title: Chloroplast proteins without cleavable transit peptides: rare exceptions or a major constituent of the chloroplast proteome?
abstract: Most chloroplast proteins (cp proteins) are nucleus-encoded, synthesized on cytosolic ribosomes as precursor proteins containing a presequence (cTP), and post-translationally imported via the Tic/Toc complex into the organelle, where the cTP is removed. Only a few unambiguous instances of cp proteins that do not require cTPs (non-canonical cp proteins) have been reported so far. However, the survey of data from large-scale proteomic studies presented here suggests that the fraction of such proteins in the total cp proteome might be as large as approximately 30%. To explore this discrepancy, we chose a representative set of 28 putative non-canonical cp proteins, and used in vitro import and Red Fluorescent Protein (RFP)-fusion assays to determine their sub-cellular destinations. Four proteins, including embryo defective 1211, glycolate oxidase 2, protein disulfide isomerase-like protein (PDII), and a putative glutathione S-transferase, could be unambiguously assigned to the chloroplast. Several others ('potential cp proteins') were found to be imported into chloroplasts in vitro, but failed to localize to the organelle when RFP was fused to their C-terminal ends. Extrapolations suggest that the fraction of cp proteins that enter the inner compartments of the organelle, although they lack a cTP, might be as large as 11.4% of the total cp proteome. Our data also support the idea that cytosolic proteins that associate with the cp outer membrane might account for false positive cp proteins obtained in earlier studies.
authors.affiliation: Lehrstuhl für Botanik, Department Biologie I, Ludwig-Maximilians-Universität München, Menzinger Str. 67, D-80638 München, Germany.
authors.forename: Ute
authors.initials: U
authors.lastname: Armbruster
authors.forename: Alexander
authors.initials: A
authors.lastname: Hertle
authors.forename: Elina
authors.initials: E
authors.lastname: Makarenko
authors.forename: Jessica
authors.initials: J
authors.lastname: Zühlke
authors.forename: Mathias
authors.initials: M
authors.lastname: Pribil
authors.forename: Angela
authors.initials: A
authors.lastname: Dietzmann
authors.forename: Ivo
authors.initials: I
authors.lastname: Schliebner
authors.forename: Elena
authors.initials: E
authors.lastname: Aseeva
authors.forename: Elena
authors.initials: E
authors.lastname: Fenino
authors.forename: Michael
authors.initials: M
authors.lastname: Scharfenberg
authors.forename: Christian
authors.initials: C
authors.lastname: Voigt
authors.forename: Dario
authors.initials: D
authors.lastname: Leister
mesh_terms: D000595:Amino Acid Sequence; D002736:Chloroplasts / Q000378:metabolism*; D018076:DNA, Complementary / Q000235:genetics; D030562:Databases, Protein; D008164:Luminescent Proteins / Q000378:metabolism; D015388:Organelles / Q000378:metabolism; D018532:Pisum sativum / Q000378:metabolism; D018515:Plant Leaves / Q000378:metabolism; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D020543:Proteome / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D036226:Seedlings / Q000378:metabolism; D014158:Transcription, Genetic; D000097573:Red Fluorescent Protein
chemical_list: D018076:DNA, Complementary; D008164:Luminescent Proteins; D010940:Plant Proteins; D020543:Proteome; D012333:RNA, Messenger; D000097573:Red Fluorescent Protein