yeast Saccharomyces A model organism

The yeast Saccharomyces cerevisiae A model organism in genetics, genomics, and systems biology

the reason that yeast could serve as a model for all eukaryotic biology derives from the facility with which the relation between gene structure and protein function can be established.” (Botstein and Fink, Science 1988).

yeast Saccharomyces A model organism

Caenorhabditis  elegans  A novel model organism





yeast has graduated from a position as the premier model for eukaryotic cell biology to become the pioneer organism … of the entirely new fields of study called “functional genomics” and “systems biology.” These new fields look beyond the functions of individual genes and proteins, focusing on how these interact and work together to determine the properties of living cells and organisms.” (Botstein and Fink, Genetics 2011)

Content

  • Yeast is a eukaryotic organism
  • The yeast genome, definitions, and gene ontology
  • Foundations for the model organism: tools for genetic tractability
  • Defining function: functional genomics and networks
  • The concepts of yeast strains and variability
  • SGD: The Saccharomyces Genome Database

Drosophila as a Model Organism

The yeast saccharomyces cerevisiae: habitat, importance, and use

  •  Yeast lives primarily on fruits, flowers, and other sugar-containing substrates
  • Free-living organism: yeast copes with a wide range of environmental conditions:
  • Temperatures from freezing to about 55°C are tolerated
  • Yeasts proliferate from 12°C to 40°C  Growth is possible from pH 2.8-8.0
  • Almost complete drying is tolerated (dry yeast)
  • Yeast can still grow and ferment at sugar concentrations of 3M (high osmotic pressure)
  • Yeast can tolerate up to 20% alcohol
  • Saccharomyces cerevisiae is the main organism in wine production besides other yeasts; the reason is the enormous fermentation capacity, low pH and high ethanol tolerance.
  • Saccharomyces cerevisiae (carlsbergensis) is the beer yeast because it ferments sugar to alcohol even in the presence of oxygen, lager yeast ferments at 8°C.
  • Saccharomyces cerevisiae is the yeast used in baking because it produces carbon dioxide from sugar very rapidly.
  • Saccharomyces cerevisiae is used to produce commercially important proteins because it can be genetically engineered, it is regarded as safe and fermentation technology is highly advanced.
  • Saccharomyces cerevisiae is the most important eukaryotic cellular model system because it can be studied by powerful genetics.
  • Pioneers of genomics, functional genomics, systems and synthetic biology employ S. cerevisiae.

Drosophila as a Model Organism

A yeast cells is about 4-7µm large

Saccharomyces cerevisiae is a eukaryote

  • Belongs to fungi, ascomycetes
  • The unicellular organism with the ability to produce pseudohyphae
  • S. cerevisiae divides by budding (hence: budding yeast) while Schizosaccharomyces pombe divides by fission (hence: fission yeast). 
  • Budding results in two cells of unequal size, a mother (old cell) and a daughter (new cell).
  • Yeast life is not indefinite; yeast cells age and mothers die after about 30-40 divisions.
  • The cell has a eukaryotic structure with different organelles:
  • Cell wall consisting of glucans, mannans, and proteins
  • Periplasmic space with hydrolytic enzymes
  • Plasma membrane consisting of a phospholipid bilayer and many different proteins
  • Nucleus with nucleolus
  • Vacuole as storage and hydrolytic organelle
  • Secretory pathway with the endoplasmic reticulum, Golgi apparatus, and secretory vesicles
  • Peroxisomes for oxidative degradation
  • Mitochondria for respiration

Life cycle of yeasts

The life cycle of yeasts: yeast has a sex life

  • Yeast cells can proliferate both as haploids (1n, one copy of each chromosome) and as diploids (2n, two copies of each chromosome).
  •  Haploid cells have one of two mating types: a or alpha (α).
  •  Two haploid cells can mate to form a zygote from which a diploid cell buds off.
  •  Under nitrogen starvation, diploid cells undergo meiosis and sporulation to form an ascus with four haploid spores.
  •  Those germinate to form haploid cells.
  •  Hence, the properties of the meiotic products can be studied directly.

Spores (survival structures




Yeast: a unicellular organism with different cell types

  • Thus, although yeast is unicellular, we can distinguish different cell types with different genetic programs:
  •  Haploid MATa versus MATα (can respond to pheromone, can mate; cannot do meiosis)
  •  Haploid versus Diploid (MATa/alpha) (cannot respond to pheromone or mate, can sporulate)
  •  Spores (survival structures)
  •  Mothers and daughters (age n+1, can switch mating type, ago 0)

Yeast sex

  • Central to sexual communication is the pheromone response signal transduction pathway.
  •  All modules of that pathway consist of components conserved from yeast to human.
  • The pathway consists of a specific pheromone receptor, that binds a- or α-factor; it belongs to the class of seven transmembrane G-protein coupled receptors, like many human hormone receptors.
  •  Binding of pheromone stimulates reorientation of the cell towards the source of the pheromone (the mating partners).
  •  Binding of pheromone also stimulates a signaling cascade, a so-called MAP (Mitogen-Activated Protein) kinase pathway, similar to many pathways in human (animals and plants).
  •  This signaling pathway causes cell cycle arrest to prepare cells for mating (cells must be synchronized in the G1 phase of the cell cycle to fuse to a diploid cell).
  •  The pathway controls the expression of genes important for mating.

Structure and Analysis of Eukaryotic Genes

Haploids and diploids in nature and laboratory

  • In nature, yeast cells always grow as diploids: increases their chance to survive mutation of an essential gene (because there is always a second gene copy).
  •  However, from time to time deleterious mutations need to be ”cleaned out” and advantageous mutations should eventually be manifested.
  •  Under nitrogen starvation, diploid cells sporulate; under favorable conditions, haploid spores germinate, provided that they have received functional copies of all essential genes.
  •  This often means that only a single spore (if any) of a tetrad survives.
  •  How to make sure that this single spore finds a mating partner to form a diploid? The answer is mating type switch!
  •  After the first division, the mother cell switches mating type and mates with its daughter to form a diploid, which then, of course, is homozygous for all genes and starts a new clone of cells.
  •  If the mating-type can be switched and diploid is the preferred form, why then sporulate and have mating types?
  •  There are probably several reasons: (1) Spores are hardy and survive harsh conditions (2) Sporulation is a way to ”clean” the genome from accumulated mutations (3) Meiosis is a way to generate new combinations of alleles, which may turn out to be advantageous (4) Sometimes cells may find a mating partner from a different population and form a new clone, with possibly advantageous allele combination.
  •  In order to do yeast genetics and to grow haploid cells in the laboratory, mating type switch must be prevented: all laboratory strains are HO mutants and can not switch.

Yeast genetics

Yeast genetics: the genetic material

  • The S. cerevisiae nuclear genome has 16 chromosomes.
  •  In addition, there is a mitochondrial genome and a plasmid, the 2μ circle.
  •  The yeast chromosomes contain centromeres and telomeres, which are simpler than those of higher eukaryotes.
  •  The haploid yeast genome consists of about 12,500 kb and was completely sequenced as early 1996 (first complete genome sequence of a eukaryote).
  •  Since then, the genomes of numerous other yeast species and many different yeast strains where sequenced.

Yeast genetics: the genetic material

  •  The yeast genome is predicted to contain about 6,600 protein-coding genes. 5056 or 76% are verified, 764 or 12% are uncharacterized and 787 or 12% are “dubious”. http://www.yeastgenome.org/cache/genomeSnapshot.html
  •  Initial definition: likely protein-coding is an ORF of at least 100 codons. Of course, there are proteins smaller than 100 codons.
  •  There are other 1,400 genetic elements (chromosome structure, RNA genes, transposons, etc).
  •  There is substantial ”gene redundancy”, which originates from an ancient genome duplication and subsequent reshuffling.
  •  About 100 million years ago, a tetraploid was formed from a diploid. Tetraploids are viable but highly unstable. Perhaps extra copies of glycolytic genes provided a selective advantage. Ca 15% duplication remains.
  •  This means that there are many genes for which closely related homologs (paralogues) exist, which often are differentially regulated and whose products are adapted to specific conditions.
  •  The most extreme example are sugar transporter genes; there are more than twenty.
  •  Only a small percentage of yeast genes has introns, very few have more than one; mapping of introns is still incomplete.
  •  The intergenic space between genes is only between 200 and 1,000bp.
  •  The largest known regulatory sequences are spread over about 2,800bp (MUC1/FLO11).
  •  This means that the yeast genome is highly compact, about 1 gene per 2kb

Eukaryotic and prokaryotic gene structure 





Extreme metabolic adaptation

  • Preferred carbon sources are glucose, fructose, and sucrose.
  • Glucose and fructose mediate a gene expression program called glucose repression; genes required for utilization of different carbon sources.
  • At high concentrations, glucose and fructose are fermented to ethanol and carbon dioxide irrespective of the presence of oxygen: Crabtree effect.
  • A fermenting yeast cell consists of ca 50% of proteins involved in just the fermentative pathway: glycolysis.
  • Together with high ethanol tolerance, the fermentative capacity may confer a selective advantage in high sugar-containing environments.

PRS423

Gene ontology

Gene Ontology, or GO, is a major bioinformatics initiative emerged from yeast functional genomics to unify the representation of gene and gene product attributes across all species. More specifically, the project aims to:

  •  Maintain and develop a controlled vocabulary of gene and gene product attributes.
  • Annotate genes and gene products, and assimilate and disseminate annotation data.
  • Provide tools for easy access to all aspects of the data provided by the project.
  • Three main attributes: molecular function, biological process, and cellular component.

Tools that made yeast the prime model organism

  •  S. cerevisiae combines many advantages of bacterial with eukaryotic genetics.
  • Yeast can be “transformed” with replicating plasmids.  Transformation is efficient, although not as efficient as in E. coli.
  • This enables genetic studies (e.g. functional complementation of mutations with yeast or heterologous genes).
  • Complementation is an outstanding tool for functional analysis.
  • Plasmids also enable system perturbation such as overexpression.
  • Since yeast does not produce plasmid in high amounts shuttle vectors are used for cloning and production in E. coli and analysis in yeast.
  • Typical plasmid copy numbers are between 1/cell (centromeric plasmids) and 20-50/cell (episomal plasmids) and up to 200/cell.
  • Yeast can be transformed with more than one plasmid at a time.
  • This enables further system perturbation but also advanced approaches such as two-hybrid and FRET analysis for protein interactions

Eukaryotic Gene Structure

Tools that made yeast the prime model organism

  •  S. cerevisiae has an incredibly efficient system for homologous recombination.
  • Homologous recombination occurs between two pieces of DNA with the same sequence, commonly between two homologous chromosomes in meiosis, but also with transformed DNA.
  • This recombination system enables genome manipulation with the highest efficiency and of extreme precision.
  • A piece of DNS generated in vitro and transformed into yeast may find its rightful place in the genome based on only a few bases sequence identity and initiate the predicted genetic change.

Tools that made yeast




The Saccharomyces Genome Database SGD

  •  A yeast community effort.
  • A unique resource on all yeast genes and proteins and access to numerous datasets and curated literature.
  • A model for other organism-specific databases.
  • The Saccharomyces Genome Database (SGD) provides comprehensive integrated biological information for the budding yeast Saccharomyces cerevisiae along with search and analysis tools to explore these data, enabling the discovery of functional relationships between sequence and gene products in fungi and higher organisms.

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yeast Saccharomyces A model organism

The yeast Saccharomyces cerevisiae A model organism in genetics, genomics, and systems biology