substrate-independent, 14:3:3 protein-mediated plastid import pathway of NADPH:protochlorophyllide oxidoreductase A

• Published in: Proceedings of the National Academy of Sciences - PNAS 2007-05, Vol.104 (20), p.8538-8543
• Main Authors: Schemenewitz, Andreas, Pollmann, Stephan, Reinbothe, Christiane, Reinbothe, Steffen
• Format: Article
• Language: English
• Subjects: 14-3-3 Proteins - metabolism; 14:3:3 protein; Animals; Antiserum; Barley; binding proteins; Binding Sites; Biochemistry; Biochemistry, Molecular Biology; Biological Sciences; Chemical synthesis; Chloroplasts; Cross-Linking Reagents - pharmacology; Enzymes; Hordeum; Hordeum vulgare; HSP70 Heat-Shock Proteins - metabolism; Immunoprecipitation; Imports; Life Sciences; Light; Membranes; Mutation - genetics; NADPH:protochlorophyllide oxidoreductase A; oxidoreductases; Oxidoreductases Acting on CH-CH Group Donors - biosynthesis; Oxidoreductases Acting on CH-CH Group Donors - metabolism; Oxygenases - metabolism; Peas; Plant cells; plant proteins; Plants; Plastids; Plastids - drug effects; Plastids - metabolism; Plastids - radiation effects; Protein Binding - drug effects; Protein Binding - radiation effects; Protein Biosynthesis - drug effects; Protein Biosynthesis - radiation effects; Protein Conformation - drug effects; Protein Conformation - radiation effects; Protein precursors; Protein Precursors - chemistry; Protein Precursors - metabolism; protein sorting; protein transport; Protein Transport - drug effects; Protein Transport - radiation effects; protein-protein interactions; Proteins; Protochlorophyllide - pharmacology; Rabbits; Substrate Specificity - drug effects; Substrate Specificity - radiation effects; Triticum aestivum; Vegetal Biology; zymogens
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.0702058104

Plastids are semiautonomous organelles that contain only limited coding information in their own DNA. Because most of their genome was transferred to the nucleus after their endosymbiotic origin, plastids must import the major part of their protein constituents from the cytosol. The exact role of cytosolic targeting factors in the regulation of plastid protein import has not been determined. Here, we report that the nucleus-encoded NADPH:protochlorophyllide (Pchlide) oxidoreductase A plastid precursor (pPORA) can use two different plastid import pathways that differ by the requirements for cytosolic 14:3:3 proteins and Hsp70. pPORA synthesized in a wheat germ lysate segregated into different precursor fractions. While import of free pPORA and only Hsp70-complexed pPORA was Pchlide-dependent and involved the previously identified Pchlide-dependent translocon, 14:3:3 protein- and Hsp70-complexed pPORA was transported into Pchlide-free chloroplasts through the Toc75-containing standard translocon at the outer chloroplast membrane/translocon at the inner chloroplast membrane machinery. A 14:3:3 protein binding site was identified in the mature region of the ³⁵S-pPORA, which governed 14:3:3 protein- and Hsp70-mediated, Pchlide-independent plastid import. Collectively, our results reveal that the import of pPORA into the plastids is tightly regulated and involves different cytosolic targeting factors and plastid envelope translocon complexes.