Yeast red pigment, protein aggregates, and amyloidoses: a review.

Estimating the amyloid level in yeast Saccharomyces, we found out that the red pigment (product of polymerization of aminoimidazole ribotide) accumulating in ade1 and ade2 mutants leads to drop of the amyloid content. We demonstrated in vitro that fibrils of several proteins grown in the presence of the red pigment stop formation at the protofibril stage and form stable aggregates due to coalescence. Also, the red pigment inhibits reactive oxygen species accumulation in cells.

Yeast red pigment modifies cloned human α-synuclein pathogenesis in Parkinson disease models in Saccharomyces cerevisiae and Drosophila melanogaster.

Recently, we identified the yeast red pigment (RP), a polymer of 1-(5'-Phosphoribosyl)-5-aminoimidazole, as a novel potential anti-amyloid agent for the therapy of neurodegenerative diseases. The purpose of this study was to further validate RP for treatment of Parkinson's disease (PD) and to clarify molecular mechanisms involved in the reduction of amyloid cytotoxicity. We investigated RP effects in vivo using Saccharomyces cerevisiae and Drosophila melanogaster PD models.

Yeast red pigment modifies Amyloid beta growth in Alzheimer disease models in both Saccharomyces cerevisiae and Drosophila melanogaster.

The effect of yeast red pigment on amyloid-β (Aβ) aggregation and fibril growth was studied in yeasts, fruit flies and in vitro. Yeast strains accumulating red pigment (red strains) contained less amyloid and had better survival rates compared to isogenic strains without red pigment accumulation (white strains). Confocal and fluorescent microscopy was used to visualise fluorescent Aβ-GFP aggregates.

[Comparative structural and functional characteristics of different forms of Saccharomyces cerevisiae red pigment and its synthetic analogue].

Structural and functional characteristics of the yeast red pigment (product of polymerization of N1-(beta-D-ribofuranosyl)-5-aminoimadazole), isolated from adel 1 mutant cells of Saccharomyces cerevisiae, its deribosylated derivatives (obtained by acid hydrolysis) and its synthetic pigment analogue (product of polymerization of N1-methyl-5-aminoimadazole in vitro) has been obtained. Products of in vitro polymerization were identified using mass spectrometry. The ability of these pigments to inhibit amyloid formation using insulin fibrils was compared.

[The effect of red pigment of Saccharomyces cerevisiae on insulin fibril formation in vitro].

The effect of the yeast red pigment, the result of polymerization of AIR, and of its low molecular weight derivative (presumably devoid of phosphoribosyl moiety) on the formation of amyloid fibrils in vitro was studied. Both the red pigment and its derivative, the result of acid hydrolysis of the original pigment, were shown to diminish the intensity of amyloid bound Thioflavine T fluorescence. Correlation between the decrease of the intensity of Thioflavine T fluorescence and the concentration of both forms of the red pigment was demonstrated.

The effect of red pigment on the amyloidization of yeast proteins.

The intensity of amyloid-bound thioflavine T fluorescence was studied in crude lysates of yeast strains carrying mutations in the ADE1 or ADE2 genes and accumulating the red pigment (a result of polymerization of aminoimidazoleribotide), and in white isogenic strains--either adenine prototrophs or carrying mutations at the first stages of purine biosynthesis. We found that the red pigment leads to a drop of amyloid content. This result, along with the data on separation of protein polymers of white and red strains in PAGE, suggests that the red pigment inhibits amyloid fibril formation.

[The effect of red pigment on amyloidization of yeast proteins].

Amyloid bound thioflavine T fluorescence was studied in the lysates of yeast strains carrying mutations in genes ADE1 or ADE2 and accumulating red pigment, a result of polymerization of aminoimidazoleribotide (an intermediate of adenine biosynthesis). The fluorescence is drastically enhanced in the case of cells grown in media containing high concentration of adenine (100 mg/l) that blocks accumulation of red pigment. Blocks at first stages of purine biosynthesis de novo also impede red pigment and lead to the same effect on thioflavine fluorescence.

[Comparison of crude lysate pellets of isogenic strains of yeast with different prion composition: identification of a set of prion-associated proteins].

A new approach: comparative analysis of proteins of the pellets of crude cell lysates of isogenic strains of Saccharomyces cerevisiae differing by their prion composition permitted to identify a large group of prion-associated proteins in yeast cells. 2D-electrophoresis followed by MALDI-analysis of a recipient [psi-] strain and of [PSI+] cytoductant led to identification of 35 proteins whose aggregation state responded to a shift of prion(s) content. Approximately half of these proteins belonged to functional groups of chaperones and enzyme involved in glucose metabolism.

Prion-associated proteins in yeast: comparative analysis of isogenic [PSI(+)] and [psi(-)] strains.

A large group of prion-associated proteins was identified in yeast cells using a new approach, comparative analysis of pellet proteins of crude cell lysates in isogenic strains of Saccharomyces cerevisiae differing by their prion composition. Two-dimensional (2D) electrophoresis followed by MALDI analysis of the pellet proteins of [PSI(+)] and [psi(-)] strains after prion elimination by GuHCl and prion transmission by cytoduction permitted identification of ca. 40 proteins whose aggregation state correlated with the change of prion(s) content.

[Estimating of changes in the amyloid and prion content of yeast cells].

An attempt was made at estimating the overall amyloid content of yeast cells by treating crude cellular lysates with thioflavin T, the agent specifically staining amyloid fibrils. We demonstrated that overproduction of the yeast chaperone Hsp104p, as well as GuHCI treatment of the [PSI+] cells led both to elimination of the [PSI+] factor and to a stable decrease of the overall amyloid content estimated by intensity of fluorescence (IF) of the thioflavin T. At the same time, overexpression of gene SUP35, coding the protein prionizable to [PSI+], led to generation of [PSI+] clones with higher IF of thioflavin T.

Bud selection and apoptosis-like degradation of nuclei in yeast heterokaryons: a KAR1 effect.

It has been shown that defects in cell fusion during mating can trigger programmed cell death in the yeast Saccharomyces cerevisiae. We wished to test whether defects in nuclear migration during cell fusion have the same effect. A partial pedigree analysis of nine kar1 x KAR1 crosses of two different types (four alpha KAR1 x a kar1 and five alpha kar1 x a KAR1 crosses) was carried out, and quantitative estimates of the frequencies of different mother/daughter (m/d) classes were obtained.

[Phylogenetic analysis of yeast genes with large stereospecific anomalies in their promoter region].

Using an original computer program, we analysed 3 yeast Saccharomyces cerevisiae genes that contain large stereospecific anomalies (SA) in their promoter regions. Homologous genes for higher eukaryotic organisms contain large SA either in the same promoter regions or in one of their introns, the involved dinucleotide and DNA helical repeat being often conserved. We suppose that both promoter and enhancer-like sequences for these genes are evolutionary related and/or are regulated by related proteins.

The impact of manipulations with cytoplasmically inherited factors on nuclear transmission and degradation in yeast heterokaryons.

Heterokaryotic zygotes in yeast provide a unique possibility to study the survival and transmission of two genetically diverse nuclei in one cell. Using partial pedigree analysis, we show that various treatments used to change cytoplasmic hereditary determinants can essentially affect nuclear transmission in yeast heterokaryons. This includes choice of nucleus to enter the first bud and incidence of various classes of mother/daughter pairs demonstrating nuclear degradation patterns in heterokaryotic zygotes.

Concealed nuclei in Saccharomyces strains.

We have found that cells derived from heterokaryons (HK) showing phenotypical traits, coded by the nucleus of one parental strain and by the cytoplasm of the other, may produce mitotic progeny in which the second nucleus is apparently present but not expressed. This 'concealed' nucleus can be forced to expression after growth of these cytoductants on proper selective media. Using a micromanipulator, the buds containing both parental nuclei were isolated in various crosses.

[Stereospecific DNA anomalies in the human chromosome 21].

Using our own original computer program, we analysed more than 10 millions b.p. of the complete nucleotide sequence in the human chromosome 21.

[Factors affecting the frequency of concealed heterokaryosis in Saccharomyces cerevisiae].

In this work, studies on the phenomenon of concealed heterokaryosis that we previously detected in the saccharomycetes yeast strains were continued. New approaches to high effectiveness of isolation of cytoductants carrying the concealed nucleus were implemented, and the composition of individual concealed heterokaryons, zygotic clones, and the first zygotic buds was analyzed by a micromanipulator. The relationship between a delay in the growth of the parental strain (a potential donor of the concealed nucleus) and a decline in the frequency of the appearance of concealed heterokaryons (HKC) was observed.

[Detection of concealed "illegitimate" nuclei in tetrad analysis of the diploid progeny of heterokaryons in Saccharomyces cerevisiae].

In this work, the studies on the previously detected phenomenon of concealed heterokaryosis in Saccharomyces cerevisiae were continued. In genetic and Southern blotting experiments, one of the nuclei in the heterokaryon was shown to be active (capable of division and ensuring the corresponding cell phenotype), whereas the other was not expressed until the heterokaryotic clone was transferred to the medium selective for this concealed nucleus. Moreover, the concealed nucleus was able to assume the active state after fusion with the second parental nucleus.

[Stereospecific anomalies in the structure of DNA from chromosomes V, XII, XIII, XIV, XV and XVI of saccharomycetes yeasts].

Using an original computer program we analysed complete nucleotide sequences of chromosomes V, XII, XIII, XIV, XV, and XVI in yeast. The full catalogue of 5 highest stereospecific anomalies and of stereospecific anomalies for 5 genes with highest CAI in each chromosome has been presented. Trains of different stereospecific anomalies, possibly showing areas of cooperative binding of different regulatory and structural proteins to DNA, are documented (see: Soidla, Lukina, 1998, 1999).

[Cryptic heterokaryons in strains of Saccharomyces cerevisiae. Conditions for detecting a cryptic nucleus].

A phenomenon discovered earlier, cryptic heterokaryosis in Saccharomyces yeast, has been further investigated. A phenotypically silent nucleus in a yeast cell may resume its expression after fusion with another parental cell. The resulting hybrid is capable of sporulation.

[Stereospecific anomalies in DNA structure of Saccharomyces chromosomes IV, VII, VIII, X and XI].

Using an original computer program we analysed complete nucleotide sequences of chromosomes IV, VII, VIII, X and XI in yeast. Data about 5 largest stereospecific anomalies in each chromosome are presented together with those for 5 genes with highest CAI in each chromosome. Clusters of different stereospecific anomalies are demonstrated, including trains of not overlapping anomalies, possibly showing areas of cooperative binding of different regulatory and structural proteins to DNA (Soidla, Lukina, 1998).

[Cryptic heterokaryons in Saccharomyces cerevisiae strains. Model experiments].

Heterokaryons capable of segregating clones of different phenotypes were obtained in saccharomycetes yeast. Clones expressing phenotypic traits encoded by the nuclear genome of one parent and the mitochondrial genome of another were shown to contain cells that carried the second phenotypically silent nucleus. The cryptic nucleus can be uncovered by growing these "cytoductants" on the corresponding medium.

[Cryptic heterokaryons in Saccharomyces cerevisiae strains: pseudopleiotropic suppression in the multiple marked YPH857 strain].

The yeast strain YPH857 carrying multiple genetic markers was shown to segregate clones that had the pleiotropic suppression phenotype. The phenotype was designated Ppsu+. This suppression involves deletion alleles of the TRP1 and HIS3 genes and an insertion in the URA3 gene.

[Stereospecific anomalies in the structure of five completely sequenced yeast chromosomes].

Using an original computer program we analysed complete nucleotide sequences of chromosomes I, II, III, VI and IX in yeast cells. As a general rule, we found large stereospecific anomalies near genes with a presumed high expression level (a full catalogue of such anomalies for 5 genes with highest CAI in each chromosome is presented). As a rule, they are also present at mobile genetic elements.