Screening yeast-based

An example of a TIE approach is that described by Desbrow et al. [7]. In this work, the endocrine disrupting activity detected in effluents of seven UK WWTPs by means of a yeast-based screening assay [52] was mainly attributed to the presence of estradiol, estrone, and ethynylestradiol. However, to assess the estrogenic activity different bioassays may be used, e.g., the yeast-based recombinant estrogen receptor-reporter assay (YES), the MCF-7 cell proliferation (E-screen), and the estrogen receptor-mediated chemically activated... 

FIGURE 5.5 Yeast-based screen for inhibitors of human PDE IV. A PDE-deficient yeast (PMY) will not utilize acetate as a sole carbon source and is sensitive to heat shock (55°C). Complementation with a human type IV PDE (PMY + PDE) expressed from a copper-dependent (CUPl) promoter reverses the mutant phenotype. Addition of type IV PDE inhibitor (rolipram) to the complemented yeast restores the mutant phenotype (PMY + PDE + rolipram). 

Fig. 8.3 Yeast-based screen for agonists or antagonists of the human estrogen receptor. The hormone estrogen (estradiol) binds to the estrogen receptor which is expressed from a gene driven by the PGK promoter. The hormone-receptor complex binds to an estrogen-responsive element (ERE) that controls the expression of the p-galactosidase reporter gene. The assay measures the activity of the enzyme using a substrate that forms a colored product on conversion.

Fig. 8.5 Yeast-based screen for 32-agonists. The Pa-adrenergic receptor (B Ar) expressed from a GALl promoter links to the mating type response via the human Gsa-subunit expressed off the CUP1 promoter, by complementation of GPA-1. The detection of signaling is by induction of the FUS promoter linked to a p-galactosidase reporter gene.

Example 2 In another recent approach to search for autophagy modulators, Rubinsztein and Schreiber used a yeast-based screen to search for small molecules... 

Fig. 2 Yeast-based functional screen for receptor-independent activators of G-protein signaling. The yeast strain indicated in A was generated as described previously (Cismowski et al. 1999, 2002) and transformed with mammalian cDNA libraries in the galactose-inducible yeast expression vector pYES2. The strategy for identifying mammalian cDNAs that promoted growth (i.e., activated G-protein signaling pathway) is indicated in B. GALlp GALl promoter)...

Fig. 3 A AGS proteins isolated in yeast-based functional screen G-protein signaling modulator (GPSM) as named by the Human Genome Nomenclature Committee. The major sites of action of AGS proteins in the context of the G-protein activation-deactivation cycle are indicated B on the right. (GPR G-protein regulatory, GEF guanine nucleotide exchange factor, GDI guanine nucleotide dissociation inhibitor, GPCR G-protein coupled receptor)...

Key Words Ras Raf yeast two-hybrid system protein interaction compound library cell-based screening. 

Of interest is a recently described yeast-based, nutrient-dependent viability screen for inhibitors of protozoal dihydrofolate reductase (DHFR) [43,44], Antiprotozoal activity of DHFR inhibitors is well known, and DHFR- yeast complemented with the DHFR gene derived from the malaria parasite P. falciparum have been used to characterize the molecular pharmacology of resistance to the antimalarial DHFR inhibitors pyrimethamine and cycloguanil [45,46], The subsequent development of a screen was based on the demonstration that the protozoal... 

Selection and screening methods are effective tools for shidying macromolecular interactions. Valuable methods are the yeast-based one-hybrid and two-hybrid systems (for smdy-ing protein-DNA and protein-protein interactions, respectively) and bacterial-based phage display methods (for studying either type of interaction). These systems have been used to... 

---