The Pathways over Time Project
A one-semester research project in comparative functional genomics
Cysteine and methionine are superimposed over a portion of “The Tree
of Life” by Gustav Klimt (1909)
Methionine and cysteine are sulfur-containing amino acids
found in the proteins of all living organisms
The metabolic pathways for synthesizing methionine and
cysteine have changed during evolution
Genome sequencing projects offer opportunities for undergraduate
research
Over 40,000 completed genomes as of
July 15, 2014
Primarily microbial sequences
Genome annotation projects predict enzyme function
based on sequence similarities to known enzymes.
Functional testing is usually missing.
This semester-long research project uses
Saccharomyces cerevisiae, the budding yeast, to
analyze the conservation of enzyme function
under the microscope
Nonpathogenic model
organism
Yeast!
in its natural environment
in our kitchens
Spanish "cerveza"
Swahili "pombe"
Yeast research has a long history
Empirical research dominated for millenia
Louis Pasteur shows that fermentation is
due to the activities of microorganisms
1866 – “Etudes sur le Vin”
Eduard Buchner shows that a cell-free
extract of yeast is able to ferment sugar
“enzyme = from yeast”
1907 – wins Nobel Prize
Ability to alternate between haploid and diploid forms made yeast an favored
organism for geneticists
Haploids and some diploids can propagate by division (1)
When stressed, haploids of opposite mating types conjugate (2) to form diploids
Diploids generally enter meiosis and form hardy, haploid spores (3)
Spores are
contained within
an ascus
Why are yeast considered model organisms?
Model organisms have many of the same processes as more
complex life forms
Model organisms have many experimental advantages
~6000 genes
~20,000-25,000 genes
Haploid and diploid forms
Diploid
Small (~4 µm) and unicellular
Large and multi-cellular
New generation every 1.5 hr
New generation every 20 yr
Simple growth requirements
Complex growth requirements
Few genes have introns
Many introns –complex splicing
Many mutant phenotypes
Phenotypes are complex
Genetic manipulation possible
Ethical issues preclude
experimentation
Yeast are able to synthesize methionine de novo
Also MET
1, 8, 7 and
13
Over 20 S. cerevisiae genes
encode proteins involved in
methionine and cysteine
synthesis (not all appear here)
Genes were identified in
genetic screens and
confirmed with molecular
methods
We will use S. cerevisiae
strains with targeted deletions
in MET and CYS genes to
analyze foreign gene function
More later……..
The strategy
Can genes from distant
organisms replace genes
that are inactivated in
budding yeast?
(credits: Tree of Life
Web project)
1. Identify potential genes
by their similarity to known
yeast genes using
bioinformatics databases
2. Clone the foreign genes
into plasmids that drive
overexpression in S.
cerevisiae
3. Express the
foreign genes in S.
cerevisiae and see if
they can compensate
for missing MET
genes
S. cerevisiae and S. pombe are yeast species that separated from
a common ancestor from 0.3-1 billion years ago
Unicellular eukaryotes
Members of the ascomycota phylum of fungi
Fission yeast
Budding yeast
Difference in lifestyle:
One buds, the other divides
to produce two equal
offspring
Shared a common ancestor
~1 billion years ago
Saccharomyces cerevisiae
(sugar fungus found in beer)
Schizosaccharomyces
pombe
Ascomycota: spores are contained within an ascus
Considerable genome remodeling has occurred since the species diverged
whole genome duplication
Genome
size (Mbp)
14 Mbp
28 Mbp
9.4 Mbp
39 Mbp
32 Mbp
30 Mbp
12.6 Mbp
You will be building on work from previous years
Students have demonstrated
conservation of several MET
gene between S. pombe and S.
cerevisiae
Let’s hope for some great results!
Poster winners participate in the Biology Dept.’s Undergraduate
Research Symposium (first study day in May)