealed an inverse correlation between IFNAR1 and FAP levels. Whereas growth of tumors in SA mice was stimulated by co-injection of wild type but not SA fibroblasts, genetic ablation of IFNAR1 in fibroblasts also accelerated tumor growth. We discuss how inactivation of IFNAR1 in CAFs acts to stimulate stromagenesis and tumor growth.
  45. Clin Cancer Res. 2020 Aug 14. pii: clincanres.1439.2020. [Epub ahead of print]
    Wang Y, Park JYP, Pacis A, Denroche RE, Jang GH, Zhang A, Cuggia A, Domecq C, Monlong J, Raitses-Gurevich M, Grant RC, Borgida A, Holter S, Stossel C, Bu S, Masoomian M, Lungu IM, Bartlett JMS, Wilson JM, Gao ZH, Riazalhosseini Y, Asselah J, Bouganim N, Cabrera T, Boucher LM, Valenti D, Biagi J, Greenwood CM, Polak P, Foulkes WD, Golan T, O'Kane GM, Fischer SE, Knox JJ, Gallinger S, Zogopoulos G.
      PURPOSE: Pancreatic ductal adenocarcinoma (PDAC) arising in patients with a germline BRCA1 or BRCA2 (gBRCA) mutation may be sensitive to platinums and poly(adenosine diphosphate-ribose) polymerase inhibitors (PARPi). However, treatment stratification based on gBRCA mutational status alone is associated with heterogeneous responses.EXPERIMENTAL DESIGN: We performed a 7-arm preclinical trial consisting of 471 mice, representing 12 unique PDAC patient-derived xenografts, of which 9 were gBRCA-mutated. From 179 patients whose PDAC was whole genome and transcriptome sequenced, we identified 21 cases with homologous recombination deficiency (HRD), and investigated prognostic biomarkers.
    RESULTS: We found that biallelic inactivation of BRCA1/BRCA2 is associated with genomic hallmarks of HRD and required for cisplatin and talazoparib (PARPi) sensitivity. However, HRD genomic hallmarks persisted in xenografts despite the emergence of therapy resistance, indicating the presence of a genomic scar. We identified tumour polyploidy and a low Ki67 index as predictors of poor cisplatin and talazoparib response. In HRD PDAC patients, tumour polyploidy and a basal-like transcriptomic subtype were independent predictors of shorter survival. To facilitate clinical assignment of transcriptomic subtype, we developed a novel pragmatic two-marker assay (GATA6:KRT17).
    CONCLUSIONS: In summary, we propose a predictive and prognostic model of gBRCA-mutated PDAC based on HRD genomic hallmarks, Ki67 index, tumour ploidy and transcriptomic subtype.
  46. Proc Natl Acad Sci U S A. 2020 Aug 19. pii: 202008672. [Epub ahead of print]
    Cai W, Huang J, Zhu Q, Li BE, Seruggia D, Zhou P, Nguyen M, Fujiwara Y, Xie H, Yang Z, Hong D, Ren P, Xu J, Pu WT, Yuan GC, Orkin SH.
      How overall principles of cell-type-specific gene regulation (the "logic") may change during ontogeny is largely unexplored. We compared transcriptomic, epigenomic, and three-dimensional (3D) genomic profiles in embryonic (EryP) and adult (EryD) erythroblasts. Despite reduced chromatin accessibility compared to EryP, distal chromatin of EryD is enriched in H3K27ac, Gata1, and Myb occupancy. EryP-/EryD-shared enhancers are highly correlated with red blood cell identity genes, whereas cell-type-specific regulation employs different cis elements in EryP and EryD cells. In contrast to EryP-specific genes, which exhibit promoter-centric regulation through Gata1, EryD-specific genes rely more on distal enhancers for regulation involving Myb-mediated enhancer activation. Gata1 HiChIP demonstrated an overall increased enhancer-promoter interactions at EryD-specific genes, whereas genome editing in selected loci confirmed distal enhancers are required for gene expression in EryD but not in EryP. Applying a metric for enhancer dependence of transcription, we observed a progressive reliance on cell-specific enhancers with increasing ontogenetic age among diverse tissues of mouse and human origin. Our findings highlight fundamental and conserved differences at distinct developmental stages, characterized by simpler promoter-centric regulation of cell-type-specific genes in embryonic cells and increased combinatorial enhancer-driven control in adult cells.
    Keywords:  GATA1; HiChIP; Myb; enhancer; erythropoiesis
  47. Gut. 2020 Aug 21. pii: gutjnl-2020-321316. [Epub ahead of print]
    Camolotto SA, Belova VK, Torre-Healy L, Vahrenkamp JM, Berrett KC, Conway H, Shea J, Stubben C, Moffitt R, Gertz J, Snyder EL.
      OBJECTIVE: Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with a 5-year survival of less than 5%. Transcriptomic analysis has identified two clinically relevant molecular subtypes of PDAC: classical and basal-like. The classical subtype is characterised by a more favourable prognosis and better response to chemotherapy than the basal-like subtype. The classical subtype also expresses higher levels of lineage specifiers that regulate endodermal differentiation, including the nuclear receptor hepatocyte nuclear factor 4 α (HNF4α). The objective of this study is to evaluate the role of HNF4α, SIX4 and SIX1 in regulating the growth and molecular subtype of PDAC.DESIGN: We manipulate the expression of HNF4α, SIX4 and SIX1 in multiple in vitro and in vivo PDAC models. We determine the consequences of manipulating these genes on PDAC growth, differentiation and molecular subtype using functional assays, gene expression analysis and cross-species comparisons with human datasets.
    RESULTS: We show that HNF4α restrains tumour growth and drives tumour cells toward an epithelial identity. Gene expression analysis of murine models and human tumours shows that HNF4α activates expression of genes associated with the classical subtype. HNF4α also directly represses SIX4 and SIX1, two mesodermal/neuronal lineage specifiers expressed in the basal-like subtype. Finally, SIX4 and SIX1 drive proliferation and regulate differentiation in HNF4α-negative PDAC.
    CONCLUSION: Our data show that HNF4α regulates the growth and molecular subtype of PDAC by multiple mechanisms, including activation of the classical gene expression programme and repression of SIX4 and SIX1, which may represent novel dependencies of the basal-like subtype.
    Keywords:  gene expression; molecular mechanisms; pancreatic cancer
  48. Cancer Res. 2020 Aug 14. pii: canres.1829.2020. [Epub ahead of print]
    Lauko A, Mu Z, Gutmann DH, Naik UP, Lathia JD.
      Tight junction (TJ) proteins are essential for mediating interactions between adjacent cells and coordinating cellular and organ responses. Initial investigations into TJ proteins and junctional adhesion molecules (JAM) in cancer suggested a tumor suppressive role where decreased expression led to increased metastasis. However, recent studies of the JAM family members JAM-A and JAM-C have expanded the roles of these proteins to include pro-tumorigenic functions, including inhibition of apoptosis and promotion of proliferation, cancer stem cell biology, and epithelial-to-mesenchymal transition. JAM function by interacting with other proteins through three distinct molecular mechanisms: direct cell-cell interaction on adjacent cells, stabilization of adjacent cell surface receptors on the same cell, and interactions between JAM and cell surface receptors expressed on adjacent cells. Collectively, these diverse interactions contribute to both the pro- and anti-tumorigenic functions of JAM. In this review, we discuss these context-dependent functions of JAM in a variety of cancers and highlight key areas that remain poorly understood, including their potentially diverse intracellular signaling networks, their roles in the tumor microenvironment, and the consequences of post-translational modifications on their function. These studies have implications in furthering our understanding of JAM in cancer and provide a paradigm for exploring additional roles of TJ proteins.
  49. Autophagy. 2020 Aug 17.
    Chen X, Li J, Kang R, Klionsky DJ, Tang D.
      Ferroptosis is an iron-dependent, non-apoptotic form of regulated cell death caused by lipid peroxidation, which is controlled by integrated oxidation and antioxidant systems. The iron-containing enzyme lipoxygenase is the main promoter of ferroptosis by producing lipid hydroperoxides, and its function relies on the activation of ACSL4-dependent lipid biosynthesis. In contrast, the selenium-containing enzyme GPX4 is currently recognized as a central repressor of ferroptosis, and its activity depends on glutathione produced from the activation of the cystine-glutamate antiporter SLC7A11. Many metabolic (especially involving iron, lipids, and amino acids) and degradation pathways (macroautophagy/autophagy and the ubiquitin-proteasome system) orchestrate the complex ferroptotic response through direct or indirect regulation of iron accumulation or lipid peroxidation. Although the detailed mechanism of membrane injury during ferroptosis remains a mystery, ESCRT III-mediated plasma membrane repair can make cells resistant to ferroptosis. Here, we review the recent rapid progress in understanding the molecular mechanisms of ferroptosis and focus on the epigenetic, transcriptional, and posttranslational regulation of this process.
  50. Elife. 2020 Aug 17. pii: e58041. [Epub ahead of print]9
    Nowinski SM, Solmonson A, Rusin SF, Maschek JA, Bensard CL, Fogarty S, Jeong MY, Lettlova S, Berg JA, Morgan JT, Ouyang Y, Naylor BC, Paulo JA, Funai K, Cox JE, Gygi SP, Winge DR, DeBerardinis RJ, Rutter J.
      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.
    Keywords:  biochemistry; chemical biology; mouse
  51. J Cell Sci. 2020 Aug 14. pii: jcs.245555. [Epub ahead of print]
    Uemura S, Mochizuki T, Amemiya K, Kurosaka G, Yazawa M, Nakamoto K, Ishikawa Y, Izawa S, Abe F.
      Mechanical stresses including high hydrostatic pressure elicit diverse physiological effects on organisms. Gtr1/Gtr2 and Ego1/Ego3, central regulators of the TOR complex 1 (TORC1) nutrient signaling pathway, are required for the growth of Saccharomyces cerevisiae cells under high pressure. Here, we showed that a pressure of 25 MPa stimulates TORC1 to promote phosphorylation of Sch9, which depends on the EGO complex (EGOC) and Pib2. Incubation of cells at this pressure aberrantly increased the glutamine and alanine levels in the ego1Δ, gtr1Δ, tor1Δ, and pib2Δ mutants, whereas the polysome profiles were unaffected. Moreover, we found that glutamine levels were reduced by combined deletions of EGO1, GTR1, TOR1, and PIB2 with GLN3 These results suggested that high pressure leads to the intracellular accumulation of amino acids. Subsequently, Pib2 loaded with glutamine stimulates the EGOC-TORC1 complex to inactivate Gln3, downregulating glutamine synthesis. Our findings illustrated the regulatory circuit that maintained the intracellular amino acid homeostasis and suggested the critical roles the EGOC-TORC1 and Pib2-TORC1 complexes played in the growth of yeast under high hydrostatic pressure.
    Keywords:  EGO complex; Glutamine; Gtr1/Gtr2; High hydrostatic pressure; Pib2; Polysome profile; TORC1
  52. Methods Mol Biol. 2020 ;2184 225-231
    Xu YXZ, Ande SR, Mishra S.
      The resident immune cells (e.g., macrophages) present in major metabolic tissues, such as adipose and liver tissues, play crucial roles in respective tissue homeostasis through cross talk with metabolic tissues, and consequently in metabolic homeostasis at the systemic level, and their dysregulation contributes to metabolic dysregulation at large, as well as many associated diseases. Moreover, the cross talk between different resident immune cells and metabolic tissues is not limited to an intra-organ level but also includes interorgan cross talk, as they work in a coordinated manner throughout the body, such as in adipose tissue, skeletal muscle, and liver. Thus, it is important to determine the impact of altered immune functions on metabolic homeostasis and vice versa, to enhance our knowledge of immunometabolic biology. Glucose and insulin tolerance tests are simple methods that enable the measurement and analysis of the overall glucose homeostasis at the systemic level. Here we describe the process of performing metabolic tests for glucose homeostasis in mice, as mouse models are often used for defining the mechanistic underpinnings of physiology and pathophysiology related to immunometabolism, and in preclinical studies.
    Keywords:  Glucose metabolism; Glucose tolerance test; Immunometabolism; Insulin tolerance test; Meta-inflammation
  53. Chronobiol Int. 2020 Aug 20. 1-21
    Allison DB, Ren G, Peliciari-Garcia RA, Mia S, McGinnis GR, Davis J, Gamble KL, Kim JA, Young ME.
      Senescence-Accelerated Mouse-Prone 8 (SAMP8) mice exhibit characteristics of premature aging, including hair loss, cognitive dysfunction, reduced physical activity, impaired metabolic homeostasis, cardiac dysfunction and reduced lifespan. Interestingly, circadian disruption can induce or augment many of these same pathologies. Moreover, previous studies have reported that SAMP8 mice exhibit abnormalities in circadian wheel-running behavior, indicating possible alterations in circadian clock function. These observations led to the hypothesis that 24 h rhythms in behavior and/or circadian clock function are altered in SAMP8 mice and that these alterations may contribute to perturbations in whole-body metabolism. Here, we report that 6-month-old SAMP8 mice exhibit a more prominent biphasic pattern in daily behaviors (food intake and physical activity) and whole-body metabolism (energy expenditure, respiratory exchange ratio), relative to SAMR1 control mice. Consistent with a delayed onset of food intake at the end of the light phase, SAMP8 mice exhibit a phase delay (1.3-1.9 h) in 24 h gene expression rhythms of major circadian clock components (bmal1, rev-erbα, per2, dbp) in peripheral tissues (liver, skeletal muscle, white adipose tissue [WAT], brown adipose tissue [BAT]). Forcing mice to consume food only during the dark period improved alignment of both whole-body metabolism and oscillations in expression of clock genes in peripheral tissues between SAMP8 and SAMR1 mice. Next, interrogation of metabolic genes revealed altered expression of thermogenesis mediators (ucp1, pgc1α, dio2) in WAT and/or BAT in SAMP8 mice. Interestingly, SAMP8 mice exhibit a decreased tolerance to an acute (5 h) cold challenge. Moreover, SAMP8 and SAMR1 mice exhibited differential responses to a chronic (1 week) decrease in ambient temperature; the greatest response in whole-body substrate selection was observed in SAMR1 mice. Collectively, these observations reveal differential behaviors (e.g. 24 h food intake patterns) in SAMP8 mice that are associated with perturbations in peripheral circadian clocks, metabolism and thermogenesis.
    Keywords:  Aging; chronobiology; metabolism; thermogenesis
  54. Proc Natl Acad Sci U S A. 2020 Aug 21. pii: 202008919. [Epub ahead of print]
    Kaspari RR, Reyna-Neyra A, Jung L, Torres-Manzo AP, Hirabara SM, Carrasco N.
      Obesity is a major health problem worldwide, given its growing incidence and its association with a variety of comorbidities. Weight gain results from an increase in energy intake without a concomitant increase in energy expenditure. To combat the obesity epidemic, many studies have focused on the pathways underlying satiety and hunger signaling, while other studies have concentrated on the mechanisms involved in energy expenditure, most notably adaptive thermogenesis. Hypothyroidism in humans is typically associated with a decreased basal metabolic rate, lower energy expenditure, and weight gain. However, hypothyroid mouse models have been reported to have a leaner phenotype than euthyroid controls. To elucidate the mechanism underlying this phenomenon, we used a drug-free mouse model of hypothyroidism: mice lacking the sodium/iodide symporter (NIS), the plasma membrane protein that mediates active iodide uptake in the thyroid. In addition to being leaner than euthyroid mice, owing in part to reduced food intake, these hypothyroid mice show signs of compensatory up-regulation of the skeletal-muscle adaptive thermogenic marker sarcolipin, with an associated increase in fatty acid oxidation (FAO). Neither catecholamines nor thyroid-stimulating hormone (TSH) are responsible for sarcolipin expression or FAO stimulation; rather, thyroid hormones are likely to negatively regulate both processes in skeletal muscle. Our findings indicate that hypothyroidism in mice results in a variety of metabolic changes, which collectively lead to a leaner phenotype. A deeper understanding of these changes may make it possible to develop new strategies against obesity.
    Keywords:  adaptive thermogenesis; food intake; hypothyroidism; sarcolipin; sodium/iodide symporter (NIS)
  55. Eur J Cell Biol. 2020 Aug;pii: S0171-9335(20)30047-9. [Epub ahead of print]99(6): 151108
    Mohamad Kamal NS, Safuan S, Shamsuddin S, Foroozandeh P.
      Cellular theory of aging states that human aging is the result of cellular aging, in which an increasing proportion of cells reach senescence. Senescence, from the Latin word senex, means "growing old," is an irreversible growth arrest which occurs in response to damaging stimuli, such as DNA damage, telomere shortening, telomere dysfunction and oncogenic stress leading to suppression of potentially dysfunctional, transformed, or aged cells. Cellular senescence is characterized by irreversible cell cycle arrest, flattened and enlarged morphology, resistance to apoptosis, alteration in gene expression and chromatin structure, expression of senescence associated- β-galactosidase (SA-β-gal) and acquisition of senescence associated secretory phenotype (SASP). In this review paper, different types of cellular senescence including replicative senescence (RS) which occurs due to telomere shortening and stress induced premature senescence (SIPS) which occurs in response to different types of stress in cells, are discussed. Biomarkers of cellular senescence and senescent assays including BrdU incorporation assay, senescence associated- β-galactosidase (SA-β-gal) and senescence-associated heterochromatin foci assays to detect senescent cells are also addressed.
    Keywords:  Aging; Cellular senescence; Replicative senescence; Senescence associated secretory phenotype (SASP); Senescence associated- β-galactosidase (SA-β-gal); Stress induced premature senescence
  56. FASEB J. 2020 Aug 17.
    Marchetti P, Fovez Q, Germain N, Khamari R, Kluza J.
      Mitochondrial metabolism must constantly adapt to stress conditions in order to maintain bioenergetic levels related to cellular functions. This absence of proper adaptation can be seen in a wide array of conditions, including cancer. Metabolic adaptation calls on mitochondrial function and draws on the mitochondrial reserve to meet increasing needs. Among mitochondrial respiratory parameters, the spare respiratory capacity (SRC) represents a particularly robust functional parameter to evaluate mitochondrial reserve. We provide an overview of potential SRC mechanisms and regulation with a focus on its particular significance in cancer cells.
    Keywords:  mitochondrial adaptation; mitochondrial dysfunction; mitochondrial flexibility; mitochondrial metabolism; mitochondrial stress; oxygen consumption; respiratory reserve capacity; spare reserve capacity; uncoupling ratio
  57. Cell Rep. 2020 Aug 18. pii: S2211-1247(20)31028-7. [Epub ahead of print]32(7): 108043
    Egozi A, Bahar Halpern K, Farack L, Rotem H, Itzkovitz S.
      The islets of Langerhans are dynamic structures that can change in size, number of cells, and molecular function in response to physiological and pathological stress. Molecular cues originating from the surrounding "peri-islet" acinar cells that could facilitate this plasticity have not been explored. Here, we combine single-molecule transcript imaging in the intact pancreas and transcriptomics to identify spatial heterogeneity of acinar cell gene expression. We find that peri-islet acinar cells exhibit a distinct molecular signature in db/db diabetic mice that includes upregulation of trypsin family genes and elevated mTOR activity. This zonated expression program seems to be induced by CCK that is secreted from islet cells. Elevated peri-islet trypsin secretion could facilitate the islet expansion observed in this model via modulation of the islet capsule matrix components. Our study highlights a molecular axis of communication between the pancreatic exocrine and endocrine compartments that may be relevant to islet expansion.
    Keywords:  acinar cells; db/db mice; diabetes; pancreas; peri-islet acinar cells; single cell heterogeneity; single molecule imaging; smFISH; zonation
  58. Autophagy. 2020 Aug 19.
    Nath P, Jena KK, Mehto S, Chauhan NR, Sahu R, Dhar K, Srinivas K, Chauhan S, Chauhan S.
      IRGM is a genetic risk factor for several autoimmune diseases. However, the mechanism of IRGM-mediated protection in autoimmunity remains undetermined. The abnormal activation of type I interferon (IFN) response is one of the significant factors in the pathogenesis of several autoimmune diseases. In our recent study, we showed that IRGM is a master suppressor of the interferon response. We found that the depletion of IRGM results in constitutively activated CGAS-STING1, DDX58/RIG-I-MAVS, and TLR3-TICAM1/TRIF signaling pathways resulting in upregulation of almost all IFN-responsive genes. Mechanistically, IRGM utilizes a two-pronged mechanism to suppress the interferon response. First, it mediates SQSTM1/p62-dependent selective macroautophagy/autophagy of nucleic acid sensor proteins, including CGAS, DDX58/RIG-I, and TLR3. Second, it facilitates the removal of defective mitochondria by mitophagy and avoids a buildup of mito-ROS and mito-damage/danger-associated molecular patterns (DAMPs). Thus, IRGM deficiency results in increased nucleic acid sensors and DAMPs engaging a vicious cycle of aberrant activation of IFN response that is known to occur in systemic autoimmune-like conditions.
    Keywords:  Autoimmunity; DAMPs; IRGM; IRGM1; RIG-I-MAVS; autophagy; cGAS-STING; mitophagy
  59. Science. 2020 Aug 20. pii: eaba6527. [Epub ahead of print]
    Lajoie MJ, Boyken SE, Salter AI, Bruffey J, Rajan A, Langan RA, Olshefsky A, Muhunthan V, Bick MJ, Gewe M, Quijano-Rubio A, Johnson J, Lenz G, Nguyen A, Pun S, Correnti CE, Riddell SR, Baker D.
      Precise cell targeting is challenging because most mammalian cell types lack a single surface marker that distinguishes them from other cells. A solution would be to target cells based on specific combinations of proteins present on their surfaces. We design colocalization-dependent protein switches (Co-LOCKR) that perform AND, OR, and NOT Boolean logic operations. These switches activate through a conformational change only when all conditions are met, generating rapid, transcription-independent responses at single-cell resolution within complex cell populations. We implement AND gates to redirect T cell specificity against tumor cells expressing two surface antigens while avoiding off-target recognition of single-antigen cells, and 3-input switches that add NOT or OR logic to avoid or include cells expressing a third antigen. Thus, de novo designed proteins can perform computations on the surface of cells, integrating multiple distinct binding interactions into a single output.
  60. Trends Cancer. 2020 Aug 13. pii: S2405-8033(20)30207-7. [Epub ahead of print]
  61. Trends Mol Med. 2020 Aug 13. pii: S1471-4914(20)30187-8. [Epub ahead of print]
    Mascharak S, desJardins-Park HE, Longaker MT.
      Recent work has revealed that fibroblasts are remarkably heterogeneous cells, but the appropriate lens through which to study this variation (lineage, phenotype, and plasticity) and its relevance to human biology remain unclear. In this opinion article, we comment on recent breakthroughs in our understanding of fibroblast heterogeneity during skin wound healing, and on open questions that must be addressed to clinically translate these findings in order to minimize scarring in patients. We emphasize the need for experimental models of wound healing that better approximate human biology, as well as comparison of scarring and regenerative phenotypes to uncover master regulators of fibrosis.
    Keywords:  fibroblast heterogeneity; lineage; plasticity; regeneration; wound healing
  62. Proc Natl Acad Sci U S A. 2020 Aug 19. pii: 202011225. [Epub ahead of print]
    Liu Z, Selby CP, Yang Y, Lindsey-Boltz LA, Cao X, Eynullazada K, Sancar A.
      The circadian clock is a global regulatory mechanism that controls the expression of 50 to 80% of transcripts in mammals. Some of the genes controlled by the circadian clock are oncogenes or tumor suppressors. Among these Myc has been the focus of several studies which have investigated the effect of clock genes and proteins on Myc transcription and MYC protein stability. Other studies have focused on effects of Myc mutation or overproduction on the circadian clock in comparison to their effects on cell cycle progression and tumorigenesis. Here we have used mice with mutations in the essential clock genes Bmal1, Cry1, and Cry2 to gain further insight into the effect of the circadian clock on this important oncogene/oncoprotein and tumorigenesis. We find that mutation of both Cry1 and Cry2, which abolishes the negative arm of the clock transcription-translation feedback loop (TTFL), causes down-regulation of c-MYC, and mutation of Bmal1, which abolishes the positive arm of TTFL, causes up-regulation of the c-MYC protein level in mouse spleen. These findings must be taken into account in models of the clock disruption-cancer connection.
    Keywords:  BMAL1; c-MYC; circadian clock; cryptochromes; transcription regulation
  63. Lancet. 2020 08 15. pii: S0140-6736(20)31318-0. [Epub ahead of print]396(10249): 499-512
    Beyer G, Habtezion A, Werner J, Lerch MM, Mayerle J.
      Chronic pancreatitis is a multifactorial, fibroinflammatory syndrome in which repetitive episodes of pancreatic inflammation lead to extensive fibrotic tissue replacement, resulting in chronic pain, exocrine and endocrine pancreatic insufficiency, reduced quality of life, and a shorter life expectancy. The incidence and prevalence of chronic pancreatitis is rising and no curative treatment is available. Using novel diagnostic algorithms, definitive chronic pancreatitis can be diagnosed by imaging criteria alone, whereas probable chronic pancreatitis requires clinical features and imaging criteria. Criteria for the diagnosis of early chronic pancreatitis are still under discussion and need prospective validation in clinical trials. Cross-sectional imaging should be used first; endoscopic ultrasound is needed only when CT or MRI are inconclusive or to plan therapeutic interventions. Management of chronic pancreatitis requires an interdisciplinary approach including primary care practitioners, gastroenterologists, surgeons, radiologists, pain specialists, and nutritional therapists. Patients with chronic pancreatitis should be seen at least once a year and re-evaluated for causal risk factors, symptom control, and complications such as malnutrition, pancreatic exocrine insufficiency, and diabetes; refer to a specialised centre if symptoms are poorly controlled or there is risk of deterioration. Scoring systems to monitor disease progression have been developed and validated internationally. Interventional treatments for pain or cholestasis should be done by specialists only, and early discussion of treatment approaches should include all medical disciplines involved in care. Throughout this Seminar, we address research needs such as staging of pancreatitis, aspects of malnutrition and pain, and cancer surveillance, to help improve the care of patients.
  64. Clin Exp Metastasis. 2020 Aug 20.
    De A, Beligala DH, Sharma VP, Burgos CA, Lee AM, Geusz ME.
      Epithelial-mesenchymal transition (EMT) is a key event preceding tumor cell metastasis that increases cell invasiveness and cancer stem cell (CSC) populations. Studies suggest that genes used in generating circadian rhythms also serve in regulating EMT. To test the role of circadian clocks in cellular EMT events two cancer cell lines were compared, one that has a well-established circadian clock, C6 from rat glioma, and one that does not, MCF-7 from human breast tumor. MCF-7 tumorsphere cultures were tested for evidence of circadian rhythms because of previously reported circadian rhythm enhancement in C6 tumorspheres shown by elevated rhythm amplitude and increased expression of circadian clock gene Per2. Bioluminescence imaging of Per2 gene expression in MCF-7 tumorspheres revealed a previously unconfirmed circadian clock in this important cancer research model. Inducing CSC generation through EMT in C6 and MCF-7 monolayer cultures revealed circadian oscillations in the size of the post-EMT CSC population, confirming that circadian rhythms are additional processes controlling this stage of cancer progression. EMT was verified by distinct cellular morphological changes and expression of stem cell proteins OCT4, nestin, MSI1, and CD133 along with EMT-related proteins ZEB1, vimentin, and TWIST. Quantifying single-cell events and behaviors through time-lapse imaging indicated the post-EMT population size was determined largely by circadian rhythms in epithelial-like cancer cells undergoing EMT. We then identified a specific phase of the circadian rhythm in Per2 gene activation as a potential target for therapeutic treatments that may suppress EMT, minimize CSCs, and limit metastasis.
    Keywords:  Cancer stem cell; Circadian rhythm; Epithelial-mesenchymal transition; OCT4; Tumorsphere
  65. Proc Natl Acad Sci U S A. 2020 Aug 12. pii: 202006375. [Epub ahead of print]
    Sun ED, Michaels TCT, Mahadevan L.
      Many complex systems experience damage accumulation, which leads to aging, manifest as an increasing probability of system collapse with time. This naturally raises the question of how to maximize health and longevity in an aging system at minimal cost of maintenance and intervention. Here, we pose this question in the context of a simple interdependent network model of aging in complex systems and show that it exhibits cascading failures. We then use both optimal control theory and reinforcement learning alongside a combination of analysis and simulation to determine optimal maintenance protocols. These protocols may motivate the rational design of strategies for promoting longevity in aging complex systems with potential applications in therapeutic schedules and engineered system maintenance.
    Keywords:  aging; control; failure; networks; repair
  66. Cell Cycle. 2020 Aug 20. 1-13
    Foster DJ, Chang HM, Haswell JR, Gregory RI, Slack FJ.
      TRIM71 is an important RNA-binding protein in development and disease, yet its direct targets have not been investigated globally. Here we describe a number of disease and developmentally-relevant TRIM71 RNA targets such as the MBNL family, LIN28B, MDM2, and TCF7L2. We describe a new role for TRIM71 as capable of positive or negative RNA regulation depending on the RNA target. We found that TRIM71 co-precipitated with IMP1 which could explain its multiple mechanisms of RNA regulation, as IMP1 is typically thought to stabilize RNAs. Deletion of the NHL domain of TRIM71 impacted its ability to bind to RNA and RNAs bound by congenital hydrocephalus-associated point mutations in the RNA-binding NHL domain of TRIM71 clustered closely with RNAs bound by the NHL deletion mutant. Our work expands the possible mechanisms by which TRIM71 may regulate RNAs and elucidates further potential RNA targets.
    Keywords:  IGF2BP1; IMP1; RIP-seq; RNABP; TRIM71; crosslinking; liver cancer
  67. Elife. 2020 08 19. pii: e58362. [Epub ahead of print]9
    Aibara S, Singh V, Modelska A, Amunts A.
      Translation of mitochondrial messenger RNA (mt-mRNA) is performed by distinct mitoribosomes comprising at least 36 mitochondria-specific proteins. How these mitoribosomal proteins assist in the binding of mt-mRNA and to what extent they are involved in the translocation of transfer RNA (mt-tRNA) is unclear. To visualize the process of translation in human mitochondria, we report ~3.0 Å resolution structure of the human mitoribosome, including the L7/L12 stalk, and eight structures of its functional complexes with mt-mRNA, mt-tRNAs, recycling factor and additional trans factors. The study reveals a transacting protein module LRPPRC-SLIRP that delivers mt-mRNA to the mitoribosomal small subunit through a dedicated platform formed by the mitochondria-specific protein mS39. Mitoribosomal proteins of the large subunit mL40, mL48, and mL64 coordinate translocation of mt-tRNA. The comparison between those structures shows dynamic interactions between the mitoribosome and its ligands, suggesting a sequential mechanism of conformational changes.
    Keywords:  RNA; cryo-EM; gene expression; human; mitochondria; molecular biophysics; ribosome; structural biology; translation
  68. J Pain. 2020 Aug 13. pii: S1526-5900(20)30061-4. [Epub ahead of print]
    Lahav Y, Levy D, Ohry A, Zeilig G, Lahav M, Golander H, Guber AC, Uziel O, Defrin R.
      Chronic pain induces a multitude of harmful effects; recently it has been suggested that chronic pain is also associated with premature aging, manifested in shortened telomere length (TL). However, evidence for this hypothesis is scarce and inconsistent. The aim was twofold: 1) Investigate whether chronic pain is associated with premature aging, and 2) Determine whether physical exercise (PE) moderates this association if it exists. Participants were 116 male subjects, with (n=67) and without chronic pain (n=49). Blood samples for TL analysis were collected and participants were interviewed and completed questionnaires. As a part of the cohort, we included people with physical disability; this variable was controlled in the analysis. The TL of individuals with chronic pain was significantly shorter than that of pain-free individuals. Regression analysis revealed a significant moderating effect of PE on chronic pain and TL, above and beyond the effects of disability, age, and weight. Whereas chronic pain was associated with shorter telomeres in participants who did not exercise, this association was non-significant among participants who did exercise. The results suggest that chronic pain is associated with premature ageing; however, PE may mitigate this association and may protect individuals against the harmful effects of chronic pain. PERSPECTIVE: The study suggest that it is important to monitor signs of premature ageing among chronic pain patients as they are at risk. However, chronic pain patients may benefit from regular physical exercise in this respect as it may moderate premature ageing.
    Keywords:  Chronic pain; Premature ageing, Telomere length, Physical exercise
  69. F1000Res. 2020 ;pii: F1000 Faculty Rev-935. [Epub ahead of print]9
    Luo X, O'Neill KL, Huang K.
      Bax and Bak, two functionally similar, pro-apoptotic proteins of the Bcl-2 family, are known as the gateway to apoptosis because of their requisite roles as effectors of mitochondrial outer membrane permeabilization (MOMP), a major step during mitochondria-dependent apoptosis. The mechanism of how cells turn Bax/Bak from inert molecules into fully active and lethal effectors had long been the focal point of a major debate centered around two competing, but not mutually exclusive, models: direct activation and indirect activation. After intensive research efforts for over two decades, it is now widely accepted that to initiate apoptosis, some of the BH3-only proteins, a subclass of the Bcl-2 family, directly engage Bax/Bak to trigger their conformational transformation and activation. However, a series of recent discoveries, using previously unavailable CRISPR-engineered cell systems, challenge the basic premise that undergirds the consensus and provide evidence for a novel and surprisingly simple model of Bax/Bak activation: the membrane (lipids)-mediated spontaneous model. This review will discuss the evidence, rationale, significance, and implications of this new model.
    Keywords:  Activator; Apoptosis; BH3 mimetics; BH3-only proteins; Bad; Bak; Bax; Bcl-2 family; Bcl-xL; Bid; Bim; Direct activation; Indirect activation; Mcl-1; Membrane-mediated Spontaneous activation; Mitochondrial outer membrane; Sensitizer; auto-activation; de novo activation; retro-translocation
  70. Nature. 2020 Aug 19.
    Yoshihara E, O'Connor C, Gasser E, Wei Z, Oh TG, Tseng TW, Wang D, Cayabyab F, Dai Y, Yu RT, Liddle C, Atkins AR, Downes M, Evans RM.
      Islets derived from stem cells hold promise as a therapy for insulin-dependent diabetes, but there remain challenges towards achieving this goal1-6. Here we generate human islet-like organoids (HILOs) from induced pluripotent stem cells and show that non-canonical WNT4 signalling drives the metabolic maturation necessary for robust ex vivo glucose-stimulated insulin secretion. These functionally mature HILOs contain endocrine-like cell types that, upon transplantation, rapidly re-establish glucose homeostasis in diabetic NOD/SCID mice. Overexpression of the immune checkpoint protein programmed death-ligand 1 (PD-L1) protected HILO xenografts such that they were able to restore glucose homeostasis in immune-competent diabetic mice for 50 days. Furthermore, ex vivo stimulation with interferon-γ induced endogenous PD-L1 expression and restricted T cell activation and graft rejection. The generation of glucose-responsive islet-like organoids that are able to avoid immune detection provides a promising alternative to cadaveric and device-dependent therapies in the treatment of diabetes.
  71. N Engl J Med. 2020 08 20. 383(8): 721-732
    Yoshino M, Kayser BD, Yoshino J, Stein RI, Reeds D, Eagon JC, Eckhouse SR, Watrous JD, Jain M, Knight R, Schechtman K, Patterson BW, Klein S.
      BACKGROUND: Some studies have suggested that in people with type 2 diabetes, Roux-en-Y gastric bypass has therapeutic effects on metabolic function that are independent of weight loss.METHODS: We evaluated metabolic regulators of glucose homeostasis before and after matched (approximately 18%) weight loss induced by gastric bypass (surgery group) or diet alone (diet group) in 22 patients with obesity and diabetes. The primary outcome was the change in hepatic insulin sensitivity, assessed by infusion of insulin at low rates (stages 1 and 2 of a 3-stage hyperinsulinemic euglycemic pancreatic clamp). Secondary outcomes were changes in muscle insulin sensitivity, beta-cell function, and 24-hour plasma glucose and insulin profiles.
    RESULTS: Weight loss was associated with increases in mean suppression of glucose production from baseline, by 7.04 μmol per kilogram of fat-free mass per minute (95% confidence interval [CI], 4.74 to 9.33) in the diet group and by 7.02 μmol per kilogram of fat-free mass per minute (95% CI, 3.21 to 10.84) in the surgery group during clamp stage 1, and by 5.39 (95% CI, 2.44 to 8.34) and 5.37 (95% CI, 2.41 to 8.33) μmol per kilogram of fat-free mass per minute in the two groups, respectively, during clamp stage 2; there were no significant differences between the groups. Weight loss was associated with increased insulin-stimulated glucose disposal, from 30.5±15.9 to 61.6±13.0 μmol per kilogram of fat-free mass per minute in the diet group and from 29.4±12.6 to 54.5±10.4 μmol per kilogram of fat-free mass per minute in the surgery group; there was no significant difference between the groups. Weight loss increased beta-cell function (insulin secretion relative to insulin sensitivity) by 1.83 units (95% CI, 1.22 to 2.44) in the diet group and by 1.11 units (95% CI, 0.08 to 2.15) in the surgery group, with no significant difference between the groups, and it decreased the areas under the curve for 24-hour plasma glucose and insulin levels in both groups, with no significant difference between the groups. No major complications occurred in either group.
    CONCLUSIONS: In this study involving patients with obesity and type 2 diabetes, the metabolic benefits of gastric bypass surgery and diet were similar and were apparently related to weight loss itself, with no evident clinically important effects independent of weight loss. (Funded by the National Institutes of Health and others; number, NCT02207777.).
  72. Cancers (Basel). 2020 Aug 12. pii: E2260. [Epub ahead of print]12(8):
    Negrini S, De Palma R, Filaci G.
      Telomerase is a reverse transcriptase that maintains telomeres length, compensating for the attrition of chromosomal ends that occurs during each replication cycle. Telomerase is expressed in germ cells and stem cells, whereas it is virtually undetectable in adult somatic cells. On the other hand, telomerase is broadly expressed in the majority of human tumors playing a crucial role in the replicative behavior and immortality of cancer cells. Several studies have demonstrated that telomerase-derived peptides are able to bind to HLA (human leukocyte antigen) class I and class II molecules and effectively activate both CD8+ and CD4+ T cells subsets. Due to its broad and selective expression in cancer cells and its significant immunogenicity, telomerase is considered an ideal universal tumor-associated antigen, and consequently, a very attractive target for anti-cancer immunotherapy. To date, different telomerase targeting immunotherapies have been studied in pre-clinical and clinical settings, these approaches include peptide vaccination and cell-based vaccination. The objective of this review paper is to discuss the role of human telomerase in cancer immunotherapy analyzing recent developments and future perspectives in this field.
    Keywords:  cancer; clinical trials; hTERT; immunotherapy; telomerase; vaccine
  73. Nat Commun. 2020 Aug 18. 11(1): 4135
    Gutiérrez-Fernández J, Kaszuba K, Minhas GS, Baradaran R, Tambalo M, Gallagher DT, Sazanov LA.
      Complex I is the first and the largest enzyme of respiratory chains in bacteria and mitochondria. The mechanism which couples spatially separated transfer of electrons to proton translocation in complex I is not known. Here we report five crystal structures of T. thermophilus enzyme in complex with NADH or quinone-like compounds. We also determined cryo-EM structures of major and minor native states of the complex, differing in the position of the peripheral arm. Crystal structures show that binding of quinone-like compounds (but not of NADH) leads to a related global conformational change, accompanied by local re-arrangements propagating from the quinone site to the nearest proton channel. Normal mode and molecular dynamics analyses indicate that these are likely to represent the first steps in the proton translocation mechanism. Our results suggest that quinone binding and chemistry play a key role in the coupling mechanism of complex I.
  74. Metabol Open. 2019 Dec;4 100020
    Grey JFE, Townley AR, Everitt NM, Campbell-Ritchie A, Wheatley SP.
      Analysis of cellular energetics is central to understanding metabolic diseases including diabetes and cancer. The two most commonly used methods to monitor cellular respiration are the Seahorse-XF system, and Glo™ assays, which are considered "gold standards". These commercial methods measure energetics indirectly and require considerable financial investment. Here we describe an alternative assay that enables accurate quantification of NADH turnover and that is affordable. This method measures resazurin reduction to resorufin at rising concentrations in the presence of purified mitochondrial extracts until NADH becomes a rate-limiting factor. This indicates the maximal level of NADH turnover in each sample and therefore infers metabolic activity. Here we compare MRC5, MCF7 and MDA231 cell lines which have differing metabolic profiles.
    Keywords:  Mitochondria; NADH; Oxidative phosphorylation
  75. BMJ Support Palliat Care. 2020 Aug 21. pii: bmjspcare-2020-002296. [Epub ahead of print]
    Cong M, Song C, Xu H, Song C, Wang C, Fu Z, Ba Y, Wu J, Xie C, Chen G, Chen Z, Zhou L, Li T, Deng L, Xin L, Yang L, Cui J, Shi H, .
      BACKGROUND: Cancer cachexia is a complex metabolic syndrome characterised by a loss of muscle with or without loss of fat mass, and is associated with high morbidity and mortality. Despite its clinical importance, there is a lack of simple tools to screen patients for cancer cachexia. The aim of this study was to evaluate and validate the patient-generated subjective global assessment (PG-SGA) as a screening tool for cancer cachexia.METHODS: This is a secondary analysis of a multicentre, cross-sectional, observational study. Cancer cachexia was diagnosed when there was weight loss ≥5% during the past 12 months and at least three of the five following conditions were present: decreased muscle strength, fatigue, anorexia, low Fat-Free Mass Index (FFMI) and abnormal laboratory findings. A quadratic discriminant analysis was conducted for the ability of PG-SGA to predict cachexia.
    RESULTS: A total of 4231 patients with cancer were included in this analysis, and 351 patients (8.3%) were diagnosed as having cachexia. The highest incidence of cachexia was found among patients with pancreatic cancer (32.5%), oesophageal cancer (21.5%) and gastric cancer (17.9%). Compared with patients without cachexia, patients with cachexia had a lower body mass index, FFMI, hand grip strength, total protein, prealbumin, albumin, haemoglobin and Karnofsky performance status (p<0.05), while they had a higher C reactive protein level and PG-SGA Score (4.71±3.71 vs 10.87±4.84, p<0.05). The best cut-off value for PG-SGA was 6.5, with 79.8% of sensitivity and 72.3% specificity for cachexia, and the area under the receiver operating characteristic curve was 0.846 (95% CI 0.826 to 0.866, p<0.001).
    CONCLUSIONS: PG-SGA is a highly specific tool that can be used to screen patients for cancer cachexia.