Yeast as a tool to identify anti-aging compounds

Abstract

In the search for interventions against aging and age-related diseases, biological screening platforms are indispensable tools to identify anti-aging compounds among large substance libraries. The budding yeast, Saccharomyces cerevisiae, has emerged as a powerful chemical and genetic screening platform, as it combines a rapid workflow with experimental amenability and the availability of a wide range of genetic mutant libraries. Given the amount of conserved genes and aging mechanisms between yeast and human, testing candidate anti-aging substances in yeast gene-deletion or overexpression collections, or de novo derived mutants, has proven highly successful in finding potential molecular targets. Yeast-based studies, for example, have led to the discovery of the polyphenol resveratrol and the natural polyamine spermidine as potential anti-aging agents. Here, we present strategies for pharmacological anti-aging screens in yeast, discuss common pitfalls and summarize studies that have used yeast for drug discovery and target identification.

Figure 1.

Advantages and limitations of yeast for anti-aging drug discovery. The budding yeast, S. cerevisiae, offers a genetically well-defined cellular environment paired with a fast and economic experimental workflow, including a variety of mutant libraries for drug target identification. Due to its unicellular nature, studying tissue-specific responses might be limited. Other constraints are limited drug metabolization and suboptimal permeability of the fungal cell wall to xenobiotics. See the main text for further details.

Figure 2.

Screening strategies for chemogenomic drug discovery. (A) Forward chemical genetics (FCG) start with a compound library, which is administered to wild type cells or engineered yeast cells expressing a protein of interest. Hits are selected based on a phenotypic readout, ending up with bioactive compounds. Targets can be identified by treating genetic libraries (loss-of-function or gain-of-function mutants) with the identified hits and reversal of the phenotype as a readout. (B) Reverse chemical genetics (RCG) start with the selection of a target of interest, which is treated with a compound library in vitro, using ligand binding or enzymatic activity as readouts. Wild type cells or genetic libraries are treated with the selected hits to identify a phenotype and/or target.

Figure 3.

Yeast as a pharmacological screening tool in forward chemical genetics. Compound libraries can be tested in wild type yeast in an unbiased manner, or in semi-biased approaches in engineered/humanized yeast strains. To investigate specific pathways which do not result in an apparent phenotype, yeast can be manipulated to translate pathway activities into growth behavior. For instance, a promoter that is regulated by the pathway of interest can be linked to an essential gene (e.g. URA3). The growth on uracil-free or 5-fluoroorotic acid (5-FOA) containing media is an indirect measure for increased or decreased pathway activity, respectively. Humanized yeast, expressing human disease-relevant genes can be used to find compounds that specifically interfere with the function of the transgene. Compounds derived from unbiased screens should be validated in multicellular models before screening for potential targets in different yeast mutant libraries. Among others these libraries include heterozygous or haploid deletion as well as overexpression collections for different genes (Gene XY). In addition, de novo derived mutants by spontaneous (*) or transposon (Tn7) mutagenesis can be used to identify loci associated with the compound's mechanism of action.

Figure 4.

Versatile transferability of yeast phenotypical screens. Yeast acts as a cellular sieve for substances which cannot pass the plasma membrane, are exported by pleiotropic drug efflux pumps (PDRs), or generally exhibit no biological activity. Chemicals toxic to yeast can be tested in human cell culture for cytotoxicity. Substances which specifically kill cancer cells are potential chemotherapeutics, while substances with no toxicity might be further tested in pathogenic fungi as potential anti-fungal drugs. Lifespan-extending compounds should be validated in multicellular model organisms (nematodes, flies, mice). Substances which only extend yeast lifespan may be considered for biotechnological applications (e.g. yeast bioreactors) where yeast survival is desired. Chemicals which inhibit growth can be further tested in both directions (e.g. when they inhibit nutrient signaling), but are less suitable for biotechnological use.