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      Xylose Metabolism and the Effect of Oxidative Stress on Lipid and Carotenoid Production in Rhodotorula toruloides: Insights for Future Biorefinery


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          The use of cell factories to convert sugars from lignocellulosic biomass into chemicals in which oleochemicals and food additives, such as carotenoids, is essential for the shift toward sustainable processes. Rhodotorula toruloides is a yeast that naturally metabolises a wide range of substrates, including lignocellulosic hydrolysates, and converts them into lipids and carotenoids. In this study, xylose, the main component of hemicellulose, was used as the sole substrate for R. toruloides, and a detailed physiology characterisation combined with absolute proteomics and genome-scale metabolic models was carried out to understand the regulation of lipid and carotenoid production. To improve these productions, oxidative stress was induced by hydrogen peroxide and light irradiation and further enhanced by adaptive laboratory evolution. Based on the online measurements of growth and CO 2 excretion, three distinct growth phases were identified during batch cultivations. Majority of the intracellular flux estimations showed similar trends with the measured protein levels and demonstrated improved NADPH regeneration, phosphoketolase activity and reduced β-oxidation, correlating with increasing lipid yields. Light irradiation resulted in 70% higher carotenoid and 40% higher lipid content compared to the optimal growth conditions. The presence of hydrogen peroxide did not affect the carotenoid production but culminated in the highest lipid content of 0.65 g/g DCW. The adapted strain showed improved fitness and 2.3-fold higher carotenoid content than the parental strain. This work presents a holistic view of xylose conversion into microbial oil and carotenoids by R. toruloides, in a process toward renewable and cost-effective production of these molecules.

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          Controlling the False Discovery Rate: A Practical and Powerful Approach to Multiple Testing

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            Emergence of robust growth laws from optimal regulation of ribosome synthesis

            Bacteria must constantly adapt their growth to changes in nutrient availability; yet despite large-scale changes in protein expression associated with sensing, adaptation, and processing different environmental nutrients, simple growth laws connect the ribosome abundance and the growth rate. Here, we investigate the origin of these growth laws by analyzing the features of ribosomal regulation that coordinate proteome-wide expression changes with cell growth in a variety of nutrient conditions in the model organism Escherichia coli. We identify supply-driven feedforward activation of ribosomal protein synthesis as the key regulatory motif maximizing amino acid flux, and autonomously guiding a cell to achieve optimal growth in different environments. The growth laws emerge naturally from the robust regulatory strategy underlying growth rate control, irrespective of the details of the molecular implementation. The study highlights the interplay between phenomenological modeling and molecular mechanisms in uncovering fundamental operating constraints, with implications for endogenous and synthetic design of microorganisms.
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              Acute Activation of Oxidative Pentose Phosphate Pathway as First-Line Response to Oxidative Stress in Human Skin Cells.

              Integrity of human skin is endangered by exposure to UV irradiation and chemical stressors, which can provoke a toxic production of reactive oxygen species (ROS) and oxidative damage. Since oxidation of proteins and metabolites occurs virtually instantaneously, immediate cellular countermeasures are pivotal to mitigate the negative implications of acute oxidative stress. We investigated the short-term metabolic response in human skin fibroblasts and keratinocytes to H2O2 and UV exposure. In time-resolved metabolomics experiments, we observed that within seconds after stress induction, glucose catabolism is routed to the oxidative pentose phosphate pathway (PPP) and nucleotide synthesis independent of previously postulated blocks in glycolysis (i.e., of GAPDH or PKM2). Through ultra-short (13)C labeling experiments, we provide evidence for multiple cycling of carbon backbones in the oxidative PPP, potentially maximizing NADPH reduction. The identified metabolic rerouting in oxidative and non-oxidative PPP has important physiological roles in stabilization of the redox balance and ROS clearance.

                Author and article information

                Front Bioeng Biotechnol
                Front Bioeng Biotechnol
                Front. Bioeng. Biotechnol.
                Frontiers in Bioengineering and Biotechnology
                Frontiers Media S.A.
                19 August 2020
                : 8
                : 1008
                [1] 1Institute of Technology, University of Tartu , Tartu, Estonia
                [2] 2Department of Materials and Bioprocess Engineering, School of Chemical Engineering, University of Campinas , Campinas, Brazil
                Author notes

                Edited by: Chris Petzold, Lawrence Berkeley National Laboratory, United States

                Reviewed by: Zongbao K. Zhao, Dalian Institute of Chemical Physics (CAS), China; John Gladden, Sandia National Laboratories (SNL), United States

                *Correspondence: Petri-Jaan Lahtvee, petri.lahtvee@ 123456ut.ee

                These authors have contributed equally to this work and share first authorship

                This article was submitted to Synthetic Biology, a section of the journal Frontiers in Bioengineering and Biotechnology

                Copyright © 2020 Pinheiro, Bonturi, Belouah, Miranda and Lahtvee.

                This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

                : 02 June 2020
                : 31 July 2020
                Page count
                Figures: 5, Tables: 1, Equations: 0, References: 66, Pages: 15, Words: 0
                Funded by: Horizon 2020 10.13039/501100007601
                Funded by: Eesti Teadusagentuur 10.13039/501100002301
                Funded by: Fundação de Amparo à Pesquisa do Estado de São Paulo 10.13039/501100001807
                Bioengineering and Biotechnology
                Original Research

                microbial oil,carotenoids,rhodotorula toruloides,genome-scale modelling,xylose,biorefinery,absolute proteomics


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