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. 2005 Mar;25(5):1912–1921. doi: 10.1128/MCB.25.5.1912-1921.2005

FIG. 5.

FIG. 5.

Cyclic mPER2 protein accumulation accompanies the ubiquitin-proteasome-mediated degradation. (A) Serum shock-induced circadian gene expressions in the FLAG-tagged mPer2 P2-#32 cell line using the Tet-Off system. Under Dox+ conditions, expression of mPer2 mRNA was very faint at all points examined. Under Dox− conditions, higher levels of mPer2 mRNA were constitutively expressed without showing circadian change. Under both conditions, the expression of dbp mRNA exhibited clear circadian oscillation. en, endogenous mPer2 mRNA; ex, exogenous mPer2 mRNA. (B) FLAG-mPER2 was immunoprecipitated (IP) with anti-FLAG antibodies, and precipitates were examined by Western blot (WB) analysis using anti-mPER2 antibodies in the P2-#32 cell line without doxycycline after serum shock. The dense mPER2 accumulation found at 6 h became faint at 18 h after serum shock. Application of MG132 inhibited the reduction of mPER2 protein at 18 h. (C) Quantitative analysis of the expression of FLAG-mPER2 described above (B). IgG, immunoglobulin G.