Sci. STKE, 5 August 2003
METHODOLOGY Find Your Partners
The yeast two-hybrid method is widely used as a screen for interactions of mammalian proteins, but yeast cells can differ from mammalian cells in critical ways. For example, yeast cells maintain very low concentrations of nitric oxide (NO), which is known to have regulatory functions in mammalian cells. Matsumoto et al. developed a modified two-hybrid screen that allowed them to hunt for NO-dependent protein interactions. They screened for partners that bound procaspase-3, a molecule that contributes to signals that cause cell death and whose activity is regulated by S-nitrosylation. Acid sphingomyelinase (ASM) was one of several proteins that showed NO-dependent association with procaspase-3. Endogenous ASM interacted with caspase-3 in a NO-dependent manner in cultured mammalian cells and inhibited activation of the protease. Thus, like phosphorylation, covalent modification of proteins by S-nitrosylation can acutely regulate cellular activity by altering interactions of proteins in signaling complexes.
Citation: Find Your Partners. Sci. STKE 2003, tw309 (2003).
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