Expression of mPER1 protein in mouse model and transcriptional regulation of mPer2

Abstract

Many physiological and behavioral activities exhibit 24-hour periodic rhythms. These rhythms exist in many organisms, prokaryote to human, and affect on many functions, basic cellular functions to higher neural activities. Organism exhibits the circadian rhythm with intrinsic cell-autonomous clock which is entrained by day and night cycle. The cell-autonomous clock consists with genetically based autoregulatory feed-back loop mechanisms. mPerl and mPer2 are major player of the feed-back loop. They contain PAS domains for dimerization and their mRNA and protein are rhythmically expressed. To understand the oscillation of the circadian clock in vivo, real-time reporting system for mPerl and mPer2 were developed in this study. By using the gene targeting method, mPerl:Luc knock-in ES cells were generated. For the dual monitoring purpose two different types of luciferase which are fire fly luciferase and rail road warm luciferase were fused at the end of exon 23. Various length of mPer2:: Luc:Neo constructs were stably transfected fibroblasts as a transcriptional model of mPer2. By means of real-time recording of bioluminescence, inducible expression of mPER1 in mPerl::Luc knock-in ES cell and oscillation of luciferase in various truncated mPerl::Luc reporter construsts were assayed. From the mPer2:: Luc transcriptional model, presence of multiple cis-acting elements which regulate phase, and robustness of oscillation of mPer2 was suggested.