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Effects of microtubule-associated protein (MAP) expression on methylmercury-induced microtubule disassembly
1Department of Biology, University of Ottawa, Ottawa, Ontario, K1N 6N5, Canada.
Abstract:
The sensitivity of microtubules (MTs) to methylmercury- (MeHg) induced disassembly was compared in undifferentiated, MAP1A- and MAP2C-transfected, and neuronally differentiated P19 Embyronal Carcinoma (EC) cells. The extent of MT disassembly was examined qualitatively by immunofluorescence microscopy and Western blotting and quantitatively by dot blotting of polymer and soluble proteins extracts. Immunofluorescence microscopy showed that MeHg disassembled MTs in a time- and dose-dependent manner and that MTs in both MAP2C-transfected and neuronally differentiated cells, but not those in MAP1A-transfected cells, were significantly more resistant to MeHg-induced MT depolymerization than those in undifferentiated cells. These results suggest that MAP2C has a greater ability to stabilize MTs against MeHg-induced disassembly than MAP1A. Surprisingly, however, when the extent of MT disassembly was assessed by Western blotting and by quantitative dot blotting, no change was observed in the amounts of tubulin, MAP2, or MAP1A, in the polymer and soluble fractions in MeHg-treated samples, compared to the control cells that were not treated. These data show that, although MeHg treatment resulted in the disassembly of MTs, they were not depolymerized as detergent-soluble subunits, but rather appeared to form insoluble tubulin-MAP oligomers or aggregates.
Insights
Methylmercury (MeHg) causes microtubule (MT) disassembly. MAP2C-transfected and differentiated cells show increased MT resistance to MeHg, suggesting MAP2C stabilizes MTs against MeHg toxicity.
Area of Science:
- Cell Biology
- Neuroscience
- Toxicology
Background:
- Microtubules (MTs) are crucial cytoskeletal components.
- Methylmercury (MeHg) is a potent neurotoxin that disrupts MTs.
- Microtubule-associated proteins (MAPs) like MAP1A and MAP2C modulate MT stability.
Purpose of the Study:
- To compare the sensitivity of microtubules to methylmercury (MeHg)-induced disassembly.
- To investigate the role of MAP1A and MAP2C in stabilizing microtubules against MeHg.
- To examine the cellular response to MeHg-induced MT disassembly in P19 cells.
Main Methods:
- Utilized undifferentiated, MAP1A- and MAP2C-transfected, and neuronally differentiated P19 Embyronal Carcinoma (EC) cells.
- Assessed MT disassembly using immunofluorescence microscopy, Western blotting, and quantitative dot blotting.
- Analyzed polymer and soluble protein fractions to quantify tubulin, MAP1A, and MAP2 levels.
Main Results:
- MeHg induced time- and dose-dependent MT disassembly.
- MAP2C-transfected and neuronally differentiated cells exhibited significantly more resistant MTs to MeHg than undifferentiated cells.
- Western blotting and dot blotting revealed no change in soluble tubulin or MAPs, indicating MeHg forms insoluble aggregates rather than soluble subunits.
Conclusions:
- MAP2C confers greater stability to microtubules against MeHg-induced disassembly than MAP1A.
- MeHg-induced MT disassembly results in the formation of insoluble tubulin-MAP oligomers or aggregates.
- These findings highlight the differential stabilization roles of MAPs against toxic insults.