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Analysis of Period 2-containing protein complexes in mouse fibroblasts

Laufzeit: 01.01.2007 - 31.12.2008

Kurzfassung


According to current models, the mammalian circadian clock consists of interwoven positive and negative feedback loops. At the core of this oscillator lie the PERIOD (Per) and CRYPTOCHROME (Cry) proteins, which act to rhythmically inhibit the heterodimeric transcription factor CLOCK:BMAL1. To gain insight into the precise molecular mechanisms of how this transcriptional inhibition occurs, we have established fibroblast cell lines that express a TAP-tagged version of the Period 2 (Per2)...According to current models, the mammalian circadian clock consists of interwoven positive and negative feedback loops. At the core of this oscillator lie the PERIOD (Per) and CRYPTOCHROME (Cry) proteins, which act to rhythmically inhibit the heterodimeric transcription factor CLOCK:BMAL1. To gain insight into the precise molecular mechanisms of how this transcriptional inhibition occurs, we have established fibroblast cell lines that express a TAP-tagged version of the Period 2 (Per2) protein, and have purified a Per2-containing protein complex from these cells. Preliminary analysis has led to the identification of several known interaction partners and factors implicated in the circadian clock (Per1, Cry1, Cry2, casein kinase 1 delta and epsilon). Hence, we expect to identify additional bona fide interaction partners of Per2 by mass spectrometric analysis of the purified protein complexes. Such potential new components of the circadian clock will subsequently be subjected to rigorous analysis for a role in the oscillator, using a wide array of assays available in the lab, both in vitro and in vivo (tissue culture, mice).» weiterlesen» einklappen

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