Light Quality-Dependent Nuclear Import of the Plant Photoreceptors Phytochrome A and B

Abstract

The phytochrome (phy) family of plant photoreceptors controls various aspects of photomorphogenesis. Overexpression of rice phyA-green fluorescent protein (GFP) and tobacco phyB-GFP fusion proteins in tobacco results in functional photoreceptors. phyA-GFP and phyB-GFP are localized in the cytosol of dark-adapted plants. In our experiments, red light treatment led to nuclear translocation of phyA-GFP and phyB-GFP, albeit with different kinetics. Red light-induced nuclear import of phyB-GFP, but not that of phyA-GFP, was inhibited by far-red light. Far-red light alone only induced nuclear translocation of phyA-GFP. These observations indicate that nuclear import of phyA-GFP is controlled by a very low fluence response, whereas translocation of phyB-GFP is regulated by a low fluence response of phytochrome. Thus, light-regulated nucleocytoplasmic partitioning of phyA and phyB is a major step in phytochrome signaling.

Light-Induced Nuclear Translocation of Endogenous Pea Phytochrome A Visualized by Immunocytochemical Procedures

“…The GFP signal was subsequently seen as dense speckling in the nucleus under both cR and cFR light and was visible in the cytosol under cR but not cFR light (Kircher et al, 1999b). The rice PhyA-GFP fusion protein also showed a strong speckled distribution in nuclei of lightgrown transgenic tobacco (Kircher et al, 1999b). These results differ from ours in the kinetics, distribution pattern, and protein stability under cR and cFR irradiation.…”

“…The authors observed speckling of the GFP signal in the cytoplasm within 2 min of pulse irradiation. The GFP signal was subsequently seen as dense speckling in the nucleus under both cR and cFR light and was visible in the cytosol under cR but not cFR light (Kircher et al, 1999b). The rice PhyA-GFP fusion protein also showed a strong speckled distribution in nuclei of lightgrown transgenic tobacco (Kircher et al, 1999b).…”

“…A biologically active Arabidopsis PhyB-GFP fusion protein localized to the nucleus in transgenic Arabidopsis under cR light (Yamaguchi et al, 1999). R light pulses induced nuclear localization of biologically active tobacco PhyB-GFP in transgenic tobacco, but this was reversible by exposure to FR light (Kircher et al, 1999b). In addition, important amounts of endogenous PhyB are detected in nuclei isolated from light-grown Arabidopsis leaves (Sakamoto and Nagatani, 1996) as well as from light-grown pea seedlings (A. Nagatani, unpublished result).…”

“…However, detailed analyses of fluence and fluence-rate dependence will be necessary to determine whether differences in effectiveness for photoconversion of PhyA in R and FR light can account for differences in its subcellular redistribution. Kircher et al (1999b) recently reported that a single 5-min pulse of R or FR light followed by 15 min of darkness was sufficient to induce detectable nuclear accumulation of rice PhyA-GFP in transgenic tobacco. The authors observed speckling of the GFP signal in the cytoplasm within 2 min of pulse irradiation.…”

“…More recently, a phenotypic analysis of transgenic Arabidopsis plants expressing a fusion of full-length Arabidopsis PHYB and GFP showed that the fusion protein was both spectrally and biologically functional in vivo and accumulated in the nucleus in a lightdependent manner (Yamaguchi et al, 1999). A parsley PhyA apoprotein (PHYA)-GFP fusion transiently expressed in parsley protoplasts was constitutively cytosolic (Kircher et al, 1999a), whereas both biologically active rice PhyA-GFP and tobacco PhyB-GFP expressed in transgenic tobacco plants showed nuclear import dependent on the quality of the light (Kircher et al, 1999b).Because GFP fluorescence does not require any cofactor or substrate, it can be used to observe the subcellular localization of fusion proteins in vivo. However, during such fusion protein studies, one often has to use a strong, nonnative promoter, such as cauliflower mosaic virus 35S, to detect clear labels from the reporter.…”