ABSTRACT
The Aß peptide is widely considered a major cause of Alzheimer's disease since it causes neuronal death in an oligomerisation-dependent manner. In order to identify new inhibitors of Aß that may be chemo preventative for Alzheimer's disease, a yeast assay that qualitatively determines the amounts and state of the human Aß42 peptide has been developed. Yeast assays such as this can be applied to studies on aggregation toxicity, autophagic responses and drug screening in Alzheimer's disease.
Subject(s)
Alzheimer Disease/metabolism , Alzheimer Disease/pathology , Amyloid beta-Peptides/chemistry , Amyloid beta-Peptides/toxicity , Autophagy/drug effects , Drug Evaluation, Preclinical/methods , Peptide Fragments/chemistry , Peptide Fragments/toxicity , Saccharomyces cerevisiae/drug effects , Alzheimer Disease/drug therapy , Amino Acid Sequence , Animals , Cell Culture Techniques , Flow Cytometry , Humans , Molecular Sequence Data , Protein Aggregates/drug effects , Saccharomyces cerevisiae/cytology , Saccharomyces cerevisiae/genetics , Transformation, GeneticABSTRACT
A family of 21 polyphenolic compounds consisting of those found naturally in danshen and their analogues were synthesized and subsequently screened for their anti-amyloidogenic activity against the amyloid beta peptide (Aß42) of Alzheimer's disease. After 24 h incubation with Aß42, five compounds reduced thioflavin T (ThT) fluorescence, indicative of their anti-amyloidogenic propensity (p < 0.001). TEM and immunoblotting analysis also showed that selected compounds were capable of hindering fibril formation even after prolonged incubations. These compounds were also capable of rescuing the yeast cells from toxic changes induced by the chemically synthesized Aß42. In a second assay, a Saccharomyces cerevisiae AHP1 deletant strain transformed with GFP fused to Aß42 was treated with these compounds and analyzed by flow cytometry. There was a significant reduction in the green fluorescence intensity associated with 14 compounds. We interpret this result to mean that the compounds had an anti-amyloid-aggregation propensity in the yeast and GFP-Aß42 was removed by proteolysis. The position and not the number of hydroxyl groups on the aromatic ring was found to be the most important determinant for the anti-amyloidogenic properties.
Subject(s)
Amyloid/drug effects , Plant Extracts/pharmacology , Polyphenols/pharmacology , Saccharomyces cerevisiae/drug effects , Small Molecule Libraries/pharmacology , Amyloid beta-Peptides/toxicity , Peptide Fragments/toxicity , Peroxiredoxins/genetics , Plant Extracts/chemistry , Polyphenols/chemistry , Saccharomyces cerevisiae/metabolism , Salvia miltiorrhiza/chemistry , Small Molecule Libraries/chemistryABSTRACT
Folate, vitamin B9, is well recognized as being essential for cell growth. The utilization of folate is common to all cells, but the source of it may be quite different. For example, mammalian cells depend on exogenous uptake of folates, while plants and microbes can synthesize them. There has been little consideration of uptake of folate in microbial cells, and studies on the effects of folates in mammalian cells, where conditions are restricted. This study shows that exogenous folates (folic acid or folinic acid), causes Candida glabrata cells suspended in water alone to undergo two cycles of cell division and to form multiple buds. The effect was limited to cells in the stationary phase and more profound in quiescent cells. These data indicate a novel response of yeast to folates that may increase the utility of yeast as a model to study folate transport and signaling.
ABSTRACT
The tendency of amyloid ß (Aß42) peptide to misfold and aggregate into insoluble amyloid fibrils in Alzheimer's disease (AD) has been well documented. Accumulation of Aß42 fibrils has been correlated with abnormal apoptosis and unscheduled cell division which can also trigger the death of neuronal cells, while oligomers can also exhibit similar activities. While investigations using chemically-synthesized Aß42 peptide have become common practice, there appear to be differences in outcomes from different preparations. In order to resolve this inconsistency, we report 2 separate methods of preparing chemically-synthesized Aß42 and we examined their effects in yeast. Hexafluoroisopropanol pretreatment caused toxicity while, ammonium hydroxide treated Aß42 induced cell proliferation in both C. glabrata and S. cerevisiae. The hexafluoroisopropanol prepared Aß42 had greater tendency to form amyloid on yeast cells as determined by thioflavin T staining followed by flow cytometry and microscopy. Both quiescent and non-quiescent cells were analyzed by these methods of peptide preparation. Non-quiescent cells were susceptible to the toxicity of Aß42 compared with quiescent cells (p < 0.005). These data explain the discrepancy in the previous publications about the effects of chemically-synthesized Aß42 on yeast cells. The effect of Aß42 on yeast cells was independent of the size of the peptide aggregates. However, the Aß42 pretreatment determined whether the molecular conformation of peptide resulted in proliferation or toxicity in yeast based assays.
Subject(s)
Amyloid beta-Peptides/chemistry , Candida glabrata/cytology , Peptide Fragments/chemistry , Saccharomyces cerevisiae/cytology , Alzheimer Disease/metabolism , Apoptosis , Benzothiazoles , Flow Cytometry , Microscopy, Confocal , Microscopy, Electron, Transmission , Neurons/metabolism , Peptides/chemistry , Propanols/chemistry , Protein Binding , Protein Conformation , Thiazoles/chemistryABSTRACT
The major molecules associated with Alzheimer's disease, the phosphorylated protein tau and the 42 amino acid peptide, amyloid-ß (Aß), have recently been analyzed in yeast. These yeast studies have provided major new insights into the effects of tau and Aß and, at the same time, offered new approaches to rapidly search for chemicals that may be involved in prevention of Alzheimer's disease. The following review summarizes the role of yeast and its contribution in Alzheimer's disease research, and highlights important studies that have been conducted in this model organism.
