The role of a protein disulfide isomerase in the circadian clock of Chlamydomonas reinhardtii
Anna Filonova and Volker Wagner
Institute of General Botany and Plant Physiology, Friedrich-Schiller-University Jena, Am Planetarium 1, 07743 Jena, Germany
Circadian rhythms are defined as biological rhythms that persist under constant conditions of light and temperature with a period of about 24 h. In Chlamydomonas reinhardtii several processes are regulated by the circadian clock, including phototaxis, chemotaxis or cell division. With a combination of heparin affinity chromatography, two-dimensional gel electrophoresis and mass spectrometry, we could identify a protein disulfide isomerase (CrPDI2) that is specifically enriched by heparin in samples taken during subjective night [1,2]. In immunoblots using anti-peptide antibodies, we revealed a higher circadian change in abundance of the heparin bound CrPDI2 than in crude extracts, suggesting that CrPDI2 may have some basic interaction partners during night phase. Indeed, CrPDI2 is present in protein complexes with an average apparent molecular mass of ~ 158 kDa. In transgenic strains that overexpress CrPDI2, we have analyzed changes in phase or period of the circadian clock using the automated rhythm of phototaxis as indicator. To start the functional characterization of the CrPDI2, we will determine the enzymatic properties of the recombinant protein. Furthermore, we will address novel aspects of the regulatory mechanism of posttranslational control of CrPDI2 via protein-protein interaction and redox signaling in the circadian clock of C. reinhardtii.
[1] Wagner et al. (2004) FEBS Letters 559, 129-135 [2] Wagner and Mittag (2009) Methods Mol Biol. 479, 1-16.
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