(A) Immunoblot analysis showing higher PRR5 protein accumulation in spy-3 mutant. Seedlings of PRR5:PRR5-GFP and PRR5:PRR5-GFP spy-3 were grown in 12 h L/12 h D photocycles. Samples were collected at indicated time points. PRR5-GFP protein abundance was detected with GFP antibody. Actin antibody and ponceau staining were used as loading controls.
(B) Quantitative analysis of PRR5 protein accumulation as shown in A with values normalized by corresponding Actin signals. Data are means ± SE from three biological replicates.
(C) PRR5 protein degradation is slower in spy-3 mutant. The cytoplasmic protein synthesis inhibitor, cycloheximide (CHX) was added at ZT9, and plants were harvested at 0, 2, 4 and 6 h after adding CHX, as indicated in the diagram. Control plants were harvested in parallel.
(D) Quantitative analysis of PRR5 protein abundance as shown in C. Data represent for means ± SE. from three individual biological replicates.