Molecular Plant Advance Access originally published online on June 7, 2007
Molecular Plant 2008 1(1):4-14; doi:10.1093/mp/ssm002
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© The Author 2007. Published by Oxford University Press on behalf of CSPP and IPPE, SIBS, CAS.
Chemically Induced and Light-Independent Cryptochrome Photoreceptor Activation
a FB Biologie-Pflanzenphysiologie, Philipps-Universität, Karl-von-Frisch-Str. 8, 35032 Marburg, Germany
b Universität Dortmund, FB Chemie-Chemische Biologie, Otto-Hahn-Str. 6, 44227 Dortmund, Germany
c University of Tennessee, Department of Biochemistry, Cellular and Molecular Biology, Walters Life Sciences M407, Knoxville, TN 37996-0840, USA
1 To whom correspondence should be addressed. E-mail batschau{at}staff.uni-marburg.de, fax 49-(0)6421-282-1545.
The cryptochrome photoreceptors of higher plants are dimeric proteins. Their N-terminal photosensory domain mediates dimerization, and the unique C-terminal extension (CCT) mediates signaling. We made use of the human FK506-binding protein (FKBP) that binds with high affinity to rapamycin or rapamycin analogs (rapalogs). The FKBP–rapamycin complex is recognized by another protein, FRB, thus allowing rapamycin-induced dimerization of two target proteins. Here we demonstrate by bioluminescence resonance energy transfer (BRET) assays the applicability of this regulated dimerization system to plants. Furthermore, we show that fusion proteins consisting of the C-terminal domain of Arabidopsis cryptochrome 2 fused to FKBP and FRB and coexpressed in Arabidopsis cells specifically induce the expression of cryptochrome-controlled reporter and endogenous genes in darkness upon incubation with the rapalog. These results demonstrate that the activation of cryptochrome signal transduction can be chemically induced in a dose-dependent fashion and uncoupled from the light signal, and provide the groundwork for gain-of-function experiments to study specifically the role of photoreceptors in darkness or in signaling cross-talk even under light conditions that activate members of all photoreceptor families.
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