Quantitative Effects of Magnesium Chloride Stress on Aggregation of Sup35p in [psi-] Yeast Cells
Abstract:[PSI+] phenotype can be transiently induced when Magnesium chloride (MgCl2) was the selective pressure in SUP35 repeat-expansion mutant [psi-] yeast strains. We further investigated [PSI+] phenotype change under different MgCl2 conditions with native Sup35p and quantified the Sup35p status changes with fluorescence recovery after photobleaching (FRAP) and semi-denaturing detergent-agarose gel electrophoresis (SDD-AGE) analysis. It was found that the [PSI+] phenotype presented a dose-dependent relationship with the concentrations of MgCl2. Furthermore, Sup35p aggregated in MgCl2 treated cells but did not form large aggregates as it does in [PSI+] cells, and the size of Sup35p aggregates showed a time-dependent relationship with the MgCl2 application. The aggregation of Sup35p strictly depended on the presence of MgCl2 stress in our strains.
Keywords: Confocal Microscopy; FRAP assay; Green Fluorescent Protein; Immo-bilon polyvinylidene difluoride sheets; Immunoblotting; Live-Time Fluorescence Analysis; Magnesium chloride; NGMC diffusion; PSI+; SDD-AGE; SDS-PAGE; SDS-insoluble Sup35p; SDS-resistant Sup35p polymers; Saccharomyces cerevisiae; Sup35p; Sup35p in [psi-] Yeast Cells; Tris-borate-EDT; Western Blot Analysis; [PSI+]; adenine; anti-Sup35p antiserum; fluorescence recovery after photobleaching; guanidine curing; guanidine-hydrochloride; isogenic [Gpsi-] cells; nonsense suppression; prion; quantitative effects; semi-denaturing detergent-agarose gel electrophoresis; stress
Document Type: Research Article
Publication date: December 1, 2010
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