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Published on: December 21, 2011
A functional yeast survival screen of tumor-derived cDNA libraries designed to identify anti-apoptotic mammalian
Moritz Eißmann1, Bettina Schwamb, Inga Maria Melzer
1Chemotherapeutisches Forschungsinstitut Georg-Speyer-Haus, Frankfurt/Main, Germany.
Abstract:
Yeast cells can be killed upon expression of pro-apoptotic mammalian proteins. We have established a functional yeast survival screen that was used to isolate novel human anti-apoptotic genes overexpressed in treatment-resistant tumors. The screening of three different cDNA libraries prepared from metastatic melanoma, glioblastomas and leukemic blasts allowed for the identification of many yeast cell death-repressing cDNAs, including 28% of genes that are already known to inhibit apoptosis, 35% of genes upregulated in at least one tumor entity and 16% of genes described as both anti-apoptotic in function and upregulated in tumors. These results confirm the great potential of this screening tool to identify novel anti-apoptotic and tumor-relevant molecules. Three of the isolated candidate genes were further analyzed regarding their anti-apoptotic function in cell culture and their potential as a therapeutic target for molecular therapy. PAICS, an enzyme required for de novo purine biosynthesis, the long non-coding RNA MALAT1 and the MAST2 kinase are overexpressed in certain tumor entities and capable of suppressing apoptosis in human cells. Using a subcutaneous xenograft mouse model, we also demonstrated that glioblastoma tumor growth requires MAST2 expression. An additional advantage of the yeast survival screen is its universal applicability. By using various inducible pro-apoptotic killer proteins and screening the appropriate cDNA library prepared from normal or pathologic tissue of interest, the survival screen can be used to identify apoptosis inhibitors in many different systems.
Insights
Researchers developed a yeast survival screen to find new human anti-apoptotic genes overexpressed in tumors. This tool identified known and novel genes, including MAST2, crucial for glioblastoma growth, offering therapeutic potential.
Area of Science:
- Molecular Biology
- Cancer Research
- Yeast Genetics
Background:
- Yeast cells can undergo apoptosis when expressing pro-apoptotic mammalian proteins.
- Identifying anti-apoptotic genes is crucial for understanding tumor resistance and developing therapies.
Purpose of the Study:
- To establish and utilize a functional yeast survival screen for isolating novel human anti-apoptotic genes.
- To identify genes overexpressed in treatment-resistant tumors that inhibit apoptosis.
- To evaluate the therapeutic potential of identified anti-apoptotic genes.
Main Methods:
- Functional yeast survival screen using cDNA libraries from metastatic melanoma, glioblastomas, and leukemic blasts.
- Analysis of isolated cDNAs for anti-apoptotic function and tumor relevance.
- In vitro cell culture assays and in vivo xenograft mouse models to validate candidate genes (PAICS, MALAT1, MAST2).
Main Results:
- The screen identified numerous yeast cell death-repressing cDNAs, including known anti-apoptotic genes and genes upregulated in tumors.
- 16% of identified genes were both anti-apoptotic and tumor-upregulated.
- PAICS, MALAT1, and MAST2 demonstrated anti-apoptotic functions and were overexpressed in specific tumor types.
- MAST2 expression was essential for glioblastoma tumor growth in a mouse model.
Conclusions:
- The yeast survival screen is a powerful and versatile tool for discovering novel anti-apoptotic and tumor-relevant molecules.
- Identified genes like MAST2 represent potential therapeutic targets for molecular cancer therapy.
- The screen's applicability extends to various biological systems by adapting pro-apoptotic triggers and cDNA libraries.
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