title: Silencing of nicotinamide nucleotide transhydrogenase impairs cellular redox homeostasis and energy metabolism in PC12 cells.
abstract: Mitochondrial NADPH generation is largely dependent on the inner-membrane nicotinamide nucleotide transhydrogenase (NNT), which catalyzes the reduction of NADP(+) to NADPH utilizing the proton gradient as the driving force and NADH as the electron donor. Small interfering RNA (siRNA) silencing of NNT in PC12 cells results in decreased cellular NADPH levels, altered redox status of the cell in terms of decreased GSH/GSSG ratios and increased H(2)O(2) levels, thus leading to an increased redox potential (a more oxidized redox state). NNT knockdown results in a decrease of oxidative phosphorylation while anaerobic glycolysis levels remain unchanged. Decreased oxidative phosphorylation was associated with a) inhibition of mitochondrial pyruvate dehydrogenase (PDH) and succinyl-CoA:3-oxoacid CoA transferase (SCOT) activity; b) reduction of NADH availability, c) decline of mitochondrial membrane potential, and d) decrease of ATP levels. Moreover, the alteration of redox status actually precedes the impairment of mitochondrial bioenergetics. A possible mechanism could be that the activation of the redox-sensitive c-Jun N-terminal kinase (JNK) and its translocation to the mitochondrion leads to the inhibition of PDH (upon phosphorylation) and induction of intrinsic apoptosis, resulting in decreased cell viability. This study supports the notion that oxidized cellular redox state and decline in cellular bioenergetics - as a consequence of NNT knockdown - cannot be viewed as independent events, but rather as an interdependent relationship coordinated by the mitochondrial energy-redox axis. Disruption of electron flux from fuel substrates to redox components due to NNT suppression induces not only mitochondrial dysfunction but also cellular disorders through redox-sensitive signaling.
authors.affiliation: Department of Pharmacology and Pharmaceutical Sciences, School of Pharmacy, University of Southern California, Los Angeles, CA 90089, USA.
authors.forename: Fei
authors.initials: F
authors.lastname: Yin
authors.forename: Harsh
authors.initials: H
authors.lastname: Sancheti
authors.forename: Enrique
authors.initials: E
authors.lastname: Cadenas
mesh_terms: D000818:Animals; D017209:Apoptosis; D004734:Energy Metabolism; D020868:Gene Silencing; D006706:Homeostasis; D006861:Hydrogen Peroxide / Q000378:metabolism; D048031:JNK Mitogen-Activated Protein Kinases / Q000502:physiology; D008928:Mitochondria / Q000378:metabolism; D009249:NADP / Q000096:biosynthesis; D009250:NADP Transhydrogenases / Q000235:genetics / Q000502:physiology*; D010084:Oxidation-Reduction; D016716:PC12 Cells; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats
chemical_list: D034741:RNA, Small Interfering; D009249:NADP; D006861:Hydrogen Peroxide; D009250:NADP Transhydrogenases; D048031:JNK Mitogen-Activated Protein Kinases