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CQ and BAF do not affect ER area or phospholipid levels in IDHwt glioma cells.

CQ and BAF restore phospholipid levels and inhibit clonogenicity of IDHmut glioma cells.

2-HG induces a reduction in ER area in IDHmut glioma cells.

Tumor metabolism is reprogrammed to meet the demands of proliferating cancer cells. In particular, cancer cells upregulate synthesis of the membrane phospholipids phosphatidylcholine (PtdCho) and phosphatidylethanolamine (PtdE) in order to allow for rapid membrane turnover. Nonetheless, we show here that, in mutant isocitrate dehydrogenase 1 (IDHmut) gliomas, which produce the oncometabolite 2-hydroxyglutarate (2-HG), PtdCho and PtdE biosynthesis is downregulated and results in lower levels of both phospholipids when compared with wild-type IDH1 cells. 2-HG inhibited collagen-4-prolyl hydroxylase activity, leading to accumulation of misfolded procollagen-IV in the endoplasmic reticulum (ER) of both genetically engineered and patient-derived IDHmut glioma models. The resulting ER stress triggered increased expression of FAM134b, which mediated autophagic degradation of the ER (ER-phagy) and a reduction in the ER area. Because the ER is the site of phospholipid synthesis, ER-phagy led to reduced PtdCho and PtdE biosynthesis. Inhibition of ER-phagy via pharmacological or molecular approaches restored phospholipid biosynthesis in IDHmut glioma cells, triggered apoptotic cell death, inhibited tumor growth, and prolonged the survival of orthotopic IDHmut glioma-bearing mice, pointing to a potential therapeutic opportunity. Glioma patient biopsies also exhibited increased ER-phagy and downregulation of PtdCho and PtdE levels in IDHmut samples compared with wild-type, clinically validating our observations. Collectively, this study provides detailed and clinically relevant insights into the functional link between oncometabolite-driven ER-phagy and phospholipid biosynthesis in IDHmut gliomas.Significance: Downregulation of phospholipid biosynthesis via ER-phagy is essential for proliferation and clonogenicity of mutant IDH1 gliomas, a finding with immediate therapeutic implications. Cancer Res; 78(9); 2290–304. ©2018 AACR.

2-HG inhibits PtdCho and PtdE biosynthesis.

Phospholipid biosynthesis is down-regulated in IDHmut glioma cells.

Silencing FAM134b, Atg5 or Atg7 restores phospholipid biosynthesis in IDHmut glioma cells.

2-HG induces ER-phagy in IDHmut glioma cells.

ER-phagy is linked to phospholipid biosynthesis in BT54 neurospheres (A-C), orthotopic IDHmut tumor xenografts (D-J) and IDHmut patient biopsies (K-Q).

Constraint-based modelling was applied to provide a mechanistic understanding of the possible metabolic origins of the ‘Kok effect’ – the change in quantum yield of net photosynthesis at low light intensity. The well-known change in quantum yield near the light-compensation point (LCP) was predicted as an emergent behaviour from a purely stoichiometric model. From our modelling results, we discovered another subtle change in quantum yield at a light intensity lower than the LCP. Our model predicted a series of changes in metabolic flux modes in central carbon metabolism associated with the changes in quantum yields. We demonstrated that the Kok effect can be explained by changes in metabolic flux modes between catabolism and photorespiration. Changes in RuBisCO carboxylation to oxygenation ratio resulted in a change in quantum yield at light intensities above the LCP, but not below the LCP, indicating the role of photorespiration in producing the Kok effect. Cellular energy demand was predicted to have no impact on the quantum yield. Our model showed that the Kok method vastly overestimates day respiration – the CO2 released by non-photorespiratory processes in illuminated leaves. The theoretical maximum quantum yield at low light intensity was higher than typical measured values, suggesting that leaf metabolism at low light may not be regulated to optimise for energetic efficiency. Our model predictions gave insights into the set of energetically optimal changes in flux modes in low light as light intensity increases from darkness.One sentence summaryThe Kok effect can be explained by the changes in flux modes between catabolism and photorespiration.

Tumor metabolism is reprogrammed to meet the demands of proliferating cancer cells. In particular, cancer cells upregulate synthesis of the membrane phospholipids phosphatidylcholine (PtdCho) and phosphatidylethanolamine (PtdE) in order to allow for rapid membrane turnover. Nonetheless, we show here that, in mutant isocitrate dehydrogenase 1 (IDHmut) gliomas, which produce the oncometabolite 2-hydroxyglutarate (2-HG), PtdCho and PtdE biosynthesis is downregulated and results in lower levels of both phospholipids when compared with wild-type IDH1 cells. 2-HG inhibited collagen-4-prolyl hydroxylase activity, leading to accumulation of misfolded procollagen-IV in the endoplasmic reticulum (ER) of both genetically engineered and patient-derived IDHmut glioma models. The resulting ER stress triggered increased expression of FAM134b, which mediated autophagic degradation of the ER (ER-phagy) and a reduction in the ER area. Because the ER is the site of phospholipid synthesis, ER-phagy led to reduced PtdCho and PtdE biosynthesis. Inhibition of ER-phagy via pharmacological or molecular approaches restored phospholipid biosynthesis in IDHmut glioma cells, triggered apoptotic cell death, inhibited tumor growth, and prolonged the survival of orthotopic IDHmut glioma-bearing mice, pointing to a potential therapeutic opportunity. Glioma patient biopsies also exhibited increased ER-phagy and downregulation of PtdCho and PtdE levels in IDHmut samples compared with wild-type, clinically validating our observations. Collectively, this study provides detailed and clinically relevant insights into the functional link between oncometabolite-driven ER-phagy and phospholipid biosynthesis in IDHmut gliomas. Significance: Downregulation of phospholipid biosynthesis via ER-phagy is essential for proliferation and clonogenicity of mutant IDH1 gliomas, a finding with immediate therapeutic implications. Cancer Res; 78(9); 2290–304. ©2018 AACR.

Messenger RNA (mRNA) plays a critical role in cellular growth and development. However, there have been limited methods available to visualize endogenous mRNA in living cells with ease. We have designed RNA-based fluorescence “turn-on” probes that target mRNA by fusing an unstable form of Spinach with target-complementary sequences. These probes have been demonstrated to be selective, stable, and capable of targeting various mRNAs for live E. coli imaging.

Photoactivatable fluorescent proteins (PA-FPs) are widely used in live single-molecule super-resolution imaging but emit substantially fewer photons than organic dyes do. Here, we show that in heavy water (D2O) instead of H2O, common PA-FPs emit 26 %–54 % more photons, effectively improving the localization precision in super-resolution imaging.

The study of macrocycles has crossed many traditional disciplines such as chemistry, physics, biology, medicine and engineering with many research areas concentrating on specific and selective molecular recognition, self-organisation and its already demonstrated and other promising applications. Compared to traditional strategies to synthesize macrocycles with widely ranging structures using such as templated cyclization or dynamic covalent bond formation, one-pot H-bonding-assisted macrocyclization has been shown to provide a simple, fast and cost-efficient method to synthesize shape-persistent H-bonded macrocycles of varying types containing an internal cavity of as large as 2.9 nm in diameter. This review will summarize the recent works on such “greener” syntheses of H-bonded macrocycles that help to create a whole new dimension of research and to offer a new bottom-up strategy for constructing functional architectures and materials.

Described in this study is a conceptually new class of five-fold-symmetric cavity-containing planar pentameric macrocycles with their interior decorated by five convergently aligned, properly spaced carbonyl oxygen atoms. These cation-binding oxygens enclose a hydrophilic lumen of 2.85 Å in radius and thus display high-affinity binding toward alkali metal cations, and possibly many other cations, too. Arising from their high-affinity recognition of metal ions, these planar macrocycles form cation- or ion-pair-induced one-dimensional columnar aggregates, and subsequently fascinating fibrillation results.

Introduction of a continuous hydrogen-bonding network suppressed the conformational flexibility of an oligomeric backbone. Cyclization of a rigidified, suitably sized oligomer led to a circular aromatic pentamer. Its crystal structure determined by X-ray crystallography reveals a pseudo five-fold symmetric planarity in the solid state, which is quite unusual among all the previously described shape-persistent macrocycles and synthetic foldamers with biased conformations enforced by noncovalent forces.

The study of macrocycles has crossed many traditional disciplines such as chemistry, physics, biology, medicine and engineering with many research areas concentrating on specific and selective molecular recognition, self-organisation and its already demonstrated and other promising applications. Compared to traditional strategies to synthesize macrocycles with widely ranging structures using such as templated cyclization or dynamic covalent bond formation, one-pot H-bonding-assisted macrocyclization has been shown to provide a simple, fast and cost-efficient method to synthesize shape-persistent H-bonded macrocycles of varying types containing an internal cavity of as large as 2.9 nm in diameter. This review will summarize the recent works on such “greener” syntheses of H-bonded macrocycles that help to create a whole new dimension of research and to offer a new bottom-up strategy for constructing functional architectures and materials.

Biomolecules such as α-helices are exclusively right-handed without contamination by their left-handed counterparts. Within the abiotic world, chiral or external perturbations have to be applied to the abiotic helical foldamers or polymers to control and bias their helical screw sense. Otherwise, it is not possible to separate racemic helical foldamers composed of achiral building blocks. By designing two complementary “sticky” groups, incorporating them into the ends of helical pentamers and taking advantage of the energetically more favored full overlap, which involves helical backbones, we succeeded in demonstrating, for the first time, a spontaneous resolution of racemic helices into their enantiopure single-handed helical forms via chiral crystallization without the use of chiral auxiliary or external stimuli.

Water-binding foldamers have been rarely studied. By orienting both H-bond donors and acceptors toward their interior, two pyridine-derived crescent-shaped folding oligoamides were found to be capable of trapping both conventional and unconventional water dimer clusters in their cavity (∼2.5 Å radius). In the unconventional water dimer cluster, the two water molecules stay in contact via an unusual H-H interaction (2.25 Å) rather than the typical H-bond.

Aided by high level ab initio computational modeling, we successfully designed and experimentally proved a new set of backbone-rigidified pyridine-based folding oligoamides that require approximately four repeating units to form a helical turn.

In response to binding to amine and ammonium guests of varying types, a pyridine-based folding oligomer displays fingerprint regions in its (1)H NMR spectra that allow for the easy identification and classification of the bound guests.

1D helical channels can be created by "sticky" end-mediated formation of 1D helical stacks composed of short cavity-containing helices and are able to host 1D chains of dichloromethane molecules containing extraordinarily short intermolecular Cl···Cl contacts of 2.524 Å in length that computationally destabilize the host-guest complex by 14.94 kcal mol(-1).

Quantitative super-resolution imaging is important for understanding and determining the exact stoichiometry of molecular complexes or oligomerization states of proteins within a cell. This has been done previously using fluorescence proteins, however these proteins require extensive computational correction due to their blinking. Organic dyes are commonly used for super-resolution imaging because of their high photon numbers, unfortunately blinking corrections for these dyes are very difficult. Here, we develop a technique for quantitative super resolution microscopy that allows us to count the absolute number of molecules using organic “caged” dyes, which do not blink. We calibrate our system and find that caged-fluorescein and caged-carborhodamine, when conjugated with SNAP-tag labeling, allow for quantitative counting of targets. We apply this technique to the characterization of the oligomerization states of chemokine receptors in mammalian cells.