TY - JOUR
T1 - Circadian oscillations of NADH redox state using a heterologous metabolic sensor in mammalian cells
AU - Huang, Guocun
AU - Zhang, Yunfeng
AU - Shan, Yongli
AU - Yang, Shuzhang
AU - Chelliah, Yogarany
AU - Wang, Han
AU - Takahashi, Joseph S.
N1 - Publisher Copyright:
© 2016 by The American Society for Biochemistry and Molecular Biology, Inc.
PY - 2016/11/11
Y1 - 2016/11/11
N2 - It is known that there are mechanistic links between circadian clocks and metabolic cycles. Reduced nicotinamide adenine dinucleotide (NADH) is a key metabolic cofactor in all living cells; however, it is not known whether levels of NADH oscillate or not. Here we employed REX, a bacterial NADH-binding protein, fused to the VP16 activator to convert intracellular endogenous redox balance into transcriptional readouts by a reporter gene in mammalian cells. EMSA results show that the DNA binding activity of both T- and S-REX::VP16 fusions is decreased with a reduced-to-oxidized cofactor ratio increase. Transient and stabilized cell lines bearing the REX::VP16 and the REX binding operator (ROP) exhibit two circadian luminescence cycles. Consistent with these results, NADH oscillations are observed in host cells, indicating REX can act as a NADH sensor to report intracellular dynamic redox homeostasis in mammalian cells in real time. NADH oscillations provide another metabolic signal for coupling the circadian clock and cellular metabolic states.
AB - It is known that there are mechanistic links between circadian clocks and metabolic cycles. Reduced nicotinamide adenine dinucleotide (NADH) is a key metabolic cofactor in all living cells; however, it is not known whether levels of NADH oscillate or not. Here we employed REX, a bacterial NADH-binding protein, fused to the VP16 activator to convert intracellular endogenous redox balance into transcriptional readouts by a reporter gene in mammalian cells. EMSA results show that the DNA binding activity of both T- and S-REX::VP16 fusions is decreased with a reduced-to-oxidized cofactor ratio increase. Transient and stabilized cell lines bearing the REX::VP16 and the REX binding operator (ROP) exhibit two circadian luminescence cycles. Consistent with these results, NADH oscillations are observed in host cells, indicating REX can act as a NADH sensor to report intracellular dynamic redox homeostasis in mammalian cells in real time. NADH oscillations provide another metabolic signal for coupling the circadian clock and cellular metabolic states.
UR - http://www.scopus.com/inward/record.url?scp=84994751024&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84994751024&partnerID=8YFLogxK
U2 - 10.1074/jbc.M116.728774
DO - 10.1074/jbc.M116.728774
M3 - Article
C2 - 27645993
AN - SCOPUS:84994751024
SN - 0021-9258
VL - 291
SP - 23906
EP - 23914
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 46
ER -