Analysis of redox landscapes and dynamics in living cells and in vivo using genetically encoded fluorescent sensors

Zou, Yejun; Wang, Aoxue; Shi, M.; Chen, Xianjun; Liu, Renmei; Li, Ting; Zhang, Chenxia; Zhang, Zhuo; Zhu, Linyong; Ju, Zhenyu; Loscalzo, Joseph; Yang, Yi; Zhao, Yuzheng

doi:10.1038/s41596-018-0042-5

Cited by 80 publications

(47 citation statements)

References 60 publications

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“…The redox environment within the cell is spatially heterogeneous, which is evident from previous analyses using organelle-targeted redox sensors [20,36]. The mitochondrial intermembrane space is characterized by a higher redox potential relative to that in the cytosol and reflects mitochondrial activity involved in ROS generation [37].…”

Section: Discussionmentioning

confidence: 99%

Visualization of the Redox Status of Cytosolic Glutathione Using the Organelle- and Cytoskeleton-Targeted Redox Sensors

et al. 2020

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Glutathione is a small thiol-containing peptide that plays a central role in maintaining cellular redox homeostasis. Glutathione serves as a physiologic redox buffer by providing thiol electrons for catabolizing harmful oxidants and reversing oxidative effects on biomolecules. Recent evidence suggests that the balance of reduced and oxidized glutathione (GSH/GSSG) defines the redox states of Cys residues in proteins and fine-tunes their stabilities and functions. To elucidate the redox balance of cellular glutathione at subcellular resolution, a number of redox-sensitive green fluorescent protein (roGFP) variants have been developed. In this study, we constructed and functionally validated organelle- and cytoskeleton-targeted roGFP and elucidated the redox status of the cytosolic glutathione at a subcellular resolution. These new redox sensors firmly established a highly reduced redox equilibrium of cytosolic glutathione, wherein significant deviation was observed among cells. By targeting the sensor to the cytosolic and lumen sides of the Golgi membrane, we identified a prominent redox gradient across the biological membrane at the Golgi body. The results demonstrated that organelle- and cytoskeleton-targeted sensors enable the assessment of glutathione oxidation near the cytosolic surfaces of different organelle membranes.

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Section: Discussionmentioning

confidence: 99%

Visualization of the Redox Status of Cytosolic Glutathione Using the Organelle- and Cytoskeleton-Targeted Redox Sensors

et al. 2020

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“…Meanwhile, it can be adapted to high-throughput chemical screening of potential compounds targeting cellular metabolism in a variety of analytical platforms, including microplate readers, flow cytometry, fluorescence microscopy and high-content imaging systems. 97,654,[667][668][669][670][671][672][673][674] 31 P-magnetic resonance spectroscopy ( 31 P-MRS) methods 31 P-MRS based NAD + assay is a noninvasive method that could quantitatively measure intracellular NAD + contents and redox state in animal and human tissues, such as brains. It provides new approach to investigate intracellular NAD + redox state and metabolism in the human tissues with the potential for translation to human application.…”

Section: Biochemical Analysismentioning

confidence: 99%

NAD+ metabolism: pathophysiologic mechanisms and therapeutic potential

Xie

Zhang

Gao

et al. 2020

Sig Transduct Target Ther

520

383

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Nicotinamide adenine dinucleotide (NAD+) and its metabolites function as critical regulators to maintain physiologic processes, enabling the plastic cells to adapt to environmental changes including nutrient perturbation, genotoxic factors, circadian disorder, infection, inflammation and xenobiotics. These effects are mainly achieved by the driving effect of NAD+ on metabolic pathways as enzyme cofactors transferring hydrogen in oxidation-reduction reactions. Besides, multiple NAD+-dependent enzymes are involved in physiology either by post-synthesis chemical modification of DNA, RNA and proteins, or releasing second messenger cyclic ADP-ribose (cADPR) and NAADP+. Prolonged disequilibrium of NAD+ metabolism disturbs the physiological functions, resulting in diseases including metabolic diseases, cancer, aging and neurodegeneration disorder. In this review, we summarize recent advances in our understanding of the molecular mechanisms of NAD+-regulated physiological responses to stresses, the contribution of NAD+ deficiency to various diseases via manipulating cellular communication networks and the potential new avenues for therapeutic intervention.

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“…Recently, Yang and colleagues showed that a highly responsive genetically encoded intracellular sensor, SoNar, can precisely report the changes of cytosolic NAD + and NADH redox states in real time, which is useful for the evaluation of glycolysis and mitochondrial respiration activities in cells . In the cytosol glycolysis produced NADH, which may be shuttled to mitochondria and oxidized, or recycled to NAD + by lactate dehydrogenase and produce lactate (Zou et al, 2018). The flux balance of these three pathways determines the cytosolic NADH/NAD + ratio.…”

Section: Introductionmentioning

confidence: 99%

Metabolic Imaging Reveals a Unique Preference of Symmetric Cell Division and Homing of Leukemia-Initiating Cells in an Endosteal Niche

Hao

Chen

et al. 2019

Cell Metabolism

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Graphical Abstract Highlights d SoNar serves as a unique tool for in vitro/in vivo studies of stem cell metabolisms d SoNar-high cells prefer glycolysis and are enriched for leukemia-initiating cells d Leukemia-initiating cells home to endosteal niches and prefer symmetric divisions d PDK2 sustains metabolic and leukemogenic activities of leukemia-initiating cells In Brief Hao et al. reveal that a genetically encoded metabolic sensor, SoNar, precisely monitors the metabolic status of leukemia-initiating cells (LICs) both in vitro and in vivo. LICs prefer homing to endosteal niches and symmetric divisions to maintain their leukemogenic activities. PDK2 fine-tunes the glycolysis and cell-fate determinations of LICs. Hao et al., 2019, Cell Metabolism 29, 950-965 April 2, 2019 ª 2018 Elsevier Inc. SUMMARYThe metabolic properties of leukemia-initiating cells (LICs) in distinct bone marrow niches and their relationships to cell-fate determinations remain largely unknown. Using a metabolic imaging system with a highly responsive genetically encoded metabolic sensor, SoNar, we reveal that SoNar-high cells are more glycolytic, enriched for higher LIC frequency, and develop leukemia much faster than SoNar-low counterparts in an MLL-AF9-induced murine acute myeloid leukemia model. SoNar-high cells mainly home to and locate in the hypoxic endosteal niche and maintain their activities through efficient symmetric division. SoNar can indicate the dynamics of metabolic changes of LICs in the endosteal niche. SoNar-high human leukemia cells or primary samples have enhanced clonogenic capacities in vitro or leukemogenesis in vivo. PDK2 fine-tunes glycolysis, homing, and symmetric division of LICs. These findings provide a unique angle for the study of metabolisms in stem cells, and may lead to development of novel strategies for cancer treatment.

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Analysis of redox landscapes and dynamics in living cells and in vivo using genetically encoded fluorescent sensors

Cited by 80 publications

References 60 publications

Visualization of the Redox Status of Cytosolic Glutathione Using the Organelle- and Cytoskeleton-Targeted Redox Sensors

Visualization of the Redox Status of Cytosolic Glutathione Using the Organelle- and Cytoskeleton-Targeted Redox Sensors

NAD+ metabolism: pathophysiologic mechanisms and therapeutic potential

Metabolic Imaging Reveals a Unique Preference of Symmetric Cell Division and Homing of Leukemia-Initiating Cells in an Endosteal Niche

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