Selective inhibition of the mouse brain Mn-SOD by methylmercury

Abstract

Changes in mRNA levels, protein contents and enzyme activities for brain Cu,Zn- and Mn-SOD by methylmercury chloride (MMC) administration, were examined, over a period of 12 days in ICR male mice. After subcutaneous administration of MMC (10 mg/kg) to mice, brain mercury content reached a maximum at 2 days and remained at that level for at least 5 days. MMC exposure resulted in a time-dependent decrease in the Mn-SOD activity: the enzyme activity at 5 days after exposure to MMC was about 60% of control level whereas this exposure was without effect on the Cu,Zn-SOD activity, indicating differential sensitivity of SOD isozymes to the metal. However, levels of mRNA and protein synthesis for Mn-SOD were unaffected by MMC administration. The direct effect of MMC on the both SOD activities were further examined with purified enzyme preparations. After each SOD isozyme (10 U) was incubated with 0.2 mM MMC for 24 h at pH 7.8, the enzyme activities for Cu,Zn- and Mn-SOD were 90% and 37% of control, respectively. Incubations at a ratio of SOD to MMC (1 : 600) for 24 h resulted in a substantial decrease in the enzyme activity of the Mn form; this isozyme-selective inactivation was noted at alkaline pH. A combination of isoelectric focusing-agarose gel electrophoresis (IEF-AGE) and synchrotron radiation X-ray fluorescence (SR-XRF) analysis revealed that Mn-SOD rather than Cu,Zn-SOD underwent modification. Furthermore, a decrease in native form of Mn-SOD protein after MMC exposure was confirmed by gel filtration chromatography. These results indicate that Mn-SOD, but not Cu,Zn-SOD, is susceptible to modification by MMC and the resulting alteration in structure appears to cause a loss of enzyme activities.

<i>S</i>-Mercuration of cellular proteins by methylmercury and its toxicological implications

“…The authors didn't determine which isozyme was involved in the enzyme loss, but eukaryotic cells contain two types of SODs, copper and zinc SOD (Cu,Zn-SOD) and manganese SOD (Mn-SOD). We later found that Mn-SOD is susceptible to being modifi ed by MeHg and that Cu,Zn-SOD is not (Shinyashiki et al, 1996). Exposing mice to MeHg at a dose of 10 mg/kg only once resulted in a time-dependent decrease in Mn-SOD activity but had little effect on Cu,Zn-SOD activity in the brain.…”

“…They found that the sensitivity of HeLa cells to MeHg was decreased by the overexpression of Mn-SOD, suggesting that the formation of superoxide anions in the mitochondria might be involved in the mechanism(s) involved in the cytotoxicity of MeHg. We have attempted to examine the mechanism involved in the relationship between Mn-SOD and MeHg (Kumagai et al, 1997;Shinyashiki et al, 1996). Levels of mRNA and protein synthesis for Mn-SOD were unaffected by MeHg administration.…”

DNA microarray analysis of human neuroblastoma SH-SY5Y cells exposed to methylmercury

“…Methylmercury (MeHg) is an electrophile that modifies cellular proteins, resulting in enzyme dysfunction and cell damage (Shinyashiki et al, 1996). We previously reported that MeHg activates NF-E2-related factor 2 (Nrf2), a transcription factor involved in cellular protection against MeHg toxicity in human neuroblastoma SH-SY5Y cells (Toyama et al, 2007(Toyama et al, , 2011.…”

Difference between kidney and liver in decreased manganese superoxide dismutase activity caused by exposure of mice to mercuric chloride

“…Mn-SOD activities were determined by reduction of acetylated cytochrome c in the presence of potassium cyanide, as described previously (Shinyashiki et al 1996). The reaction mixture (1.5 ml) consisted of 0.05 mM xanthine, 44 nM xanthine oxidase, 25 lM acetylated cytochrome c, the enzyme preparation and 50 mM potassium phosphate buffer (pH 7.8)/0.01 mM EDTA.…”

“…We have previously demonstrated that in mice subcutaneous injection of methylmercury chloride, at a dose of 10 mg/ kg, which affects the central nervous system because of its predominant accumulation there (Clarkson 1972), results in a reduction of the brain manganese SOD (Mn-SOD) activity (Shinyashiki et al 1996). The importance of Mn-SOD for protection against methylmercury toxicity was subsequently reported by Naganuma et al (1998).…”