Cell based assay for identifying sh2-domain-specific signal transducer antagonist

A method for identifying the presence in a test composition of an inhibitor of the binding of a phosphopeptide binding pair comprising either a phosphatase domain and a ligand therefor or a phosphotyrosine binding domain and a ligand therefor, which method comprises the steps of:

• (a) providing mammalian cells which contain and are capable of expressing recombinant DNA molecules encoding two fusion proteins, the first of which comprising one or more copies of each of a transcription activation domain and one member of the phosphopeptide binding pair, the second of which comprising one or more copies of each of a DNA-binding domain and the other member of the phosphopeptide binding pair, wherein the cells
• (i) further contain a reporter gene which encodes a detectable gene product and is linked to a DNA sequence permitting expression of the reporter gene in the presence of the pair of fusion proteins, and
• ii) when cultured are capable of producing measurable amounts of said detectable gene product,
• (b) culturing said cells in the presence and absence of the test composition under conditions in which, in the absence of an inhibitor of the binding of the phosphopeptide binding pair they produce measurable amounts of said detectable gene product, and
• (c) determining whether production of the detectable gene product was diminished in the presence of the test composition which would indicate the presence of an inhibitor.