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Genetically encoded fluorescent sensors reveal dynamic regulation of NADPH metabolism

Nature methods, 2017-07, Vol.14 (7), p.720-728 [Peer Reviewed Journal]

COPYRIGHT 2017 Nature Publishing Group ;Copyright Nature Publishing Group Jul 2017 ;ISSN: 1548-7091 ;EISSN: 1548-7105 ;DOI: 10.1038/nmeth.4306 ;PMID: 28581494

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Title:
Genetically encoded fluorescent sensors reveal dynamic regulation of NADPH metabolism
Author: Tao, Rongkun ; Zhao, Yuzheng ; Chu, Huanyu ; Wang, Aoxue ; Zhu, Jiahuan ; Chen, Xianjun ; Zou, Yejun ; Shi, Mei ; Liu, Renmei ; Su, Ni ; Du, Jiulin ; Zhou, Hai-Meng ; Zhu, Linyong ; Qian, Xuhong ; Liu, Haiyan ; Loscalzo, Joseph ; Yang, Yi
Subjects: AMP-Activated Protein Kinases - genetics ; AMP-Activated Protein Kinases - metabolism ; Animals ; Biosensors ; Cell activation ; Cell Survival ; Cells (biology) ; Coding ; Dehydrogenase ; Dynamic range ; Fluctuations ; Fluorescence ; Fluorescent indicators ; Fluorescent proteins ; Gene Expression Regulation - physiology ; Genetic code ; Genetics ; Glucose ; Glucose 6 phosphate dehydrogenase ; Glucose-6-phosphate ; Glucosephosphate dehydrogenase ; Homeostasis ; Humans ; Indicators ; Luminescent Proteins - chemistry ; Luminescent Proteins - genetics ; Luminescent Proteins - metabolism ; Macrophages ; Mammalian cells ; Mammals ; Metabolism ; Mice ; Mitochondria ; Models, Molecular ; NADP - metabolism ; NADPH-diaphorase ; Nicotinamide ; Nicotinamide adenine dinucleotide ; Nicotinamide adenine dinucleotide phosphate ; Oxidative Stress ; Phosphates ; Pools ; Properties ; Protein Binding ; Protein Conformation ; Protein Domains ; Protein Engineering ; Sensors
Is Part Of: Nature methods, 2017-07, Vol.14 (7), p.720-728
Description: Reduced nicotinamide adenine dinucleotide phosphate (NADPH) is essential for biosynthetic reactions and antioxidant functions; however, detection of NADPH metabolism in living cells remains technically challenging. We develop and characterize ratiometric, pH-resistant, genetically encoded fluorescent indicators for NADPH (iNap sensors) with various affinities and wide dynamic range. iNap sensors enabled quantification of cytosolic and mitochondrial NADPH pools that are controlled by cytosolic NAD kinase levels and revealed cellular NADPH dynamics under oxidative stress depending on glucose availability. We found that mammalian cells have a strong tendency to maintain physiological NADPH homeostasis, which is regulated by glucose-6-phosphate dehydrogenase and AMP kinase. Moreover, using the iNap sensors we monitor NADPH fluctuations during the activation of macrophage cells or wound response in vivo. These data demonstrate that the iNap sensors will be valuable tools for monitoring NADPH dynamics in live cells and gaining new insights into cell metabolism.
Publisher: United States: Nature Publishing Group
Language: English
Identifier: ISSN: 1548-7091
EISSN: 1548-7105
DOI: 10.1038/nmeth.4306
PMID: 28581494
Source: MEDLINE
ProQuest Central

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Genetically encoded fluorescent sensors reveal dynamic regulation of NADPH metabolism

COPYRIGHT 2017 Nature Publishing Group ;Copyright Nature Publishing Group Jul 2017 ;ISSN: 1548-7091 ;EISSN: 1548-7105 ;DOI: 10.1038/nmeth.4306 ;PMID: 28581494

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