<i>S</i>-Mercuration of ubiquitin carboxyl-terminal hydrolase L1 through Cys152 by methylmercury causes inhibition of its catalytic activity and reduction of monoubiquitin levels in SH-SY5Y cells

Abstract

-Methylmercury (MeHg) is an environmental electrophile that covalently modifies cellular proteins. In this study, we identified proteins that undergo S-mercuration by MeHg. By combining two-dimensional SDS-PAGE, atomic absorption spectrometry and ultra performance liquid chromatography mass spectrometry (UPLC/MS/MS), we revealed that ubiquitin carboxyl-terminal hydrolase L1 (UCH-L1) is a target for S-mercuration in human neuroblastoma SH-SY5Y cells exposed to MeHg (1 μM, 9 hr). The modification site of UCH-L1 by MeHg was Cys152, as determined by matrix-assisted laser desorption ionization time-of-flight mass spectrometry. MeHg was shown to inhibit the catalytic activity of recombinant human UCH-L1 in a concentration-dependent manner. Knockdown of UCH-L1 indicated that this enzyme plays a critical role in regulating mono-ubiquitin (monoUb) levels in SH-SY5Y cells and exposure of SH-SY5Y cells to MeHg caused a reduction in the level of monoUb in these cells. These observations suggest that UCH-L1 readily undergoes S-mercuration by MeHg through Cys152 and this covalent modification inhibits UCH-L1, leading to the potential disruption of the maintenance of cellular monoUb levels.

Methylmercury, an environmental electrophile capable of activation and disruption of the Akt/CREB/Bcl-2 signal transduction pathway in SH-SY5Y cells

“…This indicated that S -mercuration of MeHg, resulting in inhibition of PTEN contributes, at least partially, to the activation of Akt during exposure to lower concentrations of MeHg. We have identified other molecular targets (e.g., Mn-SOD, nNOS, arginase I, SDH, Keap1 and UCH-L1) of MeHg using cell-free systems incubated with MeHg, cultured cells exposed to MeHg and experimental animals treated MeHg 5 6 7 8 9 . Among them, UCH-L1 is a cellular protein candidate that may be associated with activation of Akt 39 40 41 .…”

“…MeHg-mediated toxicity is, at least in part, due to its electrophilic nature, which allows it to easily react with nucleophiles such as protein thiol groups, resulting in the disruption of cellular homeostasis through the S -mercuration of proteins 3 4 . We have reported that MeHg covalently modifies a variety of proteins such as manganese superoxide dismutase (Mn-SOD), neuronal nitric oxide synthase (nNOS), arginase I, sorbitol dehydrogenase (SDH), and ubiquitin C -terminal hydrolase L1 (UCH-L1), resulting in their disruption 5 6 7 8 9 . In contrast, the environmental electrophile 1,2-naphthoquinone activates protein tyrosine phosphatase (PTP) 1B/epidermal growth factor receptor (EGFR) signalling, which is involved in cell survival, and kelch-like ECH-associated protein 1 (Keap1)/nuclear factor (erythroid-derived 2)-like 2 (Nrf2) signalling, which regulates phase-II xenobiotic detoxification enzymes and phase-III transporters associated with cellular protection against this electrophile through S -arylation of PTP1B and Keap1 10 11 .…”

S-nitrosylation of UCHL1 induces its structural instability and promotes α-synuclein aggregation

“…This modification increases the amount of UCHL1 in the insoluble fraction of SH-SY5Y cells and inhibited its hydrolase activity to 60%, reducing the level of ubiquitin in the soluble fraction of SH-SY5Y cells22. S-mercuration of UCHL1 through Cys152 by methylmercury causes inhibition of its catalytic activity and reduction of monoubiquitin level in SH-SY5Y cells23. All these observations indicate that Cys152 has an important role in neuronal survival in stress condition.…”

Toxicology of organic-inorganic hybrid molecules: bio-organometallics and its toxicology

“…and metalloids. Such studies published these past three years in the Journal of Toxicological Sciences investigated the toxicity of many types of metals, including cadmium (Ohtani et al, 2013;Miura et al, 2013;Baba et al, 2014;Du et al, 2014;Park et al, 2015;Lukkhananan et al, 2015), mercury (Yoshida et al, 2013;Kim et al, 2014), methylmercury (Hwang et al, 2013a(Hwang et al, , 2013bIwai-Shimada et al, 2013;Chang et al, 2013;Hirooka et al, 2013;Kim et al, 2013;Watanabe et al, 2013;Shao et al, 2015;Toyama et al, 2015), aluminum (Zhang et al, 2013;Jinzhu et al, 2015), manganese (Fujishiro et al, 2013), uranium (Lestaevel et al, 2013), arsenic (Tokumoto et al, 2013;Wang et al, 2015), arsine (Kato et al, 2014), organic bismuth (Kohri et al, 2015), chromium (Kimura et al, 2015), and tributyltin (Oyanagi et al, 2015). Although methylmercury is one of the organic-inorganic hybrid molecules (the minimum unit!…”