No abstract

To perceive red and far-red light, plants have evolved specific photoreceptors called phytochromes. Even though the spectral properties of all phytochromes are very similar, they show a distinct mode of action. Here we describe EID1, a negatively acting component of the signaling cascade that shifts the responsiveness of the phytochrome A (phyA) signaling system associated with hypocotyl elongation from red to far-red wavelengths. EID1 is a novel nuclear F-box protein that contains a leucine zipper whose integrity is necessary for its biological function. EID1 most probably acts by targeting activated components of the phyA signaling pathway to ubiquitin-dependent proteolysis.

SummaryCullin proteins, which belong to multigenic families in all eukaryotes, associate with other proteins to form ubiquitin protein ligases (E3s) that target substrates for proteolysis by the 26S proteasome. Here, we present the molecular and genetic characterization of a plant Cullin3. In contrast to fungi and animals, the genome of the model plant Arabidopsis thaliana contains two related CUL3 genes, called CUL3A and CUL3B. We found that CUL3A is ubiquitously expressed in plants and is able to interact with the ring-finger protein RBX1. A genomic search revealed the existence of at least 76 BTB-domain proteins in Arabidopsis belonging to 11 major families. Yeast two-hybrid experiments indicate that representative members of certain families are able to physically interact with both CUL3A and CUL3B, suggesting that Arabidopsis CUL3 forms E3 protein complexes with certain BTB domain proteins. In order to determine the function of CUL3A, we used a reverse genetic approach. The cul3a null mutant flowers slightly later than the control plants. Furthermore, this mutant exhibits a reduced sensitivity of the inhibition of hypocotyl growth in far-red light and miss-expresses COP1. The viability of the mutant plants suggests functional redundancy between the two CUL3 genes in Arabidopsis.

To identify specific mutants for components of phytochrome A (phyA) signaling in Arabidopsis, we established a light program consisting of multiple treatments with alternating red and far-red light. In wild-type seedlings, irradiation with multiple red light pulses can reduce the amount of phyA, which in turn decreases the high-irradiance responses (HIRs) mediated by the subsequent treatments with far-red light. Our mutants were able to avoid this red light-dependent reduction of the HIR. Here, we describe eid1, a new recessive mutant with increased sensitivity to far-red light. The eid1 mutation maps to the top of chromosome 4. The mutants showed no change in phenotype in darkness or under continuous white light, but they exhibited an increased sensitivity to red light and an increased persistence of HIR during prolonged dark phases after multiple short pulses of far-red light. The eid1 seedlings accumulated normal amounts of phytochrome and showed no alterations in the degradation or de novo synthesis of phyA. The expression of the Eid1 phenotype requires the presence of phyA. Our data provide evidence that EID1 is a negatively acting component in the phyA-dependent HIR-signaling pathway.

The EID1-like protein 3 (EDL3) shows high similarity to EID1 (Empfindlicher im dunkelroten Licht 1), an F-box protein that functions as a negative regulator in the signalling cascade downstream of the phytochrome A photoreceptor in Arabidopsis thaliana. Analyses revealed a strong and rapid induction of EDL3 gene expression under osmotic stress, high salinity, and upon abscisic acid (ABA) application. Therefore, it was speculated that EDL3 is involved in the regulation of responses controlled by this plant hormone, which not only regulates many aspects of plant development but also integrates responses towards temperature, drought, osmotic, and salt stresses. Physiological data obtained with over-expresser lines and a conditional knock-down mutant demonstrated that EDL3 functions as a positive regulator in ABA-dependent signalling cascades that control seed germination, root growth, greening of etiolated seedlings, and transition to flowering. Results further demonstrate that EDL3 regulates anthocyanin accumulation under drought stress. The observed effects on physiological responses fit to tissue-specific expression patterns obtained with EDL3-promoter:GUS lines. Bimolecular Fluorescence Complementation assays and yeast two-hybrid analyses showed that EDL3 carries a functional F-box domain. Thus, the protein is presumed to act as a component of a ubiquitin ligase complex that specifically directs negatively acting factors in ABA signalling to degradation via the proteasome.