Regulation of a transcription factor network by Cdk1 coordinates late cell cycle gene expression A, B Cells expressing differentially tagged WT TFs (TOS4‐3FLAG.

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Regulation of a transcription factor network by Cdk1 coordinates late cell cycle gene expression A, B Cells expressing differentially tagged WT TFs (TOS4‐3FLAG HCM1‐3HA YOX1‐3V5 YHP1‐13MYC), or phosphomutant TFs (4P, tos4‐9A‐3FLAG hcm1‐15A‐3HA yox1‐9A‐3V5 yhp1‐13A‐13MYC), were arrested in G1, released, and collected at 15‐min intervals for Western blot (A) and flow cytometry (B). Levels of TFs and cyclin‐dependent kinase 1 (Cdk1) are shown in (A). Quantification of proteins levels are shown in Supplementary Fig S5A. DNA content at each time point is shown (B, top). Overlay of WT (blue) and 4P (red) at 75 and 90 min highlight the mitotic delay in 4P cells (B, bottom). A representative of three replicate experiments is shown. C–G mRNA from WT and 4P cells collected as in (A) were compared to mRNA from asynchronous WT cells. Data from one of two biological replicates are shown. Average expression of all 6,237 genes at each time point in WT (blue) and 4P (red) cells (C). Average expression of 930 cell cycle‐regulated genes (D). For further explanation and breakdown of individual groups, see Supplementary Fig S5C. Average expression of 97 genes that have Hcm1‐binding motifs in their promoters and whose expression peaks during S‐phase (E) (Pramila et al, ). Average expression of 39 MCM cluster genes (F) (Spellman et al, ). Average expression of 31 CLB2 cluster genes (G) (Spellman et al, ). Data from biological replicates of all cell cycle‐regulated genes, and lists of genes in each cluster, are included in Supplementary Dataset S1. H RT‐qPCR of representative Hcm1 targets (CIN8, HTZ1) and MCM cluster genes (DBF2, KIN3) at the indicated time points after release from G1. For each gene, mean expression from three biological replicates, with standard deviations, is plotted. Asterisks indicate that all four comparisons are statistically significant with a P‐value of ≤ 0.01 (CIN8, P = 0.01; HTZ1, P = 0.005; DBF2, P = 0.007; KIN3, P = 0.003). Data information: In all parts, wild‐type cells are graphed in blue, 4P cells are red. Source data are available online for this figure. IF THIS IMAGE HAS BEEN PROVIDED BY OR IS OWNED BY A THIRD PARTY, AS INDICATED IN THE CAPTION LINE, THEN FURTHER PERMISSION MAY BE NEEDED BEFORE ANY FURTHER USE. PLEASE CONTACT WILEY'S PERMISSIONS DEPARTMENT ON PERMISSIONS@WILEY.COM OR USE THE RIGHTSLINK SERVICE BY CLICKING ON THE 'REQUEST PERMISSIONS' LINK ACCOMPANYING THIS ARTICLE. WILEY OR AUTHOR OWNED IMAGES MAY BE USED FOR NON-COMMERCIAL PURPOSES, SUBJECT TO PROPER CITATION OF THE ARTICLE, AUTHOR, AND PUBLISHER. EMBO J, Volume: 33, Issue: 9, Pages: 1044-1060, First published: 08 April 2014, DOI: (10.1002/embj.201386877)