Biomedical Engineering Reference
In-Depth Information
pathogens mainly associated with fungal infections belong to the
Candida
(yeast) and
Aspergillus
(mold) species. Given their pathogenicity and our poor knowledge of their
genomes, metabolisms, and phenotypes, the modeling of such fungi with
S. cerevisiae
provided a fundamental contribution to the identification of novel antifungal drugs on
several occasions. As for pathogenic fungi, knowledge of the genome, metabolism,
and phenotype for
Homo sapiens
is too poor to exploit human cells directly for drug
discovery purposes. Moreover, human cells are hardly cultivable, and the tools for
their genetic and proteicmanipulation are limited. Once again,
S. cerevisiae
was found
to be a good model. The most renowned examples of the insights on mammalian cells
obtained by means of the yeast model concerns studies on cancer. Given its genetic
similarity with higher eukaryotic cells, this microorganism has been used to deduce
functional and mechanistic aspects of eukaryotic proteins and protein systems. In
addition, heterologous expression of human proteins in
S. cerevisiae
is possible, and
this approach allows for both the identification of mutations associated with diseases
and the functional characterization of aberrant human enzyme functions. In 1993,
Strand et al. [19] predicted that mutations in the human orthologs of three yeast genes
(
MSH2
,
MLH1
, and
PML1
) might be themain factors causing hereditary nonpolyposis
colorectal cancer (HPNCC). This indication was confirmed later by Fishel et al. [20],
who proposed that mutations in either hMSH2 (the ortholog of the yeast gene
MSH2
)
or hMLH1 underlie the vast majority of cases of HPNCC. Clonal complementation
has also been used to identify human protein functions. Expression of human cDNA
in
S. cerevisiae
allowed for the selection of clones expressing human genes able to
complement mutations that occurred on a selected yeast gene. In a pioneering study
using this approach, Lee and Nurse identified a human homolog (POLD1) of the
yeast
CDC2
gene involved in cell cycle progression and capable of complementing
mutations in the microorganism gene [21].
The vast majority of the screenings based on the use of
S. cerevisiae
used only
one genetic background, that of the laboratory strain S288c. Nevertheless, the use
of this strain is limited by the accumulation of genetic defects in this strain, such as
the insertion of a transposon in the gene
HAP1
, making the strain unable to sustain
proper respiratory metabolism [22]. The effects on the yeast metabolic asset given by
these genetic defects could mask the effect of a given perturbation (i.e., forbidding the
identification of compounds modulating respiration). To prevent such a limitation,
parallel use of the widely used S288c yeast strain and of a second deeply characterized
laboratory strain, W303, was proposed, having 98% genetic identity with S288c
but able to sustain proper respiratory metabolism since bearing a functional
HAP1
gene [23].
All the aforementioned qualities make yeast the best model for studies on fun-
damental cellular processes of relevance also in higher eukaryotic cells, and for
the identification of determinants of stress and drug response [24]. Gene expres-
sion in yeast cells (wild-type or mutant), measured using DNA microarrays, helps
to identify both the primary target of stimuli/compounds and any secondary con-
sequences. The primary output is a quantitative assessment of the changes in gene
expression for almost every gene in the yeast genome. The effects on gene expres-
sion resulting from chemical (compound/stimulus) and genetic (gene mutation or
