Chapter title |
A yeast-based high-throughput screen for modulators of phosphodiesterase activity.
|
---|---|
Chapter number | 14 |
Book title |
cAMP Signaling
|
Published in |
Methods in molecular biology, January 2015
|
DOI | 10.1007/978-1-4939-2537-7_14 |
Pubmed ID | |
Book ISBNs |
978-1-4939-2536-0, 978-1-4939-2537-7
|
Authors |
Ana Santos de Medeiros, Charles S Hoffman, Charles S. Hoffman |
Abstract |
Cell-based high-throughput screens (HTSs) targeting heterologously expressed proteins in yeast identify compounds that often display relevant biological activity when tested in cell culture. We developed a fission yeast-based HTS to detect small-molecule inhibitors of mammalian cyclic nucleotide phosphodiesterases (PDEs). These screens are carried out in Schizosaccharomyces pombe using a PKA-repressed fbp1-ura4 reporter whose expression due to low PKA activity prevents cells from growing in medium containing the pyrimidine analog 5-fluoro orotic acid (5FOA). We describe here the steps required to construct strains for screening and to optimize conditions for successful screens. |
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