title: Polyphenol oxidase can cross thylakoids by both the Tat and the Sec-dependent pathways: a putative role for two stromal processing sites.
abstract: Polyphenol oxidase (PPO; EC 1.10.3.2 or EC 1.14.18.1), a thylakoid-lumen protein encoded by a nuclear gene, plays a role in the defense of plants against both herbivores and pathogens. Although previously reported to be a Tat (twin-arginine-dependent translocation) protein, the import of PPO by isolated chloroplasts was inhibited by azide, a diagnostic inhibitor of the Sec-dependent pathway. Import of PPO inhibited thylakoid translocation of a Tat protein and did not affect translocation of Sec-dependent proteins. In contrast, a pre-accumulated iPPO competed with Sec-dependent but not with Tat proteins. A previously reported second processing step in the stroma removes a twin-Arg that is part of a 'Sec-avoidance' motif in the thylakoid targeting domain of PPO. When the second processing site was mutated, the import of the resulting precursor showed Sec-dependent characteristics. The PPO transit peptide could drive thylakoid translocation of a Tat protein in the dark. Azide inhibited the secretion of a PPO intermediate that lacks a twin-Arg to the periplasm of Escherichia coli, but had no effect on the export of the intermediate containing the twin-Arg. PPO is synthesized in plants in response to wound and pathogen-related signals and it is possible that when the Tat pathway is unable to translocate adequate amounts of newly synthesized PPO, translocation is diverted to the Sec-dependent pathway by processing the intermediate at the second site and removing the twin-Arg.
authors.affiliation: Department of Plant Sciences, The Hebrew University of Jerusalem, Jerusalem 91904, Israel. shai@unr.edu
authors.forename: Shai
authors.initials: S
authors.lastname: Koussevitzky
authors.forename: Emma
authors.initials: E
authors.lastname: Ne'eman
authors.forename: Smadar
authors.initials: S
authors.lastname: Peleg
authors.forename: Eitan
authors.initials: E
authors.lastname: Harel
mesh_terms: D001386:Azides / Q000494:pharmacology; D004156:Catechol Oxidase / Q000378:metabolism*; D003624:Darkness; D004926:Escherichia coli / Q000378:metabolism; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D050505:Mutant Proteins / Q000378:metabolism; D009550:Nigericin / Q000494:pharmacology; D018532:Pisum sativum / Q000187:drug effects / Q000201:enzymology*; D010940:Plant Proteins / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D021382:Protein Sorting Signals; D021381:Protein Transport / Q000187:drug effects; D020524:Thylakoids / Q000187:drug effects / Q000201:enzymology*
chemical_list: D001386:Azides; D050505:Mutant Proteins; D010940:Plant Proteins; D011498:Protein Precursors; D021382:Protein Sorting Signals; D004156:Catechol Oxidase; D009550:Nigericin