Figure 7.

PER is phosphorylated preferentially in the nucleus. A, NLS-tagged PER (PER-NLS) is predominantly localized in the nucleus. S2R+ cells were transiently transfected with wild-type per or per tagged with NLS. Cells were fixed and stained with antibody to PER, and the localization of PER was examined by confocal microscopy. Scale bar, 20 μm. B, PER-NLS is hyperphosphorylated. S2R+ cells were transiently transfected with wild-type per or per tagged with NLS. Transfected cells were lysed with or without phosphatase. The resulting lysates were examined by immunoblot analysis using indicated antibodies. Some lysates were incubated with λ-phosphatase (New England Biolabs) for 30 min at 37°C, before gel analysis.