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Dissecting glucose signalling with diversity-oriented synthesis and small-molecule microarrays

Abstract

Small molecules that alter protein function provide a means to modulate biological networks with temporal resolution. Here we demonstrate a potentially general and scalable method of identifying such molecules by application to a particular protein, Ure2p, which represses the transcription factors Gln3p and Nil1p1,2,3. By probing a high-density microarray of small molecules generated by diversity-oriented synthesis with fluorescently labelled Ure2p, we performed 3,780 protein-binding assays in parallel and identified several compounds that bind Ure2p. One compound, which we call uretupamine, specifically activates a glucose-sensitive transcriptional pathway downstream of Ure2p. Whole-genome transcription profiling and chemical epistasis demonstrate the remarkable Ure2p specificity of uretupamine and its ability to modulate the glucose-sensitive subset of genes downstream of Ure2p. These results demonstrate that diversity-oriented synthesis and small-molecule microarrays can be used to identify small molecules that bind to a protein of interest, and that these small molecules can regulate specific functions of the protein.

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Figure 1: The library synthesis and identification of uretupamine.
Figure 2: Studies in vivo, dose–response and structure–activity relationships of uretupamine.
Figure 3: Transcription profiling of treatment with uretupamine.
Figure 4: Glucose-sensitive signalling and a model of Ure2p function.

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Acknowledgements

We thank R. Melki for providing bacterially expressed Ure2p protein. F.G.K. was supported by the NIH Medical Scientist Training Program, A.F.S. by the Howard Hughes Medical Institute predoctoral fellowship, S.M.S. by the Roche and NSF predoctoral fellowships, and P.J.H. by the American Cancer Society. S.L.S. is an Investigator at the Howard Hughes Medical Institute. This research was funded by a grant from the NIGMS (GM-38627).

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Correspondence to Stuart L. Schreiber.

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Kuruvilla, F., Shamji, A., Sternson, S. et al. Dissecting glucose signalling with diversity-oriented synthesis and small-molecule microarrays. Nature 416, 653–657 (2002). https://doi.org/10.1038/416653a

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