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. 2014 Oct 21;9(10):e109693. doi: 10.1371/journal.pone.0109693

Figure 1. Rhythmic expression of Per2 in Caki-2 cells.

Figure 1

(A) All renal cancer cell lines were transfected with the Per2 promoter reporter (2 µg) and the bioluminescence was then measured using a real-time monitoring assay. Real-time monitoring of luciferase activity of the Per2 promoter showed that activity oscillated over an approximately 24-h cycle. The luciferase activities of four replicate samples are shown. These cultures showed significant circadian rhythms (Table 1). (B) mRNA levels of Per2 were determined by real-time PCR for six plates at each time point. Total RNA was extracted every 4 h, beginning 24 h after treatment with dexamethasone for one 24-h cycle, and Per2 transcripts were quantified. Error bars indicate the standard errors of the mean values (n = 6). The data from a single 24 hours after dexamethasone treatment were analyzed using the Cosinor software for rhythmicity (Table 1). (C) The structure of the Per2 promoter and an analysis of the potential transcription factor-binding motifs in this region. The 2,994-bp region contains one E-box-like sequence (CACGTT) and one HRE-like sequence (ATGTG), similar to the consensus HRE sequence (ACGTG) located upstream of the transcription start site (TSS). (D) Sequence comparisons: upper line, mouse sequence; lower line, human sequence. The nucleotide sequence of potential transcription factor-binding motifs for E-box-like sequence and HRE-like sequence are 100% conserved between mouse and human.