bims-cagime Biomed News
on Cancer, aging and metabolism
Issue of 2020‒06‒07
fifty papers selected by
Kıvanç Görgülü
Technical University of Munich


  1. Nat Chem Biol. 2020 Jun 01.
    Pinch BJ, Doctor ZM, Nabet B, Browne CM, Seo HS, Mohardt ML, Kozono S, Lian X, Manz TD, Chun Y, Kibe S, Zaidman D, Daitchman D, Yeoh ZC, Vangos NE, Geffken EA, Tan L, Ficarro SB, London N, Marto JA, Buratowski S, Dhe-Paganon S, Zhou XZ, Lu KP, Gray NS.
      Peptidyl-prolyl cis/trans isomerase NIMA-interacting 1 (Pin1) is commonly overexpressed in human cancers, including pancreatic ductal adenocarcinoma (PDAC). While Pin1 is dispensable for viability in mice, it is required for activated Ras to induce tumorigenesis, suggesting a role for Pin1 inhibitors in Ras-driven tumors, such as PDAC. We report the development of rationally designed peptide inhibitors that covalently target Cys113, a highly conserved cysteine located in the Pin1 active site. The inhibitors were iteratively optimized for potency, selectivity and cell permeability to give BJP-06-005-3, a versatile tool compound with which to probe Pin1 biology and interrogate its role in cancer. In parallel to inhibitor development, we employed genetic and chemical-genetic strategies to assess the consequences of Pin1 loss in human PDAC cell lines. We demonstrate that Pin1 cooperates with mutant KRAS to promote transformation in PDAC, and that Pin1 inhibition impairs cell viability over time in PDAC cell lines.
    DOI:  https://doi.org/10.1038/s41589-020-0550-9
  2. Cancer Discov. 2020 Jun 04. pii: CD-20-0329. [Epub ahead of print]
    Alicea GM, Rebecca VW, Goldman AR, Fane ME, Douglass SM, Behera R, Webster MR, Kugel CH, Ecker BL, Caino MC, Kossenkov AV, Tang HY, Frederick DT, Flaherty KT, Xu X, Liu Q, Gabrilovich DI, Herlyn M, Blair IA, Schug ZT, Speicher DW, Weeraratna AT.
      Older melanoma patients (>50 years old) have poorer prognoses and response rates to targeted therapy compared to young patients (<50 years old), which can be driven, in part, by the aged microenvironment. Here, we show that aged dermal fibroblasts increase the secretion of neutral lipids, especially ceramides. When melanoma cells are exposed to the aged fibroblast lipid secretome, or co-cultured with aged fibroblasts, they increase the uptake of lipids, via the fatty acid transporter, fatty acid transport protein (FATP) 2, which is upregulated in melanoma cells in the aged microenvironment and known to play roles in lipid synthesis and accumulation. We show that blocking FATP2 in melanoma cells in an aged microenvironment inhibits their accumulation of lipids, and disrupts their mitochondrial metabolism. Inhibiting FATP2 overcomes age-related resistance to BRAF/MEK inhibition in animal models, ablates tumor relapse, and significantly extends survival time in older animals.
    DOI:  https://doi.org/10.1158/2159-8290.CD-20-0329
  3. J Exp Med. 2020 Aug 03. pii: e20191920. [Epub ahead of print]217(8):
    Manzo T, Prentice BM, Anderson KG, Raman A, Schalck A, Codreanu GS, Nava Lauson CB, Tiberti S, Raimondi A, Jones MA, Reyzer M, Bates BM, Spraggins JM, Patterson NH, McLean JA, Rai K, Tacchetti C, Tucci S, Wargo JA, Rodighiero S, Clise-Dwyer K, Sherrod SD, Kim M, Navin NE, Caprioli RM, Greenberg PD, Draetta G, Nezi L.
      CD8+ T cells are master effectors of antitumor immunity, and their presence at tumor sites correlates with favorable outcomes. However, metabolic constraints imposed by the tumor microenvironment (TME) can dampen their ability to control tumor progression. We describe lipid accumulation in the TME areas of pancreatic ductal adenocarcinoma (PDA) populated by CD8+ T cells infiltrating both murine and human tumors. In this lipid-rich but otherwise nutrient-poor TME, access to using lipid metabolism becomes particularly valuable for sustaining cell functions. Here, we found that intrapancreatic CD8+ T cells progressively accumulate specific long-chain fatty acids (LCFAs), which, rather than provide a fuel source, impair their mitochondrial function and trigger major transcriptional reprogramming of pathways involved in lipid metabolism, with the subsequent reduction of fatty acid catabolism. In particular, intrapancreatic CD8+ T cells specifically exhibit down-regulation of the very-long-chain acyl-CoA dehydrogenase (VLCAD) enzyme, which exacerbates accumulation of LCFAs and very-long-chain fatty acids (VLCFAs) that mediate lipotoxicity. Metabolic reprogramming of tumor-specific T cells through enforced expression of ACADVL enabled enhanced intratumoral T cell survival and persistence in an engineered mouse model of PDA, overcoming one of the major hurdles to immunotherapy for PDA.
    DOI:  https://doi.org/10.1084/jem.20191920
  4. Cell Metab. 2020 May 26. pii: S1550-4131(20)30241-2. [Epub ahead of print]
    Grosse L, Wagner N, Emelyanov A, Molina C, Lacas-Gervais S, Wagner KD, Bulavin DV.
      The accumulation of senescent cells can drive many age-associated phenotypes and pathologies. Consequently, it has been proposed that removing senescent cells might extend lifespan. Here, we generated two knockin mouse models targeting the best-characterized marker of senescence, p16Ink4a. Using a genetic lineage tracing approach, we found that age-induced p16High senescence is a slow process that manifests around 10-12 months of age. The majority of p16High cells were vascular endothelial cells mostly in liver sinusoids (LSECs), and to lesser extent macrophages and adipocytes. In turn, continuous or acute elimination of p16High senescent cells disrupted blood-tissue barriers with subsequent liver and perivascular tissue fibrosis and health deterioration. Our data show that senescent LSECs are not replaced after removal and have important structural and functional roles in the aging organism. In turn, delaying senescence or replacement of senescent LSECs could represent a powerful tool in slowing down aging.
    Keywords:  aging; fibrosis; lineage tracing; liver sinusoid endothelial cells; liver sinusoids; p16; senescence; vascular endothelial cells
    DOI:  https://doi.org/10.1016/j.cmet.2020.05.002
  5. Nat Commun. 2020 Jun 04. 11(1): 2817
    Shankar S, Tien JC, Siebenaler RF, Chugh S, Dommeti VL, Zelenka-Wang S, Wang XM, Apel IJ, Waninger J, Eyunni S, Xu A, Mody M, Goodrum A, Zhang Y, Tesmer JJ, Mannan R, Cao X, Vats P, Pitchiaya S, Ellison SJ, Shi J, Kumar-Sinha C, Crawford HC, Chinnaiyan AM.
      Both KRAS and EGFR are essential mediators of pancreatic cancer development and interact with Argonaute 2 (AGO2) to perturb its function. Here, in a mouse model of mutant KRAS-driven pancreatic cancer, loss of AGO2 allows precursor lesion (PanIN) formation yet prevents progression to pancreatic ductal adenocarcinoma (PDAC). Precursor lesions with AGO2 ablation undergo oncogene-induced senescence with altered microRNA expression and EGFR/RAS signaling, bypassed by loss of p53. In mouse and human pancreatic tissues, PDAC progression is associated with increased plasma membrane localization of RAS/AGO2. Furthermore, phosphorylation of AGO2Y393 disrupts both the wild-type and oncogenic KRAS-AGO2 interaction, albeit under different conditions. ARS-1620 (G12C-specific inhibitor) disrupts the KRASG12C-AGO2 interaction, suggesting that the interaction is targetable. Altogether, our study supports a biphasic model of pancreatic cancer development: an AGO2-independent early phase of PanIN formation reliant on EGFR-RAS signaling, and an AGO2-dependent phase wherein the mutant KRAS-AGO2 interaction is critical for PDAC progression.
    DOI:  https://doi.org/10.1038/s41467-020-16309-2
  6. Nat Genet. 2020 Jun;52(6): 582-593
    Jiménez-Sánchez A, Cybulska P, Mager KL, Koplev S, Cast O, Couturier DL, Memon D, Selenica P, Nikolovski I, Mazaheri Y, Bykov Y, Geyer FC, Macintyre G, Gavarró LM, Drews RM, Gill MB, Papanastasiou AD, Sosa RE, Soslow RA, Walther T, Shen R, Chi DS, Park KJ, Hollmann T, Reis-Filho JS, Markowetz F, Beltrao P, Vargas HA, Zamarin D, Brenton JD, Snyder A, Weigelt B, Sala E, Miller ML.
      In metastatic cancer, the degree of heterogeneity of the tumor microenvironment (TME) and its molecular underpinnings remain largely unstudied. To characterize the tumor-immune interface at baseline and during neoadjuvant chemotherapy (NACT) in high-grade serous ovarian cancer (HGSOC), we performed immunogenomic analysis of treatment-naive and paired samples from before and after treatment with chemotherapy. In treatment-naive HGSOC, we found that immune-cell-excluded and inflammatory microenvironments coexist within the same individuals and within the same tumor sites, indicating ubiquitous variability in immune cell infiltration. Analysis of TME cell composition, DNA copy number, mutations and gene expression showed that immune cell exclusion was associated with amplification of Myc target genes and increased expression of canonical Wnt signaling in treatment-naive HGSOC. Following NACT, increased natural killer (NK) cell infiltration and oligoclonal expansion of T cells were detected. We demonstrate that the tumor-immune microenvironment of advanced HGSOC is intrinsically heterogeneous and that chemotherapy induces local immune activation, suggesting that chemotherapy can potentiate the immunogenicity of immune-excluded HGSOC tumors.
    DOI:  https://doi.org/10.1038/s41588-020-0630-5
  7. Mol Cancer Ther. 2020 Jun 04. pii: molcanther.1166.2020. [Epub ahead of print]
    Dimitrov-Markov S, Perales-Patón J, Bockorny B, Dopazo A, Muñoz M, Baños N, Bonilla V, Menendez C, Duran Y, Huang L, Perea S, Muthuswamy SK, Al-Shahrour F, Lopez-Casas PP, Hidalgo M.
      Metastasis development is the leading cause of cancer-related mortality in pancreatic ductal adenocarcinoma (PDAC) and yet, few preclinical systems to recapitulate its full spreading process are available. Thus, modeling of tumor progression to metastasis is urgently needed. In this work, we describe the generation of highly metastatic PDAC patient-derived xenograft (PDX) mouse models and subsequent single-cell RNA sequencing of circulating tumor cells (CTC), isolated by human HLA sorting, to identify altered signaling and metabolic pathways as well as potential therapeutic targets. The mouse models developed liver and lung metastasis with a high reproducibility rate. Isolated CTCs were highly tumorigenic, had metastatic potential and single-cell RNA sequencing showed that their expression profiles clustered separately from those of their matched primary and metastatic tumors and were characterized by low expression of cell cycle and extracellular matrix associated genes. CTC transcriptomics identified survivin (BIRC5), a key regulator of mitosis and apoptosis, as one of the highest upregulated genes during metastatic spread. Pharmacological inhibition of survivin with YM155 or survivin-knockdown promoted cell death in organoid models as well as anoikis, suggesting that survivin facilitates cancer cell survival in circulation. Treatment of metastatic PDX models with YM155 alone and in combination with chemotherapy hindered the metastatic development resulting in improved survival. Metastatic PDX mouse model development allowed the identification of survivin as a promising therapeutic target to prevent the metastatic dissemination in PDAC.
    DOI:  https://doi.org/10.1158/1535-7163.MCT-19-1166
  8. Nat Rev Endocrinol. 2020 Jun 03.
    Kazak L, Cohen P.
      Perturbations in metabolic processes are associated with diseases such as obesity, type 2 diabetes mellitus, certain infections and some cancers. A resurgence of interest in creatine biology is developing, with new insights into a diverse set of regulatory functions for creatine. This resurgence is primarily driven by technological advances in genetic engineering and metabolism as well as by the realization that this metabolite has key roles in cells beyond the muscle and brain. Herein, we highlight the latest advances in creatine biology in tissues and cell types that have historically received little attention in the field. In adipose tissue, creatine controls thermogenic respiration and loss of this metabolite impairs whole-body energy expenditure, leading to obesity. We also cover the various roles that creatine metabolism has in cancer cell survival and the function of the immune system. Renewed interest in this area has begun to showcase the therapeutic potential that lies in understanding how changes in creatine metabolism lead to metabolic disease.
    DOI:  https://doi.org/10.1038/s41574-020-0365-5
  9. Proc Natl Acad Sci U S A. 2020 Jun 01. pii: 201921815. [Epub ahead of print]
    Zhang L, Cao J, Dong L, Lin H.
      Precisely controlling the activation of transcription factors is crucial for physiology. After a transcription factor is activated and carries out its transcriptional activity, it also needs to be properly deactivated. Here, we report a deactivation mechanism of HIF-1 and several other oncogenic transcription factors. HIF-1 promotes the transcription of an ADP ribosyltransferase, TiPARP, which serves to deactivate HIF-1. Mechanistically, TiPARP forms distinct nuclear condensates or nuclear bodies in an ADP ribosylation-dependent manner. The TiPARP nuclear bodies recruit both HIF-1α and an E3 ubiquitin ligase HUWE1, which promotes the ubiquitination and degradation of HIF-1α. Similarly, TiPARP promotes the degradation of c-Myc and estrogen receptor. By suppressing HIF-1α and other oncogenic transcription factors, TiPARP exerts strong antitumor effects both in cell culture and in mouse xenograft models. Our work reveals TiPARP as a negative-feedback regulator for multiple oncogenic transcription factors, provides insights into the functions of protein ADP-ribosylation, and suggests activating TiPARP as an anticancer strategy.
    Keywords:  ADP-ribosylation; HIF-1; TiPARP; nuclear condensates; ubiquitination
    DOI:  https://doi.org/10.1073/pnas.1921815117
  10. Nat Methods. 2020 Jun 01.
    Leman JK, Weitzner BD, Lewis SM, Adolf-Bryfogle J, Alam N, Alford RF, Aprahamian M, Baker D, Barlow KA, Barth P, Basanta B, Bender BJ, Blacklock K, Bonet J, Boyken SE, Bradley P, Bystroff C, Conway P, Cooper S, Correia BE, Coventry B, Das R, De Jong RM, DiMaio F, Dsilva L, Dunbrack R, Ford AS, Frenz B, Fu DY, Geniesse C, Goldschmidt L, Gowthaman R, Gray JJ, Gront D, Guffy S, Horowitz S, Huang PS, Huber T, Jacobs TM, Jeliazkov JR, Johnson DK, Kappel K, Karanicolas J, Khakzad H, Khar KR, Khare SD, Khatib F, Khramushin A, King IC, Kleffner R, Koepnick B, Kortemme T, Kuenze G, Kuhlman B, Kuroda D, Labonte JW, Lai JK, Lapidoth G, Leaver-Fay A, Lindert S, Linsky T, London N, Lubin JH, Lyskov S, Maguire J, Malmström L, Marcos E, Marcu O, Marze NA, Meiler J, Moretti R, Mulligan VK, Nerli S, Norn C, Ó'Conchúir S, Ollikainen N, Ovchinnikov S, Pacella MS, Pan X, Park H, Pavlovicz RE, Pethe M, Pierce BG, Pilla KB, Raveh B, Renfrew PD, Burman SSR, Rubenstein A, Sauer MF, Scheck A, Schief W, Schueler-Furman O, Sedan Y, Sevy AM, Sgourakis NG, Shi L, Siegel JB, Silva DA, Smith S, Song Y, Stein A, Szegedy M, Teets FD, Thyme SB, Wang RY, Watkins A, Zimmerman L, Bonneau R.
      The Rosetta software for macromolecular modeling, docking and design is extensively used in laboratories worldwide. During two decades of development by a community of laboratories at more than 60 institutions, Rosetta has been continuously refactored and extended. Its advantages are its performance and interoperability between broad modeling capabilities. Here we review tools developed in the last 5 years, including over 80 methods. We discuss improvements to the score function, user interfaces and usability. Rosetta is available at http://www.rosettacommons.org.
    DOI:  https://doi.org/10.1038/s41592-020-0848-2
  11. Cell Metab. 2020 Jun 02. pii: S1550-4131(20)30247-3. [Epub ahead of print]31(6): 1154-1172.e9
    Qureshi R, Picon-Ruiz M, Aurrekoetxea-Rodriguez I, Nunes de Paiva V, D'Amico M, Yoon H, Radhakrishnan R, Morata-Tarifa C, Ince T, Lippman ME, Thaller SR, Rodgers SE, Kesmodel S, Vivanco MDM, Slingerland JM.
      Many inflammation-associated diseases, including cancers, increase in women after menopause and with obesity. In contrast to anti-inflammatory actions of 17β-estradiol, we find estrone, which dominates after menopause, is pro-inflammatory. In human mammary adipocytes, cytokine expression increases with obesity, menopause, and cancer. Adipocyte:cancer cell interaction stimulates estrone- and NFκB-dependent pro-inflammatory cytokine upregulation. Estrone- and 17β-estradiol-driven transcriptomes differ. Estrone:ERα stimulates NFκB-mediated cytokine gene induction; 17β-estradiol opposes this. In obese mice, estrone increases and 17β-estradiol relieves inflammation. Estrone drives more rapid ER+ breast cancer growth in vivo. HSD17B14, which converts 17β-estradiol to estrone, associates with poor ER+ breast cancer outcome. Estrone and HSD17B14 upregulate inflammation, ALDH1 activity, and tumorspheres, while 17β-estradiol and HSD17B14 knockdown oppose these. Finally, a high intratumor estrone:17β-estradiol ratio increases tumor-initiating stem cells and ER+ cancer growth in vivo. These findings help explain why postmenopausal ER+ breast cancer increases with obesity, and offer new strategies for prevention and therapy.
    Keywords:  17β-estradiol; ER+ breast cancer; HSD17B14; NFκB; adipocytes; cancer stem cells; cytokines; estrone; inflammation; obesity
    DOI:  https://doi.org/10.1016/j.cmet.2020.05.008
  12. Nat Commun. 2020 Jun 01. 11(1): 2714
    Balsa E, Perry EA, Bennett CF, Jedrychowski M, Gygi SP, Doench JG, Puigserver P.
      Electron transport chain (ETC) defects occurring from mitochondrial disease mutations compromise ATP synthesis and render cells vulnerable to nutrient and oxidative stress conditions. This bioenergetic failure is thought to underlie pathologies associated with mitochondrial diseases. However, the precise metabolic processes resulting from a defective mitochondrial ETC that compromise cell viability under stress conditions are not entirely understood. We design a whole genome gain-of-function CRISPR activation screen using human mitochondrial disease complex I (CI) mutant cells to identify genes whose increased function rescue glucose restriction-induced cell death. The top hit of the screen is the cytosolic Malic Enzyme (ME1), that is sufficient to enable survival and proliferation of CI mutant cells under nutrient stress conditions. Unexpectedly, this metabolic rescue is independent of increased ATP synthesis through glycolysis or oxidative phosphorylation, but dependent on ME1-produced NADPH and glutathione (GSH). Survival upon nutrient stress or pentose phosphate pathway (PPP) inhibition depends on compensatory NADPH production through the mitochondrial one-carbon metabolism that is severely compromised in CI mutant cells. Importantly, this defective CI-dependent decrease in mitochondrial NADPH production pathway or genetic ablation of SHMT2 causes strong increases in inflammatory cytokine signatures associated with redox dependent induction of ASK1 and activation of stress kinases p38 and JNK. These studies find that a major defect of CI deficiencies is decreased mitochondrial one-carbon NADPH production that is associated with increased inflammation and cell death.
    DOI:  https://doi.org/10.1038/s41467-020-16423-1
  13. Cell Metab. 2020 Jun 02. pii: S1550-4131(20)30246-1. [Epub ahead of print]31(6): 1107-1119.e12
    Marenne G, Hendricks AE, Perdikari A, Bounds R, Payne F, Keogh JM, Lelliott CJ, Henning E, Pathan S, Ashford S, Bochukova EG, Mistry V, Daly A, Hayward C, , Wareham NJ, O'Rahilly S, Langenberg C, Wheeler E, Zeggini E, Farooqi IS, Barroso I.
      Obesity is genetically heterogeneous with monogenic and complex polygenic forms. Using exome and targeted sequencing in 2,737 severely obese cases and 6,704 controls, we identified three genes (PHIP, DGKI, and ZMYM4) with an excess burden of very rare predicted deleterious variants in cases. In cells, we found that nuclear PHIP (pleckstrin homology domain interacting protein) directly enhances transcription of pro-opiomelanocortin (POMC), a neuropeptide that suppresses appetite. Obesity-associated PHIP variants repressed POMC transcription. Our demonstration that PHIP is involved in human energy homeostasis through transcriptional regulation of central melanocortin signaling has potential diagnostic and therapeutic implications for patients with obesity and developmental delay. Additionally, we found an excess burden of predicted deleterious variants involving genes nearest to loci from obesity genome-wide association studies. Genes and gene sets influencing obesity with variable penetrance provide compelling evidence for a continuum of causality in the genetic architecture of obesity, and explain some of its missing heritability.
    Keywords:  POMC; association; function; gene set; genetics; severe childhood obesity
    DOI:  https://doi.org/10.1016/j.cmet.2020.05.007
  14. Nat Med. 2020 Jun 01.
    Dziubańska-Kusibab PJ, Berger H, Battistini F, Bouwman BAM, Iftekhar A, Katainen R, Cajuso T, Crosetto N, Orozco M, Aaltonen LA, Meyer TF.
      The mucosal epithelium is a common target of damage by chronic bacterial infections and the accompanying toxins, and most cancers originate from this tissue. We investigated whether colibactin, a potent genotoxin1 associated with certain strains of Escherichia coli2, creates a specific DNA-damage signature in infected human colorectal cells. Notably, the genomic contexts of colibactin-induced DNA double-strand breaks were enriched for an AT-rich hexameric sequence motif, associated with distinct DNA-shape characteristics. A survey of somatic mutations at colibactin target sites of several thousand cancer genomes revealed notable enrichment of this motif in colorectal cancers. Moreover, the exact double-strand-break loci corresponded with mutational hot spots in cancer genomes, reminiscent of a trinucleotide signature previously identified in healthy colorectal epithelial cells3. The present study provides evidence for the etiological role of colibactin in human cancer.
    DOI:  https://doi.org/10.1038/s41591-020-0908-2
  15. Nature. 2020 Jun;582(7810): 115-118
    von Appen A, LaJoie D, Johnson IE, Trnka MJ, Pick SM, Burlingame AL, Ullman KS, Frost A.
      During cell division, remodelling of the nuclear envelope enables chromosome segregation by the mitotic spindle1. The reformation of sealed nuclei requires ESCRTs (endosomal sorting complexes required for transport) and LEM2, a transmembrane ESCRT adaptor2-4. Here we show how the ability of LEM2 to condense on microtubules governs the activation of ESCRTs and coordinated spindle disassembly. The LEM motif of LEM2 binds BAF, conferring on LEM2 an affinity for chromatin5,6, while an adjacent low-complexity domain (LCD) promotes LEM2 phase separation. A proline-arginine-rich sequence within the LCD binds to microtubules and targets condensation of LEM2 to spindle microtubules that traverse the nascent nuclear envelope. Furthermore, the winged-helix domain of LEM2 activates the ESCRT-II/ESCRT-III hybrid protein CHMP7 to form co-oligomeric rings. Disruption of these events in human cells prevented the recruitment of downstream ESCRTs, compromised spindle disassembly, and led to defects in nuclear integrity and DNA damage. We propose that during nuclear reassembly LEM2 condenses into a liquid-like phase and coassembles with CHMP7 to form a macromolecular O-ring seal at the confluence between membranes, chromatin and the spindle. The properties of LEM2 described here, and the homologous architectures of related inner nuclear membrane proteins7,8, suggest that phase separation may contribute to other critical envelope functions, including interphase repair8-13 and chromatin organization14-17.
    DOI:  https://doi.org/10.1038/s41586-020-2232-x
  16. Mol Cell. 2020 May 28. pii: S1097-2765(20)30302-6. [Epub ahead of print]
    Ali ES, Sahu U, Villa E, O'Hara BP, Gao P, Beaudet C, Wood AW, Asara JM, Ben-Sahra I.
      The RAS-ERK/MAPK (RAS-extracellular signal-regulated kinase/mitogen-activated protein kinase) pathway integrates growth-promoting signals to stimulate cell growth and proliferation, at least in part, through alterations in metabolic gene expression. However, examples of direct and rapid regulation of the metabolic pathways by the RAS-ERK pathway remain elusive. We find that physiological and oncogenic ERK signaling activation leads to acute metabolic flux stimulation through the de novo purine synthesis pathway, thereby increasing building block availability for RNA and DNA synthesis, which is required for cell growth and proliferation. We demonstrate that ERK2, but not ERK1, phosphorylates the purine synthesis enzyme PFAS (phosphoribosylformylglycinamidine synthase) at T619 in cells to stimulate de novo purine synthesis. The expression of nonphosphorylatable PFAS (T619A) decreases purine synthesis, RAS-dependent cancer cell-colony formation, and tumor growth. Thus, ERK2-mediated PFAS phosphorylation facilitates the increase in nucleic acid synthesis required for anabolic cell growth and proliferation.
    Keywords:  ERK; FGAM; MAPK; PFAS; RAS; cancer; nucleotide synthesis; posttranslational modification; purine metabolism; tumor growth
    DOI:  https://doi.org/10.1016/j.molcel.2020.05.001
  17. Nat Med. 2020 Jun 01.
    Zviran A, Schulman RC, Shah M, Hill STK, Deochand S, Khamnei CC, Maloney D, Patel K, Liao W, Widman AJ, Wong P, Callahan MK, Ha G, Reed S, Rotem D, Frederick D, Sharova T, Miao B, Kim T, Gydush G, Rhoades J, Huang KY, Omans ND, Bolan PO, Lipsky AH, Ang C, Malbari M, Spinelli CF, Kazancioglu S, Runnels AM, Fennessey S, Stolte C, Gaiti F, Inghirami GG, Adalsteinsson V, Houck-Loomis B, Ishii J, Wolchok JD, Boland G, Robine N, Altorki NK, Landau DA.
      In many areas of oncology, we lack sensitive tools to track low-burden disease. Although cell-free DNA (cfDNA) shows promise in detecting cancer mutations, we found that the combination of low tumor fraction (TF) and limited number of DNA fragments restricts low-disease-burden monitoring through the prevailing deep targeted sequencing paradigm. We reasoned that breadth may supplant depth of sequencing to overcome the barrier of cfDNA abundance. Whole-genome sequencing (WGS) of cfDNA allowed ultra-sensitive detection, capitalizing on the cumulative signal of thousands of somatic mutations observed in solid malignancies, with TF detection sensitivity as low as 10-5. The WGS approach enabled dynamic tumor burden tracking and postoperative residual disease detection, associated with adverse outcome. Thus, we present an orthogonal framework for cfDNA cancer monitoring via genome-wide mutational integration, enabling ultra-sensitive detection, overcoming the limitation of cfDNA abundance and empowering treatment optimization in low-disease-burden oncology care.
    DOI:  https://doi.org/10.1038/s41591-020-0915-3
  18. Nat Commun. 2020 Jun 03. 11(1): 2790
    Beas AO, Gordon PB, Prentiss CL, Olsen CP, Kukurugya MA, Bennett BD, Parkhurst SM, Gottschling DE.
      Age-dependent changes in metabolism can manifest as cellular lipid accumulation, but how this accumulation is regulated or impacts longevity is poorly understood. We find that Saccharomyces cerevisiae accumulate lipid droplets (LDs) during aging. We also find that over-expressing BNA2, the first Biosynthesis of NAD+ (kynurenine) pathway gene, reduces LD accumulation during aging and extends lifespan. Mechanistically, this LD accumulation during aging is not linked to NAD+ levels, but is anti-correlated with metabolites of the shikimate and aromatic amino acid biosynthesis (SA) pathways (upstream of BNA2), which produce tryptophan (the Bna2p substrate). We provide evidence that over-expressed BNA2 skews glycolytic flux from LDs towards the SA-BNA pathways, effectively reducing LDs. Importantly, we find that accumulation of LDs does not shorten lifespan, but does protect aged cells against stress. Our findings reveal how lipid accumulation impacts longevity, and how aging cell metabolism can be rewired to modulate lipid accumulation independently from longevity.
    DOI:  https://doi.org/10.1038/s41467-020-16358-7
  19. Sci Adv. 2020 Apr;6(17): eaaz6997
    Feld L, Kellerman L, Mukherjee A, Livne A, Bouchbinder E, Wolfenson H.
      Cells' ability to apply contractile forces to their environment and to sense its mechanical properties (e.g., rigidity) are among their most fundamental features. Yet, the interrelations between contractility and mechanosensing, in particular, whether contractile force generation depends on mechanosensing, are not understood. We use theory and extensive experiments to study the time evolution of cellular contractile forces and show that they are generated by time-dependent actomyosin contractile displacements that are independent of the environment's rigidity. Consequently, contractile forces are nonmechanosensitive. We further show that the force-generating displacements are directly related to the evolution of the actomyosin network, most notably to the time-dependent concentration of F-actin. The emerging picture of force generation and mechanosensitivity offers a unified framework for understanding contractility.
    DOI:  https://doi.org/10.1126/sciadv.aaz6997
  20. Autophagy. 2020 Jun 02. 1-12
    Varusai TM, Jupe S, Sevilla C, Matthews L, Gillespie M, Stein L, Wu G, D'Eustachio P, Metzakopian E, Hermjakob H.
      The 21st century has revealed much about the fundamental cellular process of autophagy. Autophagy controls the catabolism and recycling of various cellular components both as a constitutive process and as a response to stress and foreign material invasion. There is considerable knowledge of the molecular mechanisms of autophagy, and this is still growing as new modalities emerge. There is a need to investigate autophagy mechanisms reliably, comprehensively and conveniently. Reactome is a freely available knowledgebase that consists of manually curated molecular events (reactions) organized into cellular pathways (https://reactome.org/). Pathways/reactions in Reactome are hierarchically structured, graphically presented and extensively annotated. Data analysis tools, such as pathway enrichment, expression data overlay and species comparison, are also available. For customized analysis, information can also be programmatically queried. Here, we discuss the curation and annotation of the molecular mechanisms of autophagy in Reactome. We also demonstrate the value that Reactome adds to research by reanalyzing a previously published work on genome-wide CRISPR screening of autophagy components.ABBREVIATIONS: CMA: chaperone-mediated autophagy; GO: Gene Ontology; MA: macroautophagy; MI: microautophagy; MTOR: mechanistic target of rapamycin kinase; SQSTM1: sequestosome 1.
    Keywords:  Annotation; Reactome; autophagy; biocuration; curation; enrichment analysis; knowledgebase; mechanistic analysis; molecular reactions; pathways
    DOI:  https://doi.org/10.1080/15548627.2020.1761659
  21. Cell. 2020 May 22. pii: S0092-8674(20)30567-5. [Epub ahead of print]
    Martin JK, Sheehan JP, Bratton BP, Moore GM, Mateus A, Li SH, Kim H, Rabinowitz JD, Typas A, Savitski MM, Wilson MZ, Gitai Z.
      The rise of antibiotic resistance and declining discovery of new antibiotics has created a global health crisis. Of particular concern, no new antibiotic classes have been approved for treating Gram-negative pathogens in decades. Here, we characterize a compound, SCH-79797, that kills both Gram-negative and Gram-positive bacteria through a unique dual-targeting mechanism of action (MoA) with undetectably low resistance frequencies. To characterize its MoA, we combined quantitative imaging, proteomic, genetic, metabolomic, and cell-based assays. This pipeline demonstrates that SCH-79797 has two independent cellular targets, folate metabolism and bacterial membrane integrity, and outperforms combination treatments in killing methicillin-resistant Staphylococcus aureus (MRSA) persisters. Building on the molecular core of SCH-79797, we developed a derivative, Irresistin-16, with increased potency and showed its efficacy against Neisseria gonorrhoeae in a mouse vaginal infection model. This promising antibiotic lead suggests that combining multiple MoAs onto a single chemical scaffold may be an underappreciated approach to targeting challenging bacterial pathogens.
    Keywords:  Acinetobacter baumannii; Gram-negative pathogens; Neisseria gonorrhoeae; antibiotics; broad spectrum; dual-target drugs; folate metabolism; membrane disrupting
    DOI:  https://doi.org/10.1016/j.cell.2020.05.005
  22. Nat Genet. 2020 Jun 01.
    Jaeger MG, Schwalb B, Mackowiak SD, Velychko T, Hanzl A, Imrichova H, Brand M, Agerer B, Chorn S, Nabet B, Ferguson FM, Müller AC, Bergthaler A, Gray NS, Bradner JE, Bock C, Hnisz D, Cramer P, Winter GE.
      The Mediator complex directs signals from DNA-binding transcription factors to RNA polymerase II (Pol II). Despite this pivotal position, mechanistic understanding of Mediator in human cells remains incomplete. Here we quantified Mediator-controlled Pol II kinetics by coupling rapid subunit degradation with orthogonal experimental readouts. In agreement with a model of condensate-driven transcription initiation, large clusters of hypophosphorylated Pol II rapidly disassembled upon Mediator degradation. This was accompanied by a selective and pronounced disruption of cell-type-specifying transcriptional circuits, whose constituent genes featured exceptionally high rates of Pol II turnover. Notably, the transcriptional output of most other genes was largely unaffected by acute Mediator ablation. Maintenance of transcriptional activity at these genes was linked to an unexpected CDK9-dependent compensatory feedback loop that elevated Pol II pause release rates across the genome. Collectively, our work positions human Mediator as a globally acting coactivator that selectively safeguards the functionality of cell-type-specifying transcriptional networks.
    DOI:  https://doi.org/10.1038/s41588-020-0635-0
  23. Elife. 2020 Jun 03. pii: e54341. [Epub ahead of print]9
    Shipkovenska G, Durango A, Kalocsay M, Gygi SP, Moazed D.
      Heterochromatic domains containing histone H3 lysine 9 methylation (H3K9me) can be epigenetically inherited independently of underlying DNA sequence. To gain insight into the mechanisms that mediate epigenetic inheritance, we used a Schizosaccharomyces pombe inducible heterochromatin formation system to perform a genetic screen for mutations that abolish heterochromatin inheritance without affecting its establishment. We identified mutations in several pathways, including the conserved and essential Rix1-associated complex (henceforth the rixosome), which contains RNA endonuclease and polynucleotide kinase activities with known roles in ribosomal RNA processing. We show that the rixosome is required for spreading and epigenetic inheritance of heterochromatin in fission yeast. Viable rixosome mutations that disrupt its association with Swi6/HP1 fail to localize to heterochromatin, lead to accumulation of heterochromatic RNAs, and block spreading of H3K9me and silencing into actively transcribed regions. These findings reveal a new pathway for degradation of heterochromatic RNAs with essential roles in heterochromatin spreading and inheritance.
    Keywords:  H3K9 methylation; RNA degradation; Rix1; S. pombe; cell biology; chromosomes; epigenetics; gene expression; heterochromatin; rixosome
    DOI:  https://doi.org/10.7554/eLife.54341
  24. J Cell Sci. 2020 May 22. pii: jcs228114. [Epub ahead of print]133(10):
    Birgisdottir ÅB, Johansen T.
      Autophagy and endocytosis are membrane-vesicle-based cellular pathways for degradation and recycling of intracellular and extracellular components, respectively. These pathways have a common endpoint at the lysosome, where their cargo is degraded. In addition, the two pathways intersect at different stages during vesicle formation, fusion and trafficking, and share parts of the molecular machinery. Accumulating evidence shows that autophagy is dependent upon endocytosis and vice versa. The emerging joint network of autophagy and endocytosis is of vital importance for cellular metabolism and signaling, and thus also highly relevant in disease settings. In this Review, we will discuss examples of how the autophagy machinery impacts on endocytosis and cell signaling, and highlight how endocytosis regulates the different steps in autophagy in mammalian cells. Finally, we will focus on the interplay of these pathways in the quality control of their common endpoint, the lysosome.
    Keywords:  Autophagy; Endocytosis; LAP; LC3-associated phagocytosis; Lysosome; Phagophore
    DOI:  https://doi.org/10.1242/jcs.228114
  25. J Natl Cancer Inst. 2020 Jun 04. pii: djaa073. [Epub ahead of print]
    Michelakos T, Cai L, Villani V, Sabbatino F, Kontos F, Fernández-Del Castillo C, Yamada T, Neyaz A, Taylor MS, Deshpande V, Kurokawa T, Ting DT, Qadan M, Weekes CD, Allen JN, Clark JW, Hong TS, Ryan DP, Wo JY, Warshaw AL, Lillemoe KD, Ferrone S, Ferrone CR.
      BACKGROUND: Neoadjuvant FOLFIRINOX and chemoradiation have been utilized to downstage borderline and locally advanced pancreatic ductal adenocarcinoma (PDAC). Whether neoadjuvant therapy-induced tumor immune response contributes to the improved survival is unknown. Therefore, we evaluated whether neoadjuvant therapy induces an immune response towards PDAC.METHODS: Clinicopathologic variables were collected for surgically resected PDACs at the Massachusetts General Hospital (1998-2016). Neoadjuvant regimens included FOLFIRINOX with/without chemoradiation, proton chemoradiation (25Gy), photon chemoradiation (50.4Gy) or no neoadjuvant therapy. HLA class I and II expression, and immune cell infiltration (CD4+, FoxP3+, CD8+, Granzyme B+ cells and M2 macrophages) were analyzed immunohistochemically and correlated with clinicopathologic variables. The antitumor immune response was compared among neoadjuvant therapy regimens. All statistical tests were two-sided.
    RESULTS: Two hundred forty-eight PDAC patients were included. Median age was 64y; 50.0% were female. HLA-A defects were less frequent in the FOLFIRINOX cohort (p=.006). HLA class II expression was lowest in photon and highest in proton patients (p=.02). The FOLFIRINOX cohort exhibited the densest CD8+ cell infiltration (p<.001). FOLFIRINOX and proton patients had the highest CD4+ and lowest T regulatory (FoxP3+) cell density, respectively. M2 macrophage density was statistically significantly higher in the treatment-naïve group (p<.001), in which dense M2 macrophage infiltration was an independent predictor of poor OS.
    CONCLUSIONS: Neoadjuvant FOLFIRINOX with/without chemoradiation may induce immunologically relevant changes in the tumor microenvironment. It may reduce HLA-A defects, increase CD8+ cell density and decrease T regulatory cell and M2 macrophage density. Therefore, neoadjuvant FOLFIRINOX therapy may benefit from combinations with checkpoint inhibitors, which can enhance patients' antitumor immune response.
    Keywords:  FOLFIRINOX; Human Leukocyte Antigen (HLA); Immune Response; Macrophage; Pancreatic Ductal Adenocarcinoma (PDAC)
    DOI:  https://doi.org/10.1093/jnci/djaa073
  26. Cell. 2020 Jun 03. pii: S0092-8674(20)30555-9. [Epub ahead of print]
    Vaccaro A, Kaplan Dor Y, Nambara K, Pollina EA, Lin C, Greenberg ME, Rogulja D.
      The view that sleep is essential for survival is supported by the ubiquity of this behavior, the apparent existence of sleep-like states in the earliest animals, and the fact that severe sleep loss can be lethal. The cause of this lethality is unknown. Here we show, using flies and mice, that sleep deprivation leads to accumulation of reactive oxygen species (ROS) and consequent oxidative stress, specifically in the gut. ROS are not just correlates of sleep deprivation but drivers of death: their neutralization prevents oxidative stress and allows flies to have a normal lifespan with little to no sleep. The rescue can be achieved with oral antioxidant compounds or with gut-targeted transgenic expression of antioxidant enzymes. We conclude that death upon severe sleep restriction can be caused by oxidative stress, that the gut is central in this process, and that survival without sleep is possible when ROS accumulation is prevented. VIDEO ABSTRACT.
    Keywords:  antioxidants; free radicals; gut; oxidative stress; reactive oxygen species; sleep; sleep deprivation; survival
    DOI:  https://doi.org/10.1016/j.cell.2020.04.049
  27. Cell Rep. 2020 Jun 02. pii: S2211-1247(20)30671-9. [Epub ahead of print]31(9): 107701
    Becker LM, O'Connell JT, Vo AP, Cain MP, Tampe D, Bizarro L, Sugimoto H, McGow AK, Asara JM, Lovisa S, McAndrews KM, Zielinski R, Lorenzi PL, Zeisberg M, Raza S, LeBleu VS, Kalluri R.
      The mechanistic contributions of cancer-associated fibroblasts (CAFs) in breast cancer progression remain to be fully understood. While altered glucose metabolism in CAFs could fuel cancer cells, how such metabolic reprogramming emerges and is sustained needs further investigation. Studying fibroblasts isolated from patients with benign breast tissues and breast cancer, in conjunction with multiple animal models, we demonstrate that CAFs exhibit a metabolic shift toward lactate and pyruvate production and fuel biosynthetic pathways of cancer cells. The depletion or suppression of the lactate production of CAFs alter the tumor metabolic profile and impede tumor growth. The glycolytic phenotype of the CAFs is in part sustained through epigenetic reprogramming of HIF-1α and glycolytic enzymes. Hypoxia induces epigenetic reprogramming of normal fibroblasts, resulting in a pro-glycolytic, CAF-like transcriptome. Our findings suggest that the glucose metabolism of CAFs evolves during tumor progression, and their breast cancer-promoting phenotype is partly mediated by oxygen-dependent epigenetic modifications.
    Keywords:  breast cancer; cancer-associated fibroblasts; epigenetic alterations; hypoxia; metabolism
    DOI:  https://doi.org/10.1016/j.celrep.2020.107701
  28. Nature. 2020 Jun 03.
    Sulkowski PL, Oeck S, Dow J, Economos NG, Mirfakhraie L, Liu Y, Noronha K, Bao X, Li J, Shuch BM, King MC, Bindra RS, Glazer PM.
      Deregulation of metabolism and disruption of genome integrity are hallmarks of cancer1. Increased levels of the metabolites 2-hydroxyglutarate, succinate and fumarate occur in human malignancies owing to somatic mutations in the isocitrate dehydrogenase-1 or -2 (IDH1 or IDH2) genes, or germline mutations in the fumarate hydratase (FH) and succinate dehydrogenase genes (SDHA, SDHB, SDHC and SDHD), respectively2-4. Recent work has made an unexpected connection between these metabolites and DNA repair by showing that they suppress the pathway of homology-dependent repair (HDR)5,6 and confer an exquisite sensitivity to inhibitors of poly (ADP-ribose) polymerase (PARP) that are being tested in clinical trials. However, the mechanism by which these oncometabolites inhibit HDR remains poorly understood. Here we determine the pathway by which these metabolites disrupt DNA repair. We show that oncometabolite-induced inhibition of the lysine demethylase KDM4B results in aberrant hypermethylation of histone 3 lysine 9 (H3K9) at loci surrounding DNA breaks, masking a local H3K9 trimethylation signal that is essential for the proper execution of HDR. Consequently, recruitment of TIP60 and ATM, two key proximal HDR factors, is substantially impaired at DNA breaks, with reduced end resection and diminished recruitment of downstream repair factors. These findings provide a mechanistic basis for oncometabolite-induced HDR suppression and may guide effective strategies to exploit these defects for therapeutic gain.
    DOI:  https://doi.org/10.1038/s41586-020-2363-0
  29. Elife. 2020 Jun 05. pii: e55438. [Epub ahead of print]9
    Schaich MA, Sanford SL, Welfer GA, Johnson SA, Khoang TH, Opresko PL, Freudenthal BD.
      Telomerase extends telomere sequences at chromosomal ends to protect genomic DNA. During this process it must select the correct nucleotide from a pool of nucleotides with various sugars and base pairing properties, which is critically important for the proper capping of telomeric sequences by shelterin. Unfortunately, how telomerase selects correct nucleotides is unknown. Here, we determined structures of Tribolium castaneum telomerase reverse transcriptase (TERT) throughout its catalytic cycle and mapped the active site residues responsible for nucleoside selection, metal coordination, triphosphate binding, and RNA template stabilization. We found that TERT inserts a mismatch or ribonucleotide ~1 in 10,000 and ~1 in 14,000 insertion events, respectively. At biological ribonucleotide concentrations, these rates translate to ~40 ribonucleotides inserted per 10 kilobases. Human telomerase assays determined a conserved tyrosine steric gate regulates ribonucleotide insertion into telomeres. Cumulatively, our work provides insight into how telomerase selects the proper nucleotide to maintain telomere integrity.
    Keywords:  T. castaneum; biochemistry; chemical biology; fidelity; molecular biophysics; reverse transcriptase; ribonucleotide; structural biology; telomerase
    DOI:  https://doi.org/10.7554/eLife.55438
  30. EMBO Rep. 2020 Jun 02. e48686
    Han H, Tan J, Wang R, Wan H, He Y, Yan X, Guo J, Gao Q, Li J, Shang S, Chen F, Tian R, Liu W, Liao L, Tang B, Zhang Z.
      Impairment of PINK1/parkin-mediated mitophagy is currently proposed to be the molecular basis of mitochondrial abnormality in Parkinson's disease (PD). We here demonstrate that PINK1 directly phosphorylates Drp1 on S616. Drp1S616 phosphorylation is significantly reduced in cells and mouse tissues deficient for PINK1, but unaffected by parkin inactivation. PINK1-mediated mitochondrial fission is Drp1S616 phosphorylation dependent. Overexpression of either wild-type Drp1 or of the phosphomimetic mutant Drp1S616D , but not a dephosphorylation-mimic mutant Drp1S616A , rescues PINK1 deficiency-associated phenotypes in Drosophila. Moreover, Drp1 restores PINK1-dependent mitochondrial fission in ATG5-null cells and ATG7-null Drosophila. Reduced Drp1S616 phosphorylation is detected in fibroblasts derived from 4 PD patients harboring PINK1 mutations and in 4 out of 7 sporadic PD cases. Taken together, we have identified Drp1 as a substrate of PINK1 and a novel mechanism how PINK1 regulates mitochondrial fission independent of parkin and autophagy. Our results further link impaired PINK1-mediated Drp1S616 phosphorylation with the pathogenesis of both familial and sporadic PD.
    Keywords:  Parkinson’s disease; autophagy; human dermal fibroblasts; mitochondrial dynamics; parkin
    DOI:  https://doi.org/10.15252/embr.201948686
  31. Nat Commun. 2020 Jun 03. 11(1): 2795
    Cui L, Chen SY, Lerbs T, Lee JW, Domizi P, Gordon S, Kim YH, Nolan G, Betancur P, Wernig G.
      The transcription factor JUN is highly expressed in pulmonary fibrosis. Its induction in mice drives lung fibrosis, which is abrogated by administration of anti-CD47. Here, we use high-dimensional mass cytometry to profile protein expression and secretome of cells from patients with pulmonary fibrosis. We show that JUN is activated in fibrotic fibroblasts that expressed increased CD47 and PD-L1. Using ATAC-seq and ChIP-seq, we found that activation of JUN rendered promoters and enhancers of CD47 and PD-L1 accessible. We further detect increased IL-6 that amplified JUN-mediated CD47 enhancer activity and protein expression. Using an in vivo mouse model of fibrosis, we found two distinct mechanisms by which blocking IL-6, CD47 and PD-L1 reversed fibrosis, by increasing phagocytosis of profibrotic fibroblasts and by eliminating suppressive effects on adaptive immunity. Our results identify specific immune mechanisms that promote fibrosis and suggest a therapeutic approach that could be used alongside conventional anti-fibrotics for pulmonary fibrosis.
    DOI:  https://doi.org/10.1038/s41467-020-16466-4
  32. J Cell Biol. 2020 Aug 03. pii: e201903127. [Epub ahead of print]219(8):
    Yamamoto YH, Kasai A, Omori H, Takino T, Sugihara M, Umemoto T, Hamasaki M, Hatta T, Natsume T, Morimoto RI, Arai R, Waguri S, Sato M, Sato K, Bar-Nun S, Yoshimori T, Noda T, Nagata K.
      In macroautophagy, membrane structures called autophagosomes engulf substrates and deliver them for lysosomal degradation. Autophagosomes enwrap a variety of targets with diverse sizes, from portions of cytosol to larger organelles. However, the mechanism by which autophagosome size is controlled remains elusive. We characterized a novel ER membrane protein, ERdj8, in mammalian cells. ERdj8 localizes to a meshwork-like ER subdomain along with phosphatidylinositol synthase (PIS) and autophagy-related (Atg) proteins. ERdj8 overexpression extended the size of the autophagosome through its DnaJ and TRX domains. ERdj8 ablation resulted in a defect in engulfing larger targets. C. elegans, in which the ERdj8 orthologue dnj-8 was knocked down, could perform autophagy on smaller mitochondria derived from the paternal lineage but not the somatic mitochondria. Thus, ERdj8 may play a critical role in autophagosome formation by providing the capacity to target substrates of diverse sizes for degradation.
    DOI:  https://doi.org/10.1083/jcb.201903127
  33. J Immunol. 2020 Jun 01. pii: ji2000108. [Epub ahead of print]
    Sbierski-Kind J, Mai K, Kath J, Jurisch A, Streitz M, Kuchenbecker L, Babel N, Nienen M, Jürchott K, Spranger L, Jumpertz von Schwartzenberg R, Decker AM, Krüger U, Volk HD, Spranger J.
      The worldwide epidemic of overweight and obesity has led to an increase in associated metabolic comorbidities. Obesity induces chronic low-grade inflammation in white adipose tissue (WAT). However, the function and regulation of both innate and adaptive immune cells in human WAT under conditions of obesity and calorie restriction (CR) is not fully understood yet. Using a randomized interventional design, we investigated postmenopausal overweight or obese female subjects who either underwent CR for 3 mo followed by a 4-wk phase of weight maintenance or had to maintain a stable weight over the whole study period. A comprehensive immune phenotyping protocol was conducted using validated multiparameter flow cytometry analysis in blood and s.c. WAT (SAT). The TCR repertoire was analyzed by next-generation sequencing and cytokine levels were determined in SAT. Metabolic parameters were determined by hyperinsulinemic-euglycemic clamp. We found that insulin resistance correlates significantly with a shift toward the memory T cell compartment in SAT. TCR analysis revealed a diverse repertoire in SAT of overweight or obese individuals. Additionally, whereas weight loss improved systemic insulin sensitivity in the intervention group, SAT displayed no significant improvement of inflammatory parameters (cytokine levels and leukocyte subpopulations) compared with the control group. Our data demonstrate the accumulation of effector memory T cells in obese SAT and an association between systemic glucose homeostasis and inflammatory parameters in obese females. The long-standing effect of obesity-induced changes in SAT was demonstrated by preserved immune cell composition after short-term CR-induced weight loss.
    DOI:  https://doi.org/10.4049/jimmunol.2000108
  34. Dev Cell. 2020 May 27. pii: S1534-5807(20)30400-7. [Epub ahead of print]
    Hino N, Rossetti L, Marín-Llauradó A, Aoki K, Trepat X, Matsuda M, Hirashima T.
      During collective migration of epithelial cells, the migration direction is aligned over a tissue-scale expanse. Although the collective cell migration is known to be directed by mechanical forces transmitted via cell-cell junctions, it remains elusive how the intercellular force transmission is coordinated with intracellular biochemical signaling to achieve collective movements. Here, we show that intercellular coupling of extracellular signal-regulated kinase (ERK)-mediated mechanochemical feedback yields long-distance transmission of guidance cues. Mechanical stretch activates ERK through epidermal growth factor receptor (EGFR) activation, and ERK activation triggers cell contraction. The contraction of the activated cell pulls neighboring cells, evoking another round of ERK activation and contraction in the neighbors. Furthermore, anisotropic contraction based on front-rear polarization guarantees unidirectional propagation of ERK activation, and in turn, the ERK activation waves direct multicellular alignment of the polarity, leading to long-range ordered migration. Our findings reveal that mechanical forces mediate intercellular signaling underlying sustained transmission of guidance cues for collective cell migration.
    Keywords:  EGFR; ERK/MAPK; FRET; collective cell migration; front-rear polarity; intercellular signal transfer; mathematical model; mechanochemical feedback; mechanotransduction; wave propagation
    DOI:  https://doi.org/10.1016/j.devcel.2020.05.011
  35. Science. 2020 Jun 05. 368(6495): 1127-1131
    Cipponi A, Goode DL, Bedo J, McCabe MJ, Pajic M, Croucher DR, Rajal AG, Junankar SR, Saunders DN, Lobachevsky P, Papenfuss AT, Nessem D, Nobis M, Warren SC, Timpson P, Cowley M, Vargas AC, Qiu MR, Generali DG, Keerthikumar S, Nguyen U, Corcoran NM, Long GV, Blay JY, Thomas DM.
      In microorganisms, evolutionarily conserved mechanisms facilitate adaptation to harsh conditions through stress-induced mutagenesis (SIM). Analogous processes may underpin progression and therapeutic failure in human cancer. We describe SIM in multiple in vitro and in vivo models of human cancers under nongenotoxic drug selection, paradoxically enhancing adaptation at a competing intrinsic fitness cost. A genome-wide approach identified the mechanistic target of rapamycin (MTOR) as a stress-sensing rheostat mediating SIM across multiple cancer types and conditions. These observations are consistent with a two-phase model for drug resistance, in which an initially rapid expansion of genetic diversity is counterbalanced by an intrinsic fitness penalty, subsequently normalizing to complete adaptation under the new conditions. This model suggests synthetic lethal strategies to minimize resistance to anticancer therapy.
    DOI:  https://doi.org/10.1126/science.aau8768
  36. JCI Insight. 2020 Jun 04. pii: 131834. [Epub ahead of print]5(11):
    Liu Y, Lear TB, Verma M, Wang KZ, Otero PA, McKelvey AC, Dunn SR, Steer E, Bateman NW, Wu C, Jiang Y, Weathington NM, Rojas M, Chu CT, Chen BB, Mallampalli RK.
      Mitochondrial quality control is mediated by the PTEN-induced kinase 1 (PINK1), a cytoprotective protein that is dysregulated in inflammatory lung injury and neurodegenerative diseases. Here, we show that a ubiquitin E3 ligase receptor component, FBXO7, targets PINK1 for its cellular disposal. FBXO7, by mediating PINK1 ubiquitylation and degradation, was sufficient to induce mitochondrial injury and inflammation in experimental pneumonia. A computational simulation-based screen led to the identification of a small molecule, BC1464, which abrogated FBXO7 and PINK1 association, leading to increased cellular PINK1 concentrations and activities, and limiting mitochondrial damage. BC1464 exerted antiinflammatory activity in human tissue explants and murine lung inflammation models. Furthermore, BC1464 conferred neuroprotection in primary cortical neurons, human neuroblastoma cells, and patient-derived cells in several culture models of Parkinson's disease. The data highlight a unique opportunity to use small molecule antagonists that disrupt PINK1 interaction with the ubiquitin apparatus to enhance mitochondrial quality, limit inflammatory injury, and maintain neuronal viability.
    Keywords:  Cell stress; Drug therapy; Neuroscience; Parkinson’s disease
    DOI:  https://doi.org/10.1172/jci.insight.131834
  37. Nat Commun. 2020 Jun 02. 11(1): 2768
    Layton TB, Williams L, McCann F, Zhang M, Fritszche M, Colin-York H, Cabrita M, Ng MTH, Feldmann M, Samson S, Furniss D, Xie W, Nanchahal J.
      Fibrotic disorders are some of the most devastating and poorly treated conditions in developed nations, yet effective therapeutics are not identified for many of them. A major barrier for the identification of targets and successful clinical translation is a limited understanding of the human fibrotic microenvironment. Here, we construct a stromal cell atlas of human fibrosis at single cell resolution from patients with Dupuytren's disease, a localized fibrotic condition of the hand. A molecular taxonomy of the fibrotic milieu characterises functionally distinct stromal cell types and states, including a subset of immune regulatory ICAM1+ fibroblasts. In developing fibrosis, myofibroblasts exist along an activation continuum of phenotypically distinct populations. We also show that the tetraspanin CD82 regulates cell cycle progression and can be used as a cell surface marker of myofibroblasts. These findings have important implications for targeting core pathogenic drivers of human fibrosis.
    DOI:  https://doi.org/10.1038/s41467-020-16264-y
  38. Cell. 2020 May 28. pii: S0092-8674(20)30147-1. [Epub ahead of print]
    Wong PP, Muñoz-Félix JM, Hijazi M, Kim H, Robinson SD, De Luxán-Delgado B, Rodríguez-Hernández I, Maiques O, Meng YM, Meng Q, Bodrug N, Dukinfield MS, Reynolds LE, Elia G, Clear A, Harwood C, Wang Y, Campbell JJ, Singh R, Zhang P, Schall TJ, Matchett KP, Henderson NC, Szlosarek PW, Dreger SA, Smith S, Jones JL, Gribben JG, Cutillas PR, Meier P, Sanz-Moreno V, Hodivala-Dilke KM.
      Enhanced blood vessel (BV) formation is thought to drive tumor growth through elevated nutrient delivery. However, this observation has overlooked potential roles for mural cells in directly affecting tumor growth independent of BV function. Here we provide clinical data correlating high percentages of mural-β3-integrin-negative tumor BVs with increased tumor sizes but no effect on BV numbers. Mural-β3-integrin loss also enhances tumor growth in implanted and autochthonous mouse tumor models with no detectable effects on BV numbers or function. At a molecular level, mural-cell β3-integrin loss enhances signaling via FAK-p-HGFR-p-Akt-p-p65, driving CXCL1, CCL2, and TIMP-1 production. In particular, mural-cell-derived CCL2 stimulates tumor cell MEK1-ERK1/2-ROCK2-dependent signaling and enhances tumor cell survival and tumor growth. Overall, our data indicate that mural cells can control tumor growth via paracrine signals regulated by β3-integrin, providing a previously unrecognized mechanism of cancer growth control.
    Keywords:  mural cell; paracrine; β3-integrin
    DOI:  https://doi.org/10.1016/j.cell.2020.02.003
  39. Mol Cell. 2020 May 29. pii: S1097-2765(20)30306-3. [Epub ahead of print]
    Laprade H, Querido E, Smith MJ, Guérit D, Crimmins H, Conomos D, Pourret E, Chartrand P, Sfeir A.
      Extension of telomeres is a critical step in the immortalization of cancer cells. This complex reaction requires proper spatiotemporal coordination of telomerase and telomeres and remains poorly understood at the cellular level. To understand how cancer cells execute this process, we combine CRISPR genome editing and MS2 RNA tagging to image single molecules of telomerase RNA (hTR). Real-time dynamics and photoactivation experiments of hTR in Cajal bodies (CBs) reveal that hTERT controls the exit of hTR from CBs. Single-molecule tracking of hTR at telomeres shows that TPP1-mediated recruitment results in short telomere-telomerase scanning interactions, and then base pairing between hTR and telomere ssDNA promotes long interactions required for stable telomerase retention. Interestingly, POT1 OB-fold mutations that result in abnormally long telomeres in cancers act by enhancing this retention step. In summary, single-molecule imaging unveils the life cycle of telomerase RNA and provides a framework to reveal how cancer-associated mutations mechanistically drive defects in telomere homeostasis.
    Keywords:  ATM; ATR; POT1; cancer cellsĆ; fluorescent in situ hybridization; hTR RNA; single-molecule imaging; telomerase; telomeres
    DOI:  https://doi.org/10.1016/j.molcel.2020.05.005
  40. Elife. 2020 Jun 05. pii: e55913. [Epub ahead of print]9
    Hartmann J, Wong M, Gallo E, Gilmour D.
      Quantitative microscopy is becoming increasingly crucial in efforts to disentangle the complexity of organogenesis, yet adoption of the potent new toolbox provided by modern data science has been slow, primarily because it is often not directly applicable to developmental imaging data. We tackle this issue with a newly developed algorithm that uses point cloud-based morphometry to unpack the rich information encoded in 3D image data into a straightforward numerical representation. This enabled us to employ data science tools, including machine learning, to analyze and integrate cell morphology, intracellular organization, gene expression and annotated contextual knowledge. We apply these techniques to construct and explore a quantitative atlas of cellular architecture for the zebrafish posterior lateral line primordium, an experimentally tractable model of complex self-organized organogenesis. In doing so, we are able to retrieve both previously established and novel biologically relevant patterns, demonstrating the potential of our data-driven approach.
    Keywords:  cellular morphometry; computational biology; context-guided visualization; data integration; developmental biology; image analysis; lateral line primordium; morphogenesis; systems biology; zebrafish
    DOI:  https://doi.org/10.7554/eLife.55913
  41. Cell Metab. 2020 May 28. pii: S1550-4131(20)30251-5. [Epub ahead of print]
    Andres-Hernando A, Orlicky DJ, Kuwabara M, Ishimoto T, Nakagawa T, Johnson RJ, Lanaspa MA.
      Intake of fructose-containing sugars is strongly associated with metabolic syndrome. Compared with other sugars, dietary fructose is uniquely metabolized by fructokinase. However, the tissue-specific role of fructokinase in sugar-induced metabolic syndrome, and the specific roles of glucose and fructose in driving it, is not fully understood. Here, we show that in mice receiving excess fructose-glucose solutions, whole-body deletion of fructokinase, and thus full blockade of fructose metabolism, is sufficient to prevent metabolic syndrome. This protection is not only due to reduced fructose metabolism, but also due to decreased sugar intake. Furthermore, by using tissue-specific fructokinase-deficient mice, we determined that while sugar intake is controlled by intestinal fructokinase activity, metabolic syndrome is driven by fructose metabolism in the liver. Our findings show a two-pronged role for fructose metabolism in sugar-induced metabolic syndrome, one arm via the intestine that mediates sugar intake and a second arm in the liver that drives metabolic dysfunction.
    Keywords:  fructokinase; metabolic syndrome; obesity; sugar
    DOI:  https://doi.org/10.1016/j.cmet.2020.05.012
  42. Cancer Metastasis Rev. 2020 May 30.
    Singh V, Lamaze C.
      Caveolae are bulb-like invaginations made up of two essential structural proteins, caveolin-1 and cavins, which are abundantly present at the plasma membrane of vertebrate cells. Since their discovery more than 60 years ago, the function of caveolae has been mired in controversy. The last decade has seen the characterization of new caveolae components and regulators together with the discovery of additional cellular functions that have shed new light on these enigmatic structures. Early on, caveolae and/or caveolin-1 have been involved in the regulation of several parameters associated with cancer progression such as cell migration, metastasis, angiogenesis, or cell growth. These studies have revealed that caveolin-1 and more recently cavin-1 have a dual role with either a negative or a positive effect on most of these parameters. The recent discovery that caveolae can act as mechanosensors has sparked an array of new studies that have addressed the mechanobiology of caveolae in various cellular functions. This review summarizes the current knowledge on caveolae and their role in cancer development through their activity in membrane tension buffering. We propose that the role of caveolae in cancer has to be revisited through their response to the mechanical forces encountered by cancer cells during tumor mass development.
    Keywords:  Cancer; Caveolae; EHD2; Mechanosensing; Mechanotransdcution; Membrane tension
    DOI:  https://doi.org/10.1007/s10555-020-09899-2
  43. J Clin Invest. 2020 Jun 01. pii: 132876. [Epub ahead of print]130(6): 3253-3269
    Liu J, Zhang C, Wu H, Sun XX, Li Y, Huang S, Yue X, Lu SE, Shen Z, Su X, White E, Haffty BG, Hu W, Feng Z.
      Phosphoglycerate dehydrogenase (PHGDH), the first rate-limiting enzyme of serine synthesis, is frequently overexpressed in human cancer. PHGDH overexpression activates serine synthesis to promote cancer progression. Currently, PHGDH regulation in normal cells and cancer is not well understood. Parkin, an E3 ubiquitin ligase involved in Parkinson's disease, is a tumor suppressor. Parkin expression is frequently downregulated in many types of cancer, and its tumor-suppressive mechanism is poorly defined. Here, we show that PHGDH is a substrate for Parkin-mediated ubiquitination and degradation. Parkin interacted with PHGDH and ubiquitinated PHGDH at lysine 330, leading to PHGDH degradation to suppress serine synthesis. Parkin deficiency in cancer cells stabilized PHGDH and activated serine synthesis to promote cell proliferation and tumorigenesis, which was largely abolished by targeting PHGDH with RNA interference, CRISPR/Cas9 KO, or small-molecule PHGDH inhibitors. Furthermore, Parkin expression was inversely correlated with PHGDH expression in human breast cancer and lung cancer. Our results revealed PHGDH ubiquitination by Parkin as a crucial mechanism for PHGDH regulation that contributes to the tumor-suppressive function of Parkin and identified Parkin downregulation as a critical mechanism underlying PHGDH overexpression in cancer.
    Keywords:  Metabolism; Oncology; Tumor suppressors; Ubiquitin-proteosome system
    DOI:  https://doi.org/10.1172/JCI132876
  44. Aging Dis. 2020 May;11(3): 575-587
    Song HD, Kim SN, Saha A, Ahn SY, Akindehin S, Son Y, Cho YK, Kim M, Park JH, Jung YS, Lee YH.
      Aging-related adipose tissue dysfunction contributes to the progression of chronic metabolic diseases. We investigated the role of age-dependent expression of a neurotrophin, brain-derived neurotrophic factor (BDNF) in adipose tissue. Pro-BDNF expression was elevated in epididymal white adipose tissue (eWAT) with advanced age, which was associated with the reduction in sympathetic innervation. Interestingly, BDNF expression was enriched in PDGFRα+ adipocyte progenitors isolated from eWAT, with age-dependent increase in expression. In vitro pro-BDNF treatment caused apoptosis in adipocytes differentiated from C3H10T1/2 cells, and siRNA knockdown of sortilin mitigated these effects. Tamoxifen-inducible PDGFRα+ cell-specific deletion of BDNF (BDNFPdgfra KO) reduced pro-BDNF expression in eWAT, prevented age-associated declines in sympathetic innervation and mitochondrial content in eWAT, and improved insulin sensitivity. Moreover, BDNFPdgfra KO mice showed reduced expression of aging-induced inflammation and senescence markers in eWAT. Collectively, these results identified the upregulation of pro-BDNF expression in adipocyte progenitors as a feature of visceral white adipose tissue aging and suggested that inhibition of BDNF expression in adipocyte progenitors is potentially beneficial to prevent aging-related adipose tissue dysfunction.
    Keywords:  BDNF; adipocyte progenitors; adipose tissue; aging; sympathetic innervation
    DOI:  https://doi.org/10.14336/AD.2019.0810
  45. Aging (Albany NY). 2020 06 02. 12
    Long DM, Frame AK, Reardon PN, Cumming RC, Hendrix DA, Kretzschmar D, Giebultowicz JM.
      Lactate dehydrogenase (LDH) catalyzes the conversion of glycolysis-derived pyruvate to lactate. Lactate has been shown to play key roles in brain energetics and memory formation. However, lactate levels are elevated in aging and Alzheimer's disease patients, and it is not clear whether lactate plays protective or detrimental roles in these contexts. Here we show that Ldh transcript levels are elevated and cycle with diurnal rhythm in the heads of aged flies and this is associated with increased LDH protein, enzyme activity, and lactate concentrations. To understand the biological significance of increased Ldh gene expression, we genetically manipulated Ldh levels in adult neurons or glia. Overexpression of Ldh in both cell types caused a significant reduction in lifespan whereas Ldh down-regulation resulted in lifespan extension. Moreover, pan-neuronal overexpression of Ldh disrupted circadian locomotor activity rhythms and significantly increased brain neurodegeneration. In contrast, reduction of Ldh in neurons delayed age-dependent neurodegeneration. Thus, our unbiased genetic approach identified Ldh and lactate as potential modulators of aging and longevity in flies.
    Keywords:  aging; circadian rhythms; lactate; lactate dehydrogenase; lifespan; neurodegeneration
    DOI:  https://doi.org/10.18632/aging.103373
  46. Aging (Albany NY). 2020 Jun 05. 12
    Hsu WH, Shiao YJ, Chao YM, Huang YJ, Lin YL.
      Aging is a natural human process. It is uniquely individual, taking into account experiences, lifestyle habits and environmental factors. However, many disorders and syndromes, such as osteoporosis, neurodegenerative disorders, cognitive decline etc., often come with aging. The present study was designed to investigate the possible anti-aging effect of N6-(4-hydroxybenzyl)adenine riboside (T1-11), an adenosine analog isolated from Gastrodia elata, in a mouse model of aging created by D-galactose (D-gal) and the underlying mechanism, as well as explore the role of adenosine signaling in aging. T1-11 activated A2AR and suppressed D-gal- and BeSO4-induced cellular senescence in vitro. In vivo results in mice revealed that T1-11 abated D-gal-induced reactive oxygen species generation and ameliorated cognitive decline by inducing neurogenesis and lowering D-gal-caused neuron death. T1-11 could be a potent agent for postponing senility and preventing aging-related neuroinflammation and neurodegeneration.
    Keywords:  T1-11; adenosine analog; anti-neuroinflammation; neurogenesis; senescence
    DOI:  https://doi.org/10.18632/aging.103279
  47. J Cell Sci. 2020 Jun 05. pii: jcs.245589. [Epub ahead of print]
    Agarwal S, Ganesh S.
      Heat shock response (HSR) is a conserved cellular defensive response against stresses such as temperature, oxidative stress, and heavy metals. A significant group of players in HSR is the set of molecular chaperones, known as heat shock proteins (HSPs) that assist in the refolding of unfolded proteins and prevent the accumulation of damaged proteins. HSP genes are activated by the HSF1 transcription factor-a master regulator of the HSR pathway. A variety of stressors activates HSF1, but the key molecular players and the process that directly contribute to the HSF1 activation remains unclear. In this study, we show that heat shock induces perinuclear clustering of mitochondria in mammalian cells, and this clustering is essential for the activation of HSR. We also show that this perinuclear clustering of mitochondria results in the increased levels of ROS in the nucleus, leading to the activation of hypoxia-inducible factor-1α (HIF-1α). Finally, we provide evidence to suggest that HIF-1α is one of the critical regulators of HSF1 and that HIF-1α is essential for the activation of HSR during a heat shock.
    Keywords:  Chaperones; Hypoxia response; Mitochondrial transport; Oxidative stress; Stress response; Transcriptional regulation
    DOI:  https://doi.org/10.1242/jcs.245589
  48. EMBO J. 2020 Jun 03. e103812
    Sánchez-González C, Nuevo-Tapioles C, Herrero Martín JC, Pereira MP, Serrano Sanz S, Ramírez de Molina A, Cuezva JM, Formentini L.
      It is controversial whether mitochondrial dysfunction in skeletal muscle is the cause or consequence of metabolic disorders. Herein, we demonstrate that in vivo inhibition of mitochondrial ATP synthase in muscle alters whole-body lipid homeostasis. Mice with restrained mitochondrial ATP synthase activity presented intrafiber lipid droplets, dysregulation of acyl-glycerides, and higher visceral adipose tissue deposits, poising these animals to insulin resistance. This mitochondrial energy crisis increases lactate production, prevents fatty acid β-oxidation, and forces the catabolism of branched-chain amino acids (BCAA) to provide acetyl-CoA for de novo lipid synthesis. In turn, muscle accumulation of acetyl-CoA leads to acetylation-dependent inhibition of mitochondrial respiratory complex II enhancing oxidative phosphorylation dysfunction which results in augmented ROS production. By screening 702 FDA-approved drugs, we identified edaravone as a potent mitochondrial antioxidant and enhancer. Edaravone administration restored ROS and lipid homeostasis in skeletal muscle and reinstated insulin sensitivity. Our results suggest that muscular mitochondrial perturbations are causative of metabolic disorders and that edaravone is a potential treatment for these diseases.
    Keywords:  ATP synthase; Acetyl-CoA; edaravone; insulin resistance; mitochondria
    DOI:  https://doi.org/10.15252/embj.2019103812
  49. J Cell Sci. 2020 Jun 01. pii: jcs.242859. [Epub ahead of print]
    Urae S, Harita Y, Udagawa T, Ode KL, Nagahama M, Kajiho Y, Kanda S, Saito A, Ueda HR, Nangaku M, Oka A.
      Cubilin (CUBN) and amnionless (AMN), expressed in kidney and intestine, form a multiligand receptor complex called CUBAM that plays a crucial role in albumin absorption. To date, the mechanism of albumin endocytosis mediated by CUBAM remains to be elucidated. Here, we established an assay to quantitatively evaluate the albumin uptake by CUBAM using cells expressing full-length CUBN, which elucidated the crucial role of C-terminal part of CUBN and endocytosis signal motifs of AMN in albumin endocytosis. We also demonstrated that nuclear valosin-containing protein-like 2 (NVL2), an interacting protein of AMN, is involved in this process. While NVL2 was mainly localized in the nucleolus in cells without AMN expression, it was translocated to the extranuclear compartment when co-expressed with AMN. NVL2 knockdown significantly impaired the internalization of the CUBN-albumin complex in cultured cells, demonstrating an involvement of NVL2 in endocytic regulation. These findings uncover a link between membrane and nucleolar proteins which is involved in endocytic processes.
    Keywords:  Albumin; Amnionless; Cubilin; Endocytosis; Nuclear valosin-containing protein like
    DOI:  https://doi.org/10.1242/jcs.242859