Fig. 1: Phosphorylation of the Hcm1 TAD regulates fitness.

a Schematic showing previously characterized CDK consensus sites (S/T-P), domains and predicted disordered regions. D phosphodegron, FKH forkhead domain, TAD transactivation domain. IUPred2 disorder prediction shown with phosphorylation sites in purple (D) and blue (TAD). b Western blot showing expression of the indicated proteins at the timepoints shown. Strains were initially grown in galactose (Gal) to allow expression of the 3HA-tagged WT protein from the genome and were shifted to dextrose (Dex) to repress WT expression at the start of the experiment. Plasmid-expressed Hcm1 proteins were detected with an antibody that recognizes a C-terminal 3V5 tag, PSTAIRE is shown as a loading control. Representative images from n = 3 biological replicates are shown. c–e Strains expressing the indicated HCM1 alleles from plasmids were co-cultured and the percentage of cells expressing each allele was determined at the timepoints indicated. In all, 5000 cells from each timepoint were analyzed. Black and gray represent cells expressing wild-type HCM1 (c–e), blue represents cells expressing hcm1-8A (d), and red represents cells expressing hcm1-8E (e). Shown is an average of n = 3 experiments, error bars represent standard deviations.