title: Light-regulated nuclear import and degradation of Arabidopsis phytochrome-A N-terminal fragments.
abstract: The photoreceptor phytochrome-A (phyA) regulates germination and seedling establishment by mediating very low fluence (VLFR) and far-red high irradiance (FR-HIR) responses in Arabidopsis thaliana. In darkness, phyA homodimers exist in the biologically inactive Pr form and are localized in the cytoplasm. Light induces formation of the biologically active Pfr form and subsequent rapid nuclear import. PhyA Pfr, in contrast to the Pr form, is labile and has a half-life of ∼30 min. We produced transgenic plants in a phyA-201 null background that express the PHYA-yellow fluorescent protein (YFP) or the PHYA686-YFP-dimerization domain (DD) and PHYA686-YFP-DD-nuclear localization signal (NLS) or PHYA686-YFP-DD-nuclear exclusion signal (NES) fusion proteins. The PHYA686-YFP fusion proteins contained the N-terminal domain of phyA (686 amino acid residues), a short DD and the YFP. Here we report that (i) PHYA686-YFP-DD fusion protein is imported into the nucleus in a light-dependent fashion; (ii) neither of the PHYA686 fusion proteins is functional in FR-HIR and nuclear VLFR; and (iii) the phyA-dependent, blue light-induced inhibition of hypocotyl growth is mediated by the PHYA686-YFP-DD-NES but not by the PHYA686-YFP-DD-NLS and PHYA686-YFP-DD fusion proteins. We demonstrate that (i) light induces degradation of all PHYA N-terminal-containing fusion proteins and (ii) these N-terminal domain-containing fusion proteins including the constitutively nuclear PHYA686-YFP-DD-NLS and predominantly cytoplasmic PHYA686-YFP-DD-NES degrade at comparable rates but markedly more slowly than PHYA-YFP, whereas (iii) light-induced degradation of the native phyA is faster compared with PHYA-YFP.
authors.affiliation: Institute of Botany, Biology II, University of Freiburg, Schänzlestrasse 1, D-79104 Freiburg, Germany.
authors.forename: Iris
authors.initials: I
authors.lastname: Wolf
authors.forename: Stefan
authors.initials: S
authors.lastname: Kircher
authors.forename: Erzsébet
authors.initials: E
authors.lastname: Fejes
authors.forename: László
authors.initials: L
authors.lastname: Kozma-Bognár
authors.forename: Eberhard
authors.initials: E
authors.lastname: Schäfer
authors.forename: Ferenc
authors.initials: F
authors.lastname: Nagy
authors.forename: Eva
authors.initials: E
authors.lastname: Adám
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000528:radiation effects; D017360:Arabidopsis / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000378:metabolism*; D001426:Bacterial Proteins / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism*; D003001:Cloning, Molecular; D008027:Light; D008164:Luminescent Proteins / Q000378:metabolism; D051942:Phytochrome A / Q000378:metabolism*; D030821:Plants, Genetically Modified / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D018749:RNA, Plant / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction
chemical_list: D029681:Arabidopsis Proteins; D001426:Bacterial Proteins; D008164:Luminescent Proteins; C497973:PHYA protein, Arabidopsis; D051942:Phytochrome A; D018749:RNA, Plant; D011993:Recombinant Fusion Proteins; C054562:yellow fluorescent protein, Bacteria