Transcriptional activation assay of constructs containing the LexA-DBD fused to the Hcm1 TAD with the indicated phosphosite mutations or WT or empty vector (EV) controls. Black represents EV, gray represents WT, blue represents hcm1-8A , red represents hcm1-8E , and shades of green represent mutants with potential T priming sites mutated to S. Error bars represent standard deviation, shown is an average of n = 3 replicates. Significance was tested using repeated measures one-way ANOVA with Dunnett’s multiple comparisons test. Asterisks indicate significantly different from WT, **** P < 0.0001, *** P < 0.0005, ** P < 0.005, * P < 0.05. EV, AAAAAAAA P < 0.0001; SSSSWWWW P = 0.0064; SSSWWWWW P = 0.0154; WWWSWWWW P = 0.0005; EEEEEEEE P < 0.0001. Supplementary Fig. 4c confirms that all fusion proteins are expressed at similar levels.
