Mitochondrial Complex I Inhibitor Rotenone-Elicited Dopamine Redistribution from Vesicles to Cytosol in Human Dopaminergic SH-SY5Y Cells

Watabe, Masahiko; Nakaki, Toshio

doi:10.1124/jpet.107.127597

Cited by 60 publications

(52 citation statements)

References 29 publications

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“…A similar situation was observed in rat models of Parkinson's disease, whereas transfection with a yeast NADH dehydrogenase prevented the overproduction of ROS induced by rotenone (39,40). Recently, rotenone was described to stimulate ROS, protein carbonylation (61), and protein nitration, which was suggested to be a general signaling event underlying the formation of cytosolic protein aggregates, such as the Lewy bodies characteristic of Parkinson's disease (62). Rotenone also induces glutathione depletion (61; this work).…”

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confidence: 54%

Rotenone Enhances the Antifungal Properties of Staurosporine

et al. 2010

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We studied staurosporine-induced cell death in the filamentous fungus Neurospora crassa. The generation of reactive oxygen species during the process appears to be an important signaling event, since addition of the antioxidant glutathione prevents the effects of staurosporine on fungal growth. Selected mutants with mutations in respiratory chain complex I are extremely sensitive to the drug, stressing the involvement of complex I in programmed cell death. Following this finding, we determined that the complex I-specific inhibitor rotenone used in combination with staurosporine results in a synergistic and specific antifungal activity, likely through a concerted action on intracellular glutathione depletion. Paradoxically, the synergistic antifungal activity of rotenone and staurosporine is observed in N. crassa complex I mutants and in Saccharomyces cerevisiae, which lacks complex I. In addition, it is not observed when other complex I inhibitors are used instead of rotenone. These results indicate that the rotenone effect is independent of complex I inhibition. The combination of rotenone and staurosporine is effective against N. crassa as well as against the common pathogens Aspergillus fumigatus and Candida albicans, pointing to its usefulness as an antifungal agent.Programmed cell death (PCD) refers to a genetically controlled process of cellular suicide initiated by endogenous or extrinsic signals. Many of the genes involved are widely conserved from unicellular to multicellular organisms (46). Apoptosis and autophagy, with its particular characteristics, have been recognized as the main categories of PCD (27). The process of PCD is crucial for the development and homeostasis of metazoan organisms and has been implicated in a number of human disorders, including cancer and neurodegenerative and infectious diseases (3,10,25,55).The participation in PCD of mitochondria, the cellular organelles responsible for the production of most cellular ATP in eukaryotes (30), has been well established. Particularly, these organelles have a central role in the intrinsic (mitochondriondependent) pathway of apoptosis, which includes production of reactive oxygen species (ROS), membrane depolarization, ultrastructural changes, and the release of cytochrome c and other proteins (18,50,55). Drugs like staurosporine (STS), an inhibitor of protein kinases, have been used to induce the mitochondrion-dependent pathway of apoptosis (24, 35). Staurosporine (48) and derivatives have been used in clinical trials for cancer therapy (63). The complex I inhibitor rotenone too has been widely used to induce PCD and also extensively applied as a pesticide (11,39,56). Thus, these types of drugs can be employed for the acquisition of fundamental knowledge and for more practical applications, like modulation of the progression of PCD.Modulation of PCD by targeting metabolic pathways involved in the process can be exploited to the benefit of human health in several very significant situations, from cancer therapy (4, 57) to the treatment of f...

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confidence: 54%

Rotenone Enhances the Antifungal Properties of Staurosporine

et al. 2010

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“…Nitration of one or more tyrosine residues of TH was temporally associated with loss of enzymatic activity. The tyrosine nitration-mediated loss of TH activity parallels the decrease in dopamine levels, but this dopamine decrease alone is not sufficient to induce apoptosis because reserpine-induced depletion of intracellular dopamine does not lead to apoptosis (Watabe and Nakaki, 2007). Therefore, much less is known about the involvement of peroxynitrite in apoptosis induced by these toxins.…”

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confidence: 97%

Mitochondrial Complex I Inhibitor Rotenone Inhibits and Redistributes Vesicular Monoamine Transporter 2 via Nitration in Human Dopaminergic SH-SY5Y Cells

Watabe¹,

Nakaki²

2008

Mol Pharmacol

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Parkinson's disease is a progressive neurodegenerative disorder characterized by selective degeneration of nigrostriatal dopaminergic neurons. Long-term systemic mitochondrial complex I inhibition by rotenone induces selective degeneration of dopaminergic neurons in rats. We have reported dopamine redistribution from vesicles to the cytosol to play a crucial role in selective dopaminergic cell apoptosis. In the present study, we investigated how rotenone causes dopamine redistribution to the cytosol using an in vitro model of human dopaminergic SH-SY5Y cells. Rotenone stimulated nitration of the tyrosine residues of intracellular proteins. The inhibition of nitric-oxide synthase or reactive oxygen species decreased the amount of nitrotyrosine and attenuated rotenone-induced apoptosis. When we examined the intracellular localization of dopamine immunocytochemically using anti-dopamine/vesicular monoamine transporter 2 (VMAT2) antibodies and quantitatively using high-performance liquid chromatography, inhibiting nitration was found to suppress rotenone-induced dopamine redistribution from vesicles to the cytosol. We demonstrated rotenone to nitrate tyrosine residues of VMAT2 using an immunocytochemical method with anti-nitrotyrosine antibodies and biochemically with immunoprecipitation experiments. Rotenone inhibited the VMAT2 activity responsible for the uptake of dopamine into vesicles, and this inhibition was reversed by inhibiting nitration. Moreover, rotenone induced the accumulation of aggregate-like formations in the stained image of VMAT2, which was reversed by inhibiting nitration. Our findings demonstrate that nitration of the tyrosine residues of VMAT2 by rotenone leads to both functional inhibition and accumulation of aggregate-like formations of VMAT2 and consequently to the redistribution of dopamine to the cytosol and apoptosis of dopaminergic SH-SY5Y cells.

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“…4A). Using this model, we previously reported that the inhibition of rotenone-generated ROS or NO protected the cells from its toxicity (Watabe and Nakaki, 2007b;Watabe and Nakaki, 2008). Both L-NAME, an inhibitor of nitric oxide synthase (NOS), and N-acetylcysteine (NAC), a scavenger of ROS, suppressed the rotenone-induced increase in CK2 (Fig.…”

Section: Regulation Of Tubulin Acetylation By Ck2 In a Rotenone Modelmentioning

confidence: 99%

“…If this hypothesis was true, CK2 should be required for the proper formation of aggresomes. We previously investigated neuronal degeneration by using an in vitro rotenone model of human SH-SY5Y cells Watabe and Nakaki, 2007a;Watabe and Nakaki, 2007b;Watabe and Nakaki, 2008). Rotenone was used to generate oxidative stress in the cells 1521 CK2 regulates HDAC6 function and we analyzed aggresome formation.…”

Section: Regulation Of Tubulin Acetylation By Ck2 In a Rotenone Modelmentioning

confidence: 99%

Protein kinase CK2 regulates the formation and clearance of aggresomes in response to stress

Watabe

Nakaki

2011

Journal of Cell Science

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SummaryMisfolded protein aggregates elicit a stress response, and their clearance is crucial for cell survival. These aggregates are transported by cytoplasmic deacetylase HDAC6 and dynein motors to the aggresome via the microtubule network, and are removed by autophagic degradation. HDAC6 activity is necessary for both the transport and clearance of protein aggregates. However, the cellular factors that regulate HDAC6 activity remain unknown. Here we show that protein kinase CK2 is a crucial modulator of HDAC6 activity because CK2 directly phosphorylates HDAC6 and increases cytoplasmic deacetylase activity. Indeed, cells that expressed HDAC6 mutated at Ser458, a CK2-mediated phosphorylation site, failed to both form and clear aggresomes, and increased cytotoxicity. Interestingly, Ser458 is conserved only in higher primates, such as human and chimpanzee, but not in the rhesus macaque. These findings identify CK2 as a crucial protein involved in the formation and clearance of aggresomes, and hence in cell viability in response to misfolded protein stress.

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Mitochondrial Complex I Inhibitor Rotenone-Elicited Dopamine Redistribution from Vesicles to Cytosol in Human Dopaminergic SH-SY5Y Cells

Cited by 60 publications

References 29 publications

Rotenone Enhances the Antifungal Properties of Staurosporine

Rotenone Enhances the Antifungal Properties of Staurosporine

Mitochondrial Complex I Inhibitor Rotenone Inhibits and Redistributes Vesicular Monoamine Transporter 2 via Nitration in Human Dopaminergic SH-SY5Y Cells

Protein kinase CK2 regulates the formation and clearance of aggresomes in response to stress

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