Proteome analysis of Arabidopsis leaf peroxisomes reveals novel targeting peptides, metabolic pathways, and defense mechanisms.
We have established a protocol for the isolation of highly purified peroxisomes from mature Arabidopsis thaliana leaves and analyzed the proteome by complementary gel-based and gel-free approaches. Seventy-eight nonredundant proteins were identified, of which 42 novel proteins had previously not been associated with plant peroxisomes. Seventeen novel proteins carried predicted peroxisomal targeting signals (PTS) type 1 or type 2; 11 proteins contained PTS-related peptides. Peroxisome targeting was supported for many novel proteins by in silico analyses and confirmed for 11 representative full-length fusion proteins by fluorescence microscopy. The targeting function of predicted and unpredicted signals was investigated and SSL>, SSI>, and ASL> were established as novel functional PTS1 peptides. In contrast with the generally accepted confinement of PTS2 peptides to the N-terminal domain, the bifunctional transthyretin-like protein was demonstrated to carry internally a functional PTS2. The novel enzymes include numerous enoyl-CoA hydratases, short-chain dehydrogenases, and several enzymes involved in NADP and glutathione metabolism. Seven proteins, including beta-glucosidases and myrosinases, support the currently emerging evidence for an important role of leaf peroxisomes in defense against pathogens and herbivores. The data provide new insights into the biology of plant peroxisomes and improve the prediction accuracy of peroxisome-targeted proteins from genome sequences.
Reumann
Sigrun
S
Department of Plant Biochemistry, Georg-August-University of Goettingen, Albrecht-von-Haller-Institute for Plant Sciences, D-37077 Goettingen, Germany. sreuman@gwdg.de
Babujee
Lavanya
L
Ma
Changle
C
Wienkoop
Stephanie
S
Siemsen
Tanja
T
Antonicelli
Gerardo E
GE
Rasche
Nicolas
N
Lüder
Franziska
F
Weckwerth
Wolfram
W
Jahn
Olaf
O
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000737:chemistry / Q000378:metabolism*; D019295:Computational Biology; D015180:Electrophoresis, Gel, Two-Dimensional; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D000074435:Peroxisome-Targeting Signal 1 Receptor; D020675:Peroxisomes / Q000378:metabolism*; D018515:Plant Leaves / Q000166:cytology / Q000378:metabolism*; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
D029681:Arabidopsis Proteins; D010455:Peptides; D000074435:Peroxisome-Targeting Signal 1 Receptor; D020543:Proteome; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins