YODA: Software to facilitate high-throughput analysis of chronological life span,growth rate,and survival in budding yeast

Background  

The budding yeast Saccharomyces cerevisiae is one of the most widely studied model organisms in aging-related science. Although several genetic modifiers of yeast longevity have been identified, the utility of this system for longevity studies has been limited by a lack of high-throughput assays for quantitatively measuring survival of individual yeast cells during aging.     

Diverse modes of reproduction in the marine free-living ciliate <Emphasis Type="Italic">Glauconema trihymene</Emphasis>

Background  

Most free-living ciliates reproduce by equal fission or budding during vegetative growth. In certain ciliates, reproduction occurs inside the cyst wall, viz. reproductive cysts, but more complex reproductive strategies have generally been thought to be confined to parasitic or symbiotic species, e.g. Radiophrya spp.     

Time scale and dimension analysis of a budding yeast cell cycle model

Background  

The progress through the eukaryotic cell division cycle is driven by an underlying molecular regulatory network. Cell cycle progression can be considered as a series of irreversible transitions from one steady state to another in the correct order. Although this view has been put forward some time ago, it has not been quantitatively proven yet. Bifurcation analysis of a model for the budding yeast cell cycle has identified only two different steady states (one for G1 and one for mitosis) using cell mass as a bifurcation parameter. By analyzing the same model, using different methods of dynamical systems theory, we provide evidence for transitions among several different steady states during the budding yeast cell cycle.     

Germinating fission yeast spores delay in G1 in response to UV irradiation

Background  

Checkpoint mechanisms prevent cell cycle transitions until previous events have been completed or damaged DNA has been repaired. In fission yeast, checkpoint mechanisms are known to regulate entry into mitosis, but so far no checkpoint inhibiting S phase entry has been identified.     

Checkpoint independence of most DNA replication origins in fission yeast

Background

In budding yeast, the replication checkpoint slows progress through S phase by inhibiting replication origin firing. In mammals, the replication checkpoint inhibits both origin firing and replication fork movement. To find out which strategy is employed in the fission yeast, Schizosaccharomyces pombe, we used microarrays to investigate the use of origins by wild-type and checkpoint-mutant strains in the presence of hydroxyurea (HU), which limits the pool of deoxyribonucleoside triphosphates (dNTPs) and activates the replication checkpoint. The checkpoint-mutant cells carried deletions either of rad3 (which encodes the fission yeast homologue of ATR) or cds1 (which encodes the fission yeast homologue of Chk2).

Results

Our microarray results proved to be largely consistent with those independently obtained and recently published by three other laboratories. However, we were able to reconcile differences between the previous studies regarding the extent to which fission yeast replication origins are affected by the replication checkpoint. We found (consistent with the three previous studies after appropriate interpretation) that, in surprising contrast to budding yeast, most fission yeast origins, including both early- and late-firing origins, are not significantly affected by checkpoint mutations during replication in the presence of HU. A few origins (~3%) behaved like those in budding yeast: they replicated earlier in the checkpoint mutants than in wild type. These were located primarily in the heterochromatic subtelomeric regions of chromosomes 1 and 2. Indeed, the subtelomeric regions defined by the strongest checkpoint restraint correspond precisely to previously mapped subtelomeric heterochromatin. This observation implies that subtelomeric heterochromatin in fission yeast differs from heterochromatin at centromeres, in the mating type region, and in ribosomal DNA, since these regions replicated at least as efficiently in wild-type cells as in checkpoint-mutant cells.

Conclusion

The fact that ~97% of fission yeast replication origins – both early and late – are not significantly affected by replication checkpoint mutations in HU-treated cells suggests that (i) most late-firing origins are restrained from firing in HU-treated cells by at least one checkpoint-independent mechanism, and (ii) checkpoint-dependent slowing of S phase in fission yeast when DNA is damaged may be accomplished primarily by the slowing of replication forks.     

Analysis of the splicing machinery in fission yeast: a comparison with budding yeast and mammals

Based on genetic and bioinformatic analysis, 80 proteins from the newly sequenced Schizosaccharomyces pombe genome appear to be splicing factors. The fission yeast splicing factors were compared to those of Homo sapiens and Saccharomyces cerevisiae in order to determine the extent of conservation or divergence that has occurred over the billion years of evolution that separate these organisms. Our results indicate that many of the factors present in all three organisms have been well conserved throughout evolution. It is calculated that 38% of the fission yeast splicing factors are more similar to the human proteins than to the budding yeast proteins (>10% more similar or similar over a greater region). Many of the factors in this category are required for recognition of the 3′ splice site. Ten fission yeast splicing factors, including putative regulatory factors, have human homologs, but no apparent budding yeast homologs based on sequence data alone. Many of the budding yeast factors that are absent in fission yeast are associated with the U1 and U4/U6.U5 snRNP. Collectively the data presented in this survey indicate that of the two yeasts, S.pombe contains a splicing machinery more closely reflecting the archetype of a spliceosome.     

Modeling the evolution of a classic genetic switch

Background  

The regulatory network underlying the yeast galactose-use pathway has emerged as a model system for the study of regulatory network evolution. Evidence has recently been provided for adaptive evolution in this network following a whole genome duplication event. An ancestral gene encoding a bi-functional galactokinase and co-inducer protein molecule has become subfunctionalized as paralogous genes (GAL1 and GAL3) in Saccharomyces cerevisiae, with most fitness gains being attributable to changes in cis-regulatory elements. However, the quantitative functional implications of the evolutionary changes in this regulatory network remain unexplored.     

What makes species unique? The contribution of proteins with obscure features

Background  

Proteins with obscure features (POFs), which lack currently defined motifs or domains, represent between 18% and 38% of a typical eukaryotic proteome. To evaluate the contribution of this class of proteins to the diversity of eukaryotes, we performed a comparative analysis of the predicted proteomes derived from 10 different sequenced genomes, including budding and fission yeast, worm, fly, mosquito, Arabidopsis, rice, mouse, rat, and human.     

PhiC31 recombination system demonstrates heritable germinal transmission of site-specific excision from the <Emphasis Type="Italic">Arabidopsis</Emphasis> genome

Background  

The large serine recombinase phiC31 from broad host range Streptomyces temperate phage, catalyzes the site-specific recombination of two recognition sites that differ in sequence, typically known as attachment sites attB and attP. Previously, we characterized the phiC31 catalytic activity and modes of action in the fission yeast Schizosaccharomyces pombe.     

Experimental annotation of the human pathogen <Emphasis Type="Italic">Candida albicans</Emphasis> coding and noncoding transcribed regions using high-resolution tiling arrays

Background  

Compared to other model organisms and despite the clinical relevance of the pathogenic yeast Candida albicans, no comprehensive analysis has been done to provide experimental support of its in silico-based genome annotation.     

Doxycycline-regulated gene expression in the opportunistic fungal pathogen Aspergillus fumigatus

Background  

Although Aspergillus fumigatus is an important human fungal pathogen there are few expression systems available to study the contribution of specific genes to the growth and virulence of this opportunistic mould. Regulatable promoter systems based upon prokaryotic regulatory elements in the E. coli tetracycline-resistance operon have been successfully used to manipulate gene expression in several organisms, including mice, flies, plants, and yeast. However, the system has not yet been adapted for Aspergillus spp.     

The fission yeast DNA structure checkpoint protein Rad26ATRIP/LCD1/UVSD accumulates in the cytoplasm following microtubule destabilization

Background  

DNA structure checkpoints are conserved eukaryotic signal transduction pathways that help preserve genomic integrity. Upon detecting checkpoint signals such as stalled replication forks or double-stranded DNA breaks, these pathways coordinate appropriate stress responses. Members of the PI-3 kinase related kinase (PIKK) family are essential elements of DNA structure checkpoints. In fission yeast, the Rad3 PIKK and its regulatory subunit Rad26 coordinate the detection of checkpoint signals with pathway outputs.