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. 2016 Sep 1;27(17):2735–2741. doi: 10.1091/mbc.E16-03-0192

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© 2016 Howard and Tansey. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

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FIGURE 3: — The effect of proteasome inhibition on Gcn4 activity is not mediated via the ArgR repressor or changes in nucleosome occupancy. (A) GCN4 ARG80 (GHY010), gcn4 ARG80 (GHY004), GCN4 arg80 (GHY081), and gcn4 arg80 (GHY079) strains were grown to log phase in minimal medium and treated with DMSO or MG132. After 1 h, strains were treated with DMSO or SM for 1.5 h and RNA harvested, and ARG1 mRNA levels were quantified by RT-qPCR. Each qPCR was normalized to the SM-treated GCN4 ARG80 sample. Error bars represent SEM. n = 4. (B) GCN4 yeast (GHY010) were grown to log phase at 30°C in minimal medium, treated with DMSO or MG132, and induced with SM or a DMSO control for an additional 1.5 h, and nucleosome occupancy was mapped by MNase digestion, coupled with tiled primer sets spanning ARG1. qPCR data were normalized to the signal from a GAL1-10 promoter-localized nucleosome. Data points represent an average of two independent experiments. (C) As in B, except monitoring nucleosome positioning surrounding the HIS4 locus. Data points represent an average of two independent experiments.