Fig. 2. In the fwd1^RIP strains, FRQ is hyperphosphorylated, and the FRQ protein levels are higher due to increased protein stability. (A) Western blot analysis showing that, in the fwd1^RIP strains, FRQ was mostly hyperphosphorylated and its levels were higher than those of the wild-type strains. Cultures harvested in LL or DD29 were used. The arrows indicate the hyperphosphorylated FRQ species observed only in the fwd1^RIP strains. (B and C) Western blot analyses showing that FRQ was more stable in the fwd1^RIP strain after LD transition or cycloheximide (CHX) treatment (10 µg/ml). Cultures were first grown in LL for 1 day prior to the addition of CHX or were transferred into constant darkness and harvested at the indicated time. Densitometric analysis of the western blot results of (B) is shown in (C).