Application of yeast to study the tau and amyloid-β abnormalities of Alzheimer's disease

The major molecules associated with Alzheimer's disease, the phosphorylated protein tau and the 42 amino acid peptide, amyloid-β (Aβ), have recently been analyzed in yeast. These yeast studies have provided major new insights into the effects of tau and Aβ 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

Estimation of in vitro prebiotic activity of individual and mixed dietary fibres

Background: Epidemiological studies have demonstrated a link between dietary fibre deficiency and prevalence of many &ldquo;Western diseases&rdquo; particularly colon related diseases. Many of the health benefits associated with dietary fibre are attributed to their prebiotic effect. However, not all fibres have the same prebiotic potential or the same impact on colon health.Objective: To examine the in vitro fermentation properties of individual and mixed dietary fibres by measuring fermentation byproducts over time.Design: Wheat bran and guar gum were selected for this study. Individual and mixed dietary fibres were added to batch fermentation system and were inoculated with fresh faecal inoculum (n= 4). Positive (inulin) and negative (no substrate) fermenters were also used to determine the differences. The pH of the five fermenters was adjusted to a baseline of 5.5 and 6.8 representing the pH of the proximal and distal sections of the colon respectively. Samples were drawn out of the fermenters at 0, 3, 9 and 24 hours for the analysis of pH, ammonia and short chain fatty acids (SCFAs).Outcomes: There were no significant differences in the pH levels at various time points between fermenters adjusted to pH 5.5 at baseline. However, in fermenters adjusted to pH 6.8 the pH of the fermenter containing wheat bran increased over the time (24h (P = 0.017)) due to production of a high amount of ammonia. The total SCFAs production was greater in fermenters containing combined fibres.Conclusion: There is a large inter-individual variation in the prebiotic effect of all types of dietary fibres, however, in the present study, dietary fibre combinations showed greater prebiotic potential compared with the individual fibres.<br /

Yeast as a platform for identification of chemo-protectors of Alzheimer&#039;s disease

The complexity of cellular pathways in neurodegenerative disease has limited the understanding of the molecular mechanisms underlying Alzheimer&amp;rsquo;s disease (AD). Therefore, to gain insight into AD and to devise potential therapeutic approaches, simple models are employed. Saccharomyces cerevisiae is currently utilised for analysing cellular toxicity and protein aggregation in neurodegenerative diseases including AD. This study has therefore focused on improvements and validation of the yeast screening model and assays to generate a more suitable disease model that can be used successfully for designing therapeutic strategies and interventions which correlate with in vitro and mammalian testing systems. This thesis investigates and shows for the first time that the pretreatment method of synthetic A&amp;beta;42 peptide determines its activity and ultimately how it interacts with yeast cells. The effects of A&amp;beta;42 appear to be limited to the yeast cell wall and interactions with the amyloidogenic cell wall proteins. S. cerevisiae was found to be more resistant than Candida glabrata to the effect of A&amp;beta;42 peptide. It was identified that conformational changes in the peptide due to preparation methods, determine its fate on toxicity and proliferation. Hexafluoroisopropanol pretreated A&amp;beta;42 had a greater tendency to aggregate on yeast cells as determined by thioflavin T staining followed by flow cytometry and microscopy. Quiescent cells were found to be more resistant to the toxicity of A&amp;beta;42 peptide than non-quiescent cells. Also, a preparation which results in toxicity to PC12 neuronal cells caused proliferation of S. cerevisiae and C. glabrata cells. This further indicated that extracellular toxicity assay in yeast using chemically-synthesised A&amp;beta;42 peptide results in a different outcome. My study shows for the first time that exogenous folate (folic acid or folinic acid) causes C. glabrata cells suspended in water to undergo two cycles of cell division and to form multiple buds. This effect was limited to cells in stationary phase and was more profound in quiescent cells. This study further exploited the cellular uptake of folate by utilising a putative folate transporter (YJL163C) and examined its role. Folate uptake appeared normal in YJL163C overexpressed yeast cells and a deletant mutant strain. Furthermore, the YJL163C deletion did not abolish folate-stimulated cell division. However, YJL163C overexpression in a diploid resulted in meiotic cell divisions. The A&amp;beta;42 was toxic to the YJL163C deletion stain suggesting that the YJL163C putative transporter was not a unique receptor for A&amp;beta;42. Another novel outcome of this study was the identification of the most suitable isomers of bio-inspired compounds with anti-amyloidogenic properties using both in vivo assays and in vitro assays. The positions of the hydroxyl moieties on the aromatic rings were found to be the major determinant of their potency rather than number of hydroxyl groups. This data implied that any yeast screening assays should be complemented with conventional in vitro assays to ensure their validity prior to high-throughput analyses. Another major development was the identification that the GFP-A&amp;beta;42 fusion system, which was utilised as a screening platform for identification of anti-oligomerisation of compounds, could only be used when expressed in AHP1 deletant mutant strain while no effect was observed in the wild-type strain. The data from this study highlights the pitfalls that exist in current yeast model systems and how they should be improved for future investigations

Exogenous folates stimulate growth and budding of Candida glabrata

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

Anti-amyloidogenic properties of some phenolic compounds

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 (A beta(42)) of Alzheimer&#039;s disease. After 24 h incubation with A beta(42), five compounds reduced thioflavin T (ThT) fluorescence, indicative of their anti-amyloidogenic propensity (p &lt; 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 A beta(42). In a second assay, a Saccharomyces cerevisiae AHP1 deletant strain transformed with GFP fused to A beta(42) 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-A beta(42) 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

Yeast as a model for studies on Abeta aggregation toxicity in Alzheimer&#039;s disease, autophagic responses, and drug screening

The Abeta peptide is widely considered a major cause of Alzheimer&#039;s disease since it causes neuronal death in an oligomerisation-dependent manner. In order to identify new inhibitors of Abeta that may be chemo preventative for Alzheimer&#039;s disease, a yeast assay that qualitatively determines the amounts and state of the human Abeta42 peptide has been developed. Yeast assays such as this can be applied to studies on aggregation toxicity, autophagic responses and drug screening in Alzheimer&#039;s disease