FHY1 Mediates Nuclear Import of the Light-Activated Phytochrome A Photoreceptor

et al. (2008) FHY1 Mediates Nuclear Import of the Light-Activated Phytochrome A Photoreceptor. PLoS Genet 4(8): e1000143. doi:10.1371/journal.pgen.1000143 FHY1 Mediates Nuclear Import of the Light-Activated Phytochrome A Photoreceptor Thierry Genoud 0 Fabian Schweizer 0 Anke Tscheuschler 0 Dimitry Debrieux 0 Jorge J. Casal 0 Eberhard Scha fer 0 Andreas Hiltbrunner 0 Christian Fankhauser 0 Gregory P. Copenhaver, The University of North Carolina at Chapel Hill, United States of America 0 1 Centre for Integrative Genomics, University of Lausanne, Lausanne, Switzerland, 2 Institut fu r Biologie II/Botanik, Albert Ludwigs Universita t , Freiburg, Germany, 3 IFEVA , Facultad de Agronom a, Universidad de Buenos Aires , Buenos Aires, Argentina, 4 Consejo Nacional de Investigaciones Cient ficas y Te cnicas (CONICET), Buenos Aires, Argentina, 5 BIOSS , Centre for Biological Signalling Studies, University of Freiburg , Freiburg , Germany The phytochrome (phy) family of photoreceptors is of crucial importance throughout the life cycle of higher plants. Lightinduced nuclear import is required for most phytochrome responses. Nuclear accumulation of phyA is dependent on two related proteins called FHY1 (Far-red elongated HYpocotyl 1) and FHL (FHY1 Like), with FHY1 playing the predominant function. The transcription of FHY1 and FHL are controlled by FHY3 (Far-red elongated HYpocotyl 3) and FAR1 (FAr-red impaired Response 1), a related pair of transcription factors, which thus indirectly control phyA nuclear accumulation. FHY1 and FHL preferentially interact with the light-activated form of phyA, but the mechanism by which they enable photoreceptor accumulation in the nucleus remains unsolved. Sequence comparison of numerous FHY1-related proteins indicates that only the NLS located at the N-terminus and the phyA-interaction domain located at the C-terminus are conserved. We demonstrate that these two parts of FHY1 are sufficient for FHY1 function. phyA nuclear accumulation is inhibited in the presence of high levels of FHY1 variants unable to enter the nucleus. Furthermore, nuclear accumulation of phyA becomes light- and FHY1-independent when an NLS sequence is fused to phyA, strongly suggesting that FHY1 mediates nuclear import of light-activated phyA. In accordance with this idea, FHY1 and FHY3 become functionally dispensable in seedlings expressing a constitutively nuclear version of phyA. Our data suggest that the mechanism uncovered in Arabidopsis is conserved in higher plants. Moreover, this mechanism allows us to propose a model explaining why phyA needs a specific nuclear import pathway. - Funding: This work was supported by grants from the Swiss National Science Foundation and HFSP to CF (SNF 3100A0-112638; HFSP RGY0016/2004-C), the Deutsche Forschungsgemeinschaft (DFG) to ES (SFB 592 and SFB 746), the Agencia Nacional de Promocio n Cientfica y Tecnolo gica to JJC (BID 1728 OC-AR PICT32492) and a postdoctoral fellowship from HFSP to AH (LT00631/2003-C). Competing Interests: The authors have declared that no competing interests exist. Plants are sessile organisms and therefore have to adapt growth and development to the environmental conditions at their site of germination. Light is one of the most important factors directing such adaptive responses and it is involved in many developmental steps throughout the life of plants [1,2]. To detect intensity, quality (wavelength) and direction of incident light plants have evolved a set of photoreceptors monitoring red/far-red (R/FR), blue/UV-A and UV-B [37]. The phytochrome family of red/far-red photoreceptors plays a key role in seed germination, leaf and stem development, circadian rhythms, shade avoidance and induction of flowering [8]. Although in higher plants phytochromes are not the primary photoreceptors controlling phototropism and chloroplast movements, the phytochromes modulate these responses [911]. Phytochromes are homodimeric chromoproteins containing the linear tetrapyrole phytochromobilin as chromophore. They photoconvert between two spectrally distinct forms: the redlight-absorbing Pr and the biologically active far-red lightabsorbing Pfr form [3,12]. As the absorption spectra of the two forms overlap the photoconversion is not complete in either direction. Irradiation with light therefore results in a wavelengthspecific equilibrium between the Pr and Pfr forms, with only ,2% Pfr in far-red light and ,85% Pfr in red light [13]. Under natural conditions the Pfr/Pr ratio differs dramatically depending on the position of the plant within the community (canopy shade versus open environment) [14,15]. In Arabidopsis the phytochrome gene family consists of five members (PHYAE), among which PHYA and PHYB play the most prominent functions [16]. phyB is the major red light receptor and mediates the red/far-red reversible low fluence response (LFR). Other members of the phytochrome family contribute to responses primarily controlled by phyB. In contrast, responses to continuous far-red light (high irradiance response, HIR) and to single light pulse of very low fluence light (VLFR) depend exclusively on phyA [1,3,12]. Photoreceptor mutants have reduced fitness but only the phyA mutant is conditionally lethal, highlighting the importance of this photoreceptor [17,18]. Its functional importance is further revealed by the high degree of sequence conservation among all angiosperms [19]. phyA is also crucial for the modulation of phototropin responses such as the enhancement of phototropism [10,11]. In response to changes in the environment, animals can take shelter while the sessile plants must adapt to the prevalent conditions. Great plasticity in growth and development are striking examples of how plants cope with a changing environment. In plants, light is both a source of energy and an essential informational cue perceived by several classes of photoreceptors. Phytochrome-mediated light signaling is particularly well studied, because these photoreceptors control all aspects of the plant life cycle. The phytochromes are cytoplasmic in the dark and must enter the nucleus upon light activation to initiate signal transduction. How this important light-regulated event is achieved is poorly understood. Here we describe the function of an evolutionary conserved protein called FHY1 for Far-red elongated HYpocotyl 1. We demonstrate that FHY1 interacts with a light-activated phytochrome in the cytoplasm, allowing the complex to be transported into the nucleus. Interestingly, if this phytochrome can enter the nucleus by another mechanism, FHY1 is no longer required for seedling development, indicating that a major function of FHY1 is to chaperone an activated phytochrome into the nucleus. Our experiments suggest that this mechanism uncovered in Arabidopsis is widely conserved among flowering plants. The subcellular localization of phytochromes is tightly regulated by light. They localize to the cytosol in the dark but transloca (...truncated)