title: A role of Toc33 in the protochlorophyllide-dependent plastid import pathway of NADPH:protochlorophyllide oxidoreductase (POR) A.
abstract: NADPH:protochlorophyllide oxidoreductase (POR) A is a key enzyme of chlorophyll biosynthesis in angiosperms. It is nucleus-encoded, synthesized as a larger precursor in the cytosol and imported into the plastids in a substrate-dependent manner. Plastid envelope membrane proteins, called protochlorophyllide-dependent translocon proteins, Ptcs, have been identified that interact with pPORA during import. Among them are a 16-kDa ortholog of the previously characterized outer envelope protein Oep16 (named Ptc16) and a 33-kDa protein (Ptc33) related to the GTP-binding proteins Toc33 and Toc34 of Arabidopsis. In the present work, we studied the interactions and roles of Ptc16 and Ptc33 during pPORA import. Radiolabeled Ptc16/Oep16 was synthesized from a corresponding cDNA and imported into isolated Arabidopsis plastids. Crosslinking experiments revealed that import of 35S-Oep16/Ptc16 is stimulated by GTP. 35S-Oep16/Ptc16 forms larger complexes with Toc33 but not Toc34. Plastids of the ppi1 mutant of Arabidopsis lacking Toc33, were unable to import pPORA in darkness but imported the small subunit precursor of ribulose-1,5-bisphosphate carboxylase/oxygenase (pSSU), precursor ferredoxin (pFd) as well as pPORB which is a close relative of pPORA. In white light, partial suppressions of pSSU, pFd and pPORB import were observed. Our results unveil a hitherto unrecognized role of Toc33 in pPORA import and suggest photooxidative membrane damage, induced by excess Pchlide accumulating in ppi1 chloroplasts because of the lack of pPORA import, to be the cause of the general drop of protein import.
authors.affiliation: Université Joseph Fourier et Centre National de la Recherche Scientifique (CNRS), UMR5575, CERMO, BP53, F-38041 Grenoble Cedex 9, France. steffen.reinbothe@ujf-grenoble.fr
authors.forename: Steffen
authors.initials: S
authors.lastname: Reinbothe
authors.forename: Stephan
authors.initials: S
authors.lastname: Pollmann
authors.forename: Armin
authors.initials: A
authors.lastname: Springer
authors.forename: Richard Johari
authors.initials: RJ
authors.lastname: James
authors.forename: Gabrielle
authors.initials: G
authors.lastname: Tichtinsky
authors.forename: Christiane
authors.initials: C
authors.lastname: Reinbothe
mesh_terms: D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000502:physiology*; D001693:Biological Transport, Active; D008565:Membrane Proteins / Q000502:physiology*; D044925:Oxidoreductases Acting on CH-CH Group Donors / Q000378:metabolism*; D018087:Plastids / Q000378:metabolism*; D013379:Substrate Specificity
chemical_list: D029681:Arabidopsis Proteins; D008565:Membrane Proteins; C114786:Toc33 protein, Arabidopsis; D044925:Oxidoreductases Acting on CH-CH Group Donors; C023886:protochlorophyllide reductase