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. 2020 May 8;11:2271. doi: 10.1038/s41467-020-16107-w

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© The Author(s) 2020

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 3 — a Schematic displaying how TFs and Tup1–Cyc8 could interact at the IME1 promoter. TF = transcription factor. b Diploid cells harbouring TUP1-AID and V5-tagged TFs (YAP6-V5, FW4214; SOK2-V5, FW4218; PHD1-V5, FW5056; MOT3-V5, FW4229; NRG1-V5, FW4230; NRG2-V5, FW5055; SKO1-V5, FW4224) were grown to exponential phase. As a control, TUP1-AID (FW5057) cells were also included, which also harbour a V5 tag. Cells were either treated with IAA or DMSO, and the binding of each transcription factor was determined by ChIP and normalised over HMR. Mean of n = 2 is shown. c Binding of Tup1 is affected in cells lacking transcription factor binding motifs in the IME1 promoter. Diploid cells with a chromosomal deletion of the IME1 locus, and an integrated plasmid that contained the full IME1 gene fused with sfGFP at the amino terminus and the wild-type promoter (pIME1-WT, FW5370) or the same construct with all the candidate motif sequences mutated (pIME1-bsΔ, FW5372) were used for the analysis. These cells also expressed TUP1-V5. The binding of Tup1 was determined by ChIP. Mean and SEM of n = 3 are shown. d Expression of Ime1 during entry into meiosis in pIME1-WT and pIME1-bsΔ cells. Strains described in c were grown till saturation in rich medium (YPD), grown for an additional 16–18 h in pre-sporulation medium (BYTA), and subsequently shifted to sporulation medium (SPO). The levels of Ime1 expression were determined by imaging and quantifying the fluorescent signals generated by sfGFP-Ime1 in single cells (n = 50 cells per sample). The mean of signals detected and error bars representing the 95% confidence interval are displayed. Unpaired parametric two-tailed Welch’s t test with 95% confidence was used and p values (** = ≤0.01, *** = ≤0.001) are indicated.