Abstract
The finding that the well-known mammalian P53 protein can act as a transcription factor (TF) in the yeast S. cerevisiae has allowed for the development of different functional assays to study the impacts of 1) binding site [i.e., response element (RE)] sequence variants on P53 transactivation specificity or 2) TP53 mutations, co-expressed cofactors, or small molecules on P53 transactivation activity. Different basic and translational research applications have been developed. Experimentally, these approaches exploit two major advantages of the yeast model. On one hand, the ease of genome editing enables quick construction of qualitative or quantitative reporter systems by exploiting isogenic strains that differ only at the level of a specific P53-RE to investigate sequence-specificity of P53-dependent transactivation. On the other hand, the availability of regulated systems for ectopic P53 expression allows the evaluation of transactivation in a wide range of protein expression. Reviewed in this report are extensively used systems that are based on color reporter genes, luciferase, and the growth of yeast to illustrate their main methodological steps and to critically assess their predictive power. Moreover, the extreme versatility of these approaches can be easily exploited to study different TFs including P63 and P73, which are other members of TP53 gene family.
| Original language | English |
|---|---|
| Article number | e59071 |
| Journal | Journal of Visualized Experiments |
| Volume | 2019 |
| Issue number | 150 |
| DOIs | |
| Publication status | Published - Aug 1 2019 |
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Keywords
- Cancer Research
- Functional assay
- Growth inhibition
- Issue 150
- P53
- Response element
- Transcription
- Yeast
ASJC Scopus subject areas
- Neuroscience(all)
- Chemical Engineering(all)
- Biochemistry, Genetics and Molecular Biology(all)
- Immunology and Microbiology(all)
Cite this
Yeast as a chassis for developing functional assays to study human P53. / Monti, Paola; Bosco, Bartolomeo; Gomes, Sara; Saraiva, Lucilia; Fronza, Gilberto; Inga, Alberto.
In: Journal of Visualized Experiments, Vol. 2019, No. 150, e59071, 01.08.2019.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Yeast as a chassis for developing functional assays to study human P53
AU - Monti, Paola
AU - Bosco, Bartolomeo
AU - Gomes, Sara
AU - Saraiva, Lucilia
AU - Fronza, Gilberto
AU - Inga, Alberto
PY - 2019/8/1
Y1 - 2019/8/1
N2 - The finding that the well-known mammalian P53 protein can act as a transcription factor (TF) in the yeast S. cerevisiae has allowed for the development of different functional assays to study the impacts of 1) binding site [i.e., response element (RE)] sequence variants on P53 transactivation specificity or 2) TP53 mutations, co-expressed cofactors, or small molecules on P53 transactivation activity. Different basic and translational research applications have been developed. Experimentally, these approaches exploit two major advantages of the yeast model. On one hand, the ease of genome editing enables quick construction of qualitative or quantitative reporter systems by exploiting isogenic strains that differ only at the level of a specific P53-RE to investigate sequence-specificity of P53-dependent transactivation. On the other hand, the availability of regulated systems for ectopic P53 expression allows the evaluation of transactivation in a wide range of protein expression. Reviewed in this report are extensively used systems that are based on color reporter genes, luciferase, and the growth of yeast to illustrate their main methodological steps and to critically assess their predictive power. Moreover, the extreme versatility of these approaches can be easily exploited to study different TFs including P63 and P73, which are other members of TP53 gene family.
AB - The finding that the well-known mammalian P53 protein can act as a transcription factor (TF) in the yeast S. cerevisiae has allowed for the development of different functional assays to study the impacts of 1) binding site [i.e., response element (RE)] sequence variants on P53 transactivation specificity or 2) TP53 mutations, co-expressed cofactors, or small molecules on P53 transactivation activity. Different basic and translational research applications have been developed. Experimentally, these approaches exploit two major advantages of the yeast model. On one hand, the ease of genome editing enables quick construction of qualitative or quantitative reporter systems by exploiting isogenic strains that differ only at the level of a specific P53-RE to investigate sequence-specificity of P53-dependent transactivation. On the other hand, the availability of regulated systems for ectopic P53 expression allows the evaluation of transactivation in a wide range of protein expression. Reviewed in this report are extensively used systems that are based on color reporter genes, luciferase, and the growth of yeast to illustrate their main methodological steps and to critically assess their predictive power. Moreover, the extreme versatility of these approaches can be easily exploited to study different TFs including P63 and P73, which are other members of TP53 gene family.
KW - Cancer Research
KW - Functional assay
KW - Growth inhibition
KW - Issue 150
KW - P53
KW - Response element
KW - Transcription
KW - Yeast
UR - http://www.scopus.com/inward/record.url?scp=85070827874&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85070827874&partnerID=8YFLogxK
U2 - 10.3791/59071
DO - 10.3791/59071
M3 - Article
AN - SCOPUS:85070827874
VL - 2019
JO - Journal of Visualized Experiments
JF - Journal of Visualized Experiments
SN - 1940-087X
IS - 150
M1 - e59071
ER -