The phytochrome family of red/far-red (R/FR)-responsive photoreceptors plays a key role throughout the life cycle of plants . Arabidopsis has five phytochromes, phyA-phyE, among which phyA and phyB play the most predominant functions . Light-regulated nuclear accumulation of the phytochromes is an important regulatory step of this pathway, but to this date no factor specifically required for this event has been identified . Among all phyA signaling mutants, fhy1 and fhy3 (far-red elongated hypocotyl 1 and 3) have the most severe hyposensitive phenotype, indicating that they play particularly important roles . FHY1 is a small plant-specific protein of unknown function localized both in the nucleus and the cytoplasm . Here we show that FHY1 is specifically required for the light-regulated nuclear accumulation of phyA but not phyB. Moreover, phyA accumulation is only slightly affected in fhy3, indicating that the diminished nuclear accumulation of phyA observed in fhy1 seedlings is not simply a general consequence of reduced phyA signaling. By in vitro pull-down and yeast two-hybrid analyses, we demonstrate that FHY1 physically interacts with phyA, preferentially in its active Pfr form. Furthermore, FHY1 and phyA colocalize in planta. We therefore identify the first component required for light-regulated phytochrome nuclear accumulation.

SummaryEmpfindlicher im Dunkelroten Licht 1 (EID1) is an F-box protein that functions as a negative regulator in phytochrome A (phyA)-specific light signalling. F-box proteins are components of SCF ubiquitin ligase complexes that target proteins for degradation in the proteasome. Here we present further characterization of EID1 at the expression level, and show that it regulates photomorphogenesis in seedlings, rosette leaf development and flowering. Data on transcript expression patterns indicate that EID1 is expressed during all stages of Arabidopsis development and exhibits no light response. Microscope studies demonstrate that EID1 is localized to the nucleus, where it can form speckles under continuous far-red light that resemble clastosomes. To characterize the composition and formation of SCF EID1 complexes further, we used twohybrid and bridge assays in yeast and in planta. EID1 interacts specifically with several Arabidopsis Skp1-like (ASK) proteins and Cullin1 to form stable dimeric and trimeric complexes. Our results support a two-step association process in which the F-box protein binds first to the ASK adaptor, forming a unit which then associates with the catalytic core of the SCF complex. Finally, our data indicate that the EID1 target interaction domain is composed of two independent modules.

SummaryPhotomorphogenesis of higher plants is regulated by photoreceptors including the red/far-red lightabsorbing phytochromes, blue-UV/A sensing cryptochromes and as yet uncharacterized UV/B receptors. Speci®c phototransduction pathways that are controlled by either individual or interacting photoreceptors mediate regulation. Phytochrome B (phyB) is the major red light-sensing photoreceptor. Phototransduction mediated by this light sensor has been shown to include light-dependent nuclear import and interaction of phyB with transcription factor-like proteins in the nucleus. Here we report that nuclear import of phyB and physiological responses regulated by this photoreceptor exhibit very similar wavelength-and¯uence rate-dependence. Nuclear import of phyB is insensitive to single red, blue and far-red light pulses. It is induced by continuous red light and to a lesser extent by continuous blue light, whereas far-red light is completely ineffective. The data presented indicate that light-dependent partitioning of phyB exhibits features characteristic of blue light responsiveness ampli®cation, a phenomenon that is thought to be mediated by interaction of phyB with CRY1.