Figure 4. Genetically increasing casein kinase I (CKI) levels or activity rescues the Δprd-2 long period phenotype.

Representative race tubes (RTs) from ras-1^bd P_qa-2-ck-1a single (pink) and ras-1^bd P_qa-2-ck-1a Δprd-2 double (yellow) mutants are shown with growth using the indicated concentrations of quinic acid (QA) to drive expression of ck-1a. All results are shown in a scatterplot, where each dot represents one RT’s free running period length. ras-1^bd controls (black) had an average period of 22.5 ± 0.5 hr (N = 12), and period length was not significantly affected by QA concentration (ANOVA p=0.297). ras-1^bd Δprd-2 controls (blue) had an average period of 25.4 ± 0.4 hr (N = 10), and period length was not significantly affected by QA concentration (ANOVA p=0.093). Period length of ras-1^bd P_qa-2-ck-1a single mutants (pink) was significantly altered across QA levels (ANOVA p=3.6 × 10⁻⁶), and the average period at 10⁻¹ M QA was 24.3 ± 0.5 hr (N = 4). Period length of ras-1^bd P_qa-2-ck-1a Δprd-2 double mutants (yellow) was also significantly affected by QA levels (ANOVA p=8.1 × 10⁻⁸), and the average period at 10⁻¹ M QA was 25.4 ± 0.4 hr (N = 4). The double mutant period length was not genetically additive at high levels of QA induction (A). A hyperactive CKI allele was constructed by expressing the shortest isoform only (CKI^SHORT). 96-well plate luciferase assays were used to measure the circadian period length. Traces represent the average of three technical replicates across four biological replicate experiments for: ras-1^bd controls (gray, τ = 21.7 ± 0.3 hr), ras-1^bd Δprd-2 (blue, τ = 25.7 ± 0.6 hr), ras-1^bd CKI^SHORT (pink, τ = 17.4 ± 0.3 hr), and ras-1^bd CKI^SHORT Δprd-2 double mutants (yellow, τ = 18.2 ± 0.3). CKI^SHORT is completely epistatic to Δprd-2 in double mutants (B).