bims-glucam 2020-09-13 papers

Cell Metab. 2020 Aug 31. pii: S1550-4131(20)30424-1. [Epub ahead of print]

Glutamine Metabolism Controls Stem Cell Fate Reversibility and Long-Term Maintenance in the Hair Follicle.

Christine S Kim, Xiaolei Ding, Kira Allmeroth, Leah C Biggs, Olivia I Kolenc, Nina L'Hoest, Carlos Andrés Chacón-Martínez, Christian Edlich-Muth, Patrick Giavalisco, Kyle P Quinn, Martin S Denzel, Sabine A Eming, Sara A Wickström.

Stem cells reside in specialized niches that are critical for their function. Upon activation, hair follicle stem cells (HFSCs) exit their niche to generate the outer root sheath (ORS), but a subset of ORS progeny returns to the niche to resume an SC state. Mechanisms of this fate reversibility are unclear. We show that the ability of ORS cells to return to the SC state requires suppression of a metabolic switch from glycolysis to oxidative phosphorylation and glutamine metabolism that occurs during early HFSC lineage progression. HFSC fate reversibility and glutamine metabolism are regulated by the mammalian target of rapamycin complex 2 (mTORC2)-Akt signaling axis within the niche. Deletion of mTORC2 results in a failure to re-establish the HFSC niche, defective hair follicle regeneration, and compromised long-term maintenance of HFSCs. These findings highlight the importance of spatiotemporal control of SC metabolic states in organ homeostasis.

Keywords: Akt; Hif1; cell fate; glutamine; hair follicle; hypoxia; mTOR; mTORC2; metabolism; stem cell

DOI: https://doi.org/10.1016/j.cmet.2020.08.011

Cancer Res. 2020 Sep 09. pii: canres.0600.2020. [Epub ahead of print]

5-fluorouracil enhances the anti-tumor activity of the glutaminase inhibitor CB-839 against PIK3CA-mutant colorectal cancers.

PIK3CA encodes the p110α catalytic subunit of PI3 kinase and is frequently mutated in human cancers, including ~30% of colorectal cancer (CRC). Oncogenic mutations in PIK3CA render CRCs more dependent on glutamine. Here we report that the glutaminase inhibitor CB-839 preferentially inhibits xenograft growth of PIK3CA-mutant, but not wild-type (WT), CRCs. Moreover, the combination of CB-839 and 5-fluorouracil (5-FU) induces PIK3CA-mutant tumor regression in xenograft models. CB-839 treatment increased reactive oxygen species and caused nuclear translocation of Nrf2, which in turn up-regulated mRNA expression of uridine phosphorylase 1 (UPP1). UPP1 facilitated the conversion of 5-FU to its active compound, thereby enhancing the inhibition of thymidylate synthase. Consistently, knockout of UPP1 abrogated the tumor inhibitory effect of combined CB-839 and 5-FU administration. A phase I clinical trial showed that the combination of CB-839 and capecitabine, a prodrug of 5-FU, was well tolerated at biologically-active doses. Although not designed to test efficacy, an exploratory analysis of the phase I data showed a trend that PIK3CA-mutant CRC patients might derive greater benefit from this treatment strategy as compared to PIK3CA WT CRC patients. These results effectively demonstrate that targeting glutamine metabolism may be an effective approach for treating patients with PIK3CA-mutant CRCs and warrants further clinical evaluation.

DOI: https://doi.org/10.1158/0008-5472.CAN-20-0600

PLoS One. 2020 ;15(9): e0237981

Cancer proteome and metabolite changes linked to SHMT2.

Serine hydroxymethyltransferase 2 (SHMT2) converts serine plus tetrahydrofolate (THF) into glycine plus methylene-THF and is upregulated at the protein level in lung and other cancers. In order to better understand the role of SHMT2 in cancer a model system of HeLa cells engineered for inducible over-expression or knock-down of SHMT2 was characterized for cell proliferation and changes in metabolites and proteome as a function of SHMT2. Ectopic over-expression of SHMT2 increased cell proliferation in vitro and tumor growth in vivo. Knockdown of SHMT2 expression in vitro caused a state of glycine auxotrophy and accumulation of phosphoribosylaminoimidazolecarboxamide (AICAR), an intermediate of folate/1-carbon-pathway-dependent de novo purine nucleotide synthesis. Decreased glycine in the HeLa cell-based xenograft tumors with knocked down SHMT2 was potentiated by administration of the anti-hyperglycinemia agent benzoate. However, tumor growth was not affected by SHMT2 knockdown with or without benzoate treatment. Benzoate inhibited cell proliferation in vitro, but this was independent of SHMT2 modulation. The abundance of proteins of mitochondrial respiration complexes 1 and 3 was inversely correlated with SHMT2 levels. Proximity biotinylation in vivo (BioID) identified 48 mostly mitochondrial proteins associated with SHMT2 including the mitochondrial enzymes Acyl-CoA thioesterase (ACOT2) and glutamate dehydrogenase (GLUD1) along with more than 20 proteins from mitochondrial respiration complexes 1 and 3. These data provide insights into possible mechanisms through which elevated SHMT2 in cancers may be linked to changes in metabolism and mitochondrial function.

DOI: https://doi.org/10.1371/journal.pone.0237981

iScience. 2020 Aug 20. pii: S2589-0042(20)30671-4. [Epub ahead of print]23(9): 101479

Kinetic Trans-omic Analysis Reveals Key Regulatory Mechanisms for Insulin-Regulated Glucose Metabolism in Adipocytes.

Satoshi Ohno, Lake-Ee Quek, James R Krycer, Katsuyuki Yugi, Akiyoshi Hirayama, Satsuki Ikeda, Futaba Shoji, Kumi Suzuki, Tomoyoshi Soga, David E James, Shinya Kuroda.

Insulin regulates glucose metabolism through thousands of regulatory mechanisms; however, which regulatory mechanisms are keys to control glucose metabolism remains unknown. Here, we performed kinetic trans-omic analysis by integrating isotope-tracing glucose flux and phosphoproteomic data from insulin-stimulated adipocytes and built a kinetic mathematical model to identify key allosteric regulatory and phosphorylation events for enzymes. We identified nine reactions regulated by allosteric effectors and one by enzyme phosphorylation and determined the regulatory mechanisms for three of these reactions. Insulin stimulated glycolysis by promoting Glut4 activity by enhancing phosphorylation of AS160 at S595, stimulated fatty acid synthesis by promoting Acly activity through allosteric activation by glucose 6-phosphate or fructose 6-phosphate, and stimulated glutamate synthesis by alleviating allosteric inhibition of Gls by glutamate. Most of glycolytic reactions were regulated by amounts of substrates and products. Thus, phosphorylation or allosteric modulator-based regulation of only a few key enzymes was sufficient to change insulin-induced metabolism.

Keywords: Biological Sciences; Mathematical Biosciences; Metabolic Flux Analyisis; Metabolomics; Omics; Proteomics; Systems Biology

DOI: https://doi.org/10.1016/j.isci.2020.101479

Free Radic Biol Med. 2020 Sep 05. pii: S0891-5849(20)31233-8. [Epub ahead of print]

S-nitrosocysteine and glutathione depletion synergize to induce cell death in human tumor cells: insights into the redox and cytotoxic mechanisms.

Alaa Knany, Rotem Engelman, Hiba Abu Hariri, Shyam Biswal, Haguy Wolfenson, Moran Benhar.

Nitric oxide (NO)-dependent signaling and cytotoxic effects are mediated in part via protein S-nitrosylation. The magnitude and duration of S-nitrosylation are governed by the two main thiol reducing systems, the glutathione (GSH) and thioredoxin (Trx) antioxidant systems. In recent years, approaches have been developed to harness the cytotoxic potential of NO/nitrosylation to inhibit tumor cell growth. However, progress in this area has been hindered by insufficient understanding of the balance and interplay between cellular nitrosylation, other oxidative processes and the GSH/Trx systems. In addition, the mechanistic relationship between thiol redox imbalance and cancer cell death is not fully understood. Herein, we explored the redox and cellular effects induced by the S-nitrosylating agent, S-nitrosocysteine (CysNO), in GSH-sufficient and -deficient human tumor cells. We used L-buthionine-sulfoximine (BSO) to induce GSH deficiency, and employed redox, biochemical and cellular assays to interrogate molecular mechanisms. We found that, under GSH-sufficient conditions, a CysNO challenge (100-500 μM) results in a marked yet reversible increase in protein S-nitrosylation in the absence of appreciable S-oxidation. In contrast, under GSH-deficient conditions, CysNO induces elevated and sustained levels of both S-nitrosylation and S-oxidation. Experiments in various cancer cell lines showed that administration of CysNO or BSO alone commonly induce minimal cytotoxicity whereas BSO/CysNO combination therapy leads to extensive cell death. Studies in HeLa cancer cells revealed that treatment with BSO/CysNO results in dual inhibition of the GSH and Trx systems, thereby amplifying redox stress and causing cellular dysfunction. In particular, BSO/CysNO induced rapid oxidation and collapse of the actin cytoskeletal network, followed by loss of mitochondrial function, leading to profound and irreversible decrease in ATP levels. Further observations indicated that BSO/CysNO-induced cell death occurs via a caspase-independent mechanism that involves multiple stress-induced pathways. The present findings provide new insights into the relationship between cellular nitrosylation/oxidation, thiol antioxidant defenses and cell death. These results may aid future efforts to develop NO/redox-based anticancer approaches.

Keywords: Thiols; cancer; cell death; glutathione; nitrosylation; oxidation; thioredoxin

DOI: https://doi.org/10.1016/j.freeradbiomed.2020.08.026

Dev Cell. 2020 Aug 31. pii: S1534-5807(20)30666-3. [Epub ahead of print]

Selective Lysosome Membrane Turnover Is Induced by Nutrient Starvation.

Chan Lee, Lilian Lamech, Eleanor Johns, Michael Overholtzer.

Lysosome function is essential for cellular homeostasis, but quality-control mechanisms that maintain healthy lysosomes remain poorly characterized. Here, we developed a method to measure lysosome turnover and use this to identify a selective mechanism of membrane degradation that involves lipidation of the autophagy protein LC3 onto lysosomal membranes and the formation of intraluminal vesicles through microautophagy. This mechanism is induced in response to metabolic stress resulting from glucose starvation or by treatment with pharmacological agents that induce osmotic stress on lysosomes. Cells lacking ATG5, an essential component of the LC3 lipidation machinery, show reduced ability to regulate lysosome size and degradative capacity in response to activation of this mechanism. These findings identify a selective mechanism of lysosome membrane turnover that is induced by stress and uncover a function for LC3 lipidation in regulating lysosome size and activity through microautophagy.

Keywords: ATG5; LAP; LC3; ammonium; autophagy; glucose; glutamine; lysosome; metabolism; microautophagy

DOI: https://doi.org/10.1016/j.devcel.2020.08.008