HY5 stability and activity in arabidopsis is regulated by phosphorylation in its COP1 binding domain.

Arabidopsis HY5 is a bZIP transcription factor that promotes photomorphogenesis. Previous studies suggested that COP1, a negative regulator of photomorphogenesis, directly interacts with nuclear HY5 and targets it for proteasome-mediated degradation. Light negatively regulates the nuclear level of COP1 and thus permits HY5 accumulation. Here we report that HY5 abundance peaks in early seedling development, consistent with its role in promoting photomorphogenesis. HY5 acts exclusively within a complex and exists in two isoforms, resulting from phosphorylation within its COP1 binding domain by a light- regulated kinase activity. Unphosphorylated HY5 shows stronger interaction with COP1, is the preferred substrate for degradation, has higher affinity to target promoters and is physiologically more active than the phosphorylated version. Therefore, HY5 phosphorylation provides an added level of light-mediated regulation of HY5 stability and activity besides nuclear COP1 levels. Regulated HY5 phosphorylation not only provides abundant and physiologically more active unphosphorylated HY5 in the light, but also helps to maintain a small pool of less active phosphorylated HY5 in the dark, which could be essential for a rapid initial response during dark-to-light transition.

Targeted destabilization of HY5 during light-regulated development of Arabidopsis.

Arabidopsis seedlings display contrasting developmental patterns depending on the ambient light. Seedlings grown in the light develop photomorphogenically, characterized by short hypocotyls and expanded green cotyledons. In contrast, seedlings grown in darkness become etiolated, with elongated hypocotyls and dosed cotyledons on an apical hook. Light signals, perceived by multiple photoreceptors and transduced to downstream regulators, dictate the extent of photomorphogenic development in a quantitative manner. Two key downstream components, COP1 and HY5, act antagonistically in regulating seedling development. HY5 is a bZIP transcription factor that binds directly to the promoters of light-inducible genes, promoting their expression and photomorphogenic development. COP1 is a RING-finger protein with WD-40 repeats whose nuclear abundance is negatively regulated by light. COP1 interacts directly with HY5 in the nucleus to regulate its activity negatively. Here we show that the abundance of HY5 is directly correlated with the extent of photomorphogenic development, and that the COP1-HY5 interaction may specifically target HY5 for proteasome-mediated degradation in the nucleus.

The role of COP1 in repression of Arabidopsis photomorphogenic development.

Photomorphogenic development in Arabidopsis is regulated by the key repressor COP1, which interacts with specific transcription factors in the nucleus to modulate their activities. In the dark, COP1 accumulates in the nucleus and represses photomorphogenic development. Light diminishes the nuclear accumulation of COP1 and abrogates its repressor activity. A number of cellular components are involved in light-dependent nucleocytoplasmic partitioning of COP1, including the multisubunit COP9 complexes and at least three well-characterized photoreceptors. This review discusses current understanding of the mechanisms of COP1 action.

Multiple photoreceptors mediate the light-induced reduction of GUS-COP1 from Arabidopsis hypocotyl nuclei.

The subcellular localization of COP1, a key photomorphogenic repressor, is regulated by light in Arabidopsis seedlings. Photoreceptor loss-of-function mutants and dominant gain-of-function overexpression transgenes were both used to analyze the influences of the three photoreceptors, phyA, phyB, and CRY1, on the light-regulated subcellular localization of COP1. Through a semiquantitative analysis of the nuclear abundance of GUS-COP1 in the various genetic backgrounds, the specific roles of the individual photoreceptors have been established. The data suggest that multiple photoreceptors influence the light-regulated subcellular localization of COP1 in white light. Under specific wavelengths of light, phyA, phyB, and CRY1 each play critical roles in mediating far-red, red, and blue light signals, respectively. Our data also support an interdependency between CRY1 and the phytochromes in mediating the light-regulated subcellular localization of COP1 and thus seedling development.

Genetic and developmental control of nuclear accumulation of COP1, a repressor of photomorphogenesis in Arabidopsis.

Using a beta-glucuronidase (GUS) reporter-COP1 fusion transgene, it was shown previously that Arabidopsis COP1 acts within the nucleus as a repressor of seedling photomorphogenic development and that high inactivation of COP1 was accompanied by a reduction of COP1 nuclear abundance (A.G. von Arnim, X.-W. Deng [1994] Cell 79: 1035-1045). Here we report that the GUS-COP1 fusion transgene can completely rescue the defect of cop1 mutations and thus is fully functional during seedling development. The kinetics of GUS-COP1 relocalization in a cop1 null mutant background during dark/light transitions imply that the regulation of the functional nuclear COP1 level plays a role in stably maintaining a committed seedling's developmental fate rather than in causing such a commitment. Analysis of GUS-COP1 cellular localization in mutant hypocotyls of all pleiotropic COP/DET/FUS loci revealed that nuclear localization of GUS-COP1 was diminished under both dark and light conditions in all mutants tested, whereas nuclear localization was not affected in the less pleiotropic cop4 mutant. Using both the brassinosteroid-deficient mutant det2 and brassinosteroid treatment of wild-type seedlings, we have demonstrated that brassinosteroid does not control the hypocotyl cell elongation through regulation nuclear localization of COP1. The growth regulator cytokinin, which also dramatically reduced hypocotyl cell elongation in the absence of light, did not prevent GUS-COP1 nuclear localization in dark-grown seedlings. Our results suggest that all of the previously characterized pleiotropic COP/DET/FUS loci are required for the proper nuclear localization of the COP1 protein in the dark, whereas the less pleiotropic COP/DET loci or plant regulators tested are likely to act either downstream of COP1 or by independent pathways.

The COP9 complex, a novel multisubunit nuclear regulator involved in light control of a plant developmental switch.

Arabidopsis COP9 is a component of a large protein complex that is essential for the light control of a developmental switch and whose conformation or size is modulated by light. The complex is acidic, binds heparin, and is localized within the nucleus. Biochemical purification of the complex to near homogeneity revealed that it contains 12 distinct subunits. One of the other subunits is COP11, mutations in which result in a phenotype identical to cop9 mutants. The COP9 complex may act to regulate the nuclear abundance of COP1, an established repressor of photomorphogenic development. During the biogenesis of the COP9 complex, a certain degree of prior subunit association is a prerequisite for proper nuclear translocation. Since both COP9 and COP11 have closely related human counterparts, the COP9 complex probably represents a conserved developmental regulator in higher eukaryotes.