Direct interaction and determination of binding domains among peroxisomal import factors in Arabidopsis thaliana.
We analyzed the role of Arabidopsis orthologues of human Pex14p, Pex5p and Pex7p that are central components of peroxisomal protein import machinery. Immunoblot analysis showed that AtPex14p and AtPex5p were present in most organs in Arabidopsis, suggesting that these factors play a role in the main protein import pathways for plant peroxisomes. Two-hybrid analysis showed that AtPex14p interacted with AtPex5p, but not with AtPex7p. In addition, AtPex7p was bound to AtPex5p, indicating that the PTS2 pathway depends on the PTS1 pathway in Arabidopsis. Further analysis showed that the nine WXXXF/Y repeats in the amino acids 231K-450D and 1M-230V of AtPex5p are bound to two N-terminal domains, amino acids 58I-65L and 78R-97R of AtPex14p and the C-terminal amino acids 266Y-317S of AtPex7p, respectively. Since the binding domains of AtPex5p to AtPex14p and AtPex7p do not overlap, AtPex14p, AtPex5p and AtPex7p might form their complex and function cooperatively in peroxisomal protein import.
Nito
Kazumasa
K
Department of Cell Biology, National Institute for Basic Biology, Okazaki, 444-8585 Japan.
Hayashi
Makoto
M
Nishimura
Mikio
M
D020816:Amino Acid Motifs / Q000235:genetics; D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism; D029681:Arabidopsis Proteins; D001665:Binding Sites / Q000235:genetics; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D018506:Gene Expression Regulation, Plant; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008954:Models, Biological; D008969:Molecular Sequence Data; D000074437:Peroxisomal Targeting Signal 2 Receptor; D000074435:Peroxisome-Targeting Signal 1 Receptor; D020675:Peroxisomes / Q000235:genetics* / Q000378:metabolism; D011485:Protein Binding; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012097:Repressor Proteins; D017386:Sequence Homology, Amino Acid; D020798:Two-Hybrid System Techniques
D029681:Arabidopsis Proteins; D002352:Carrier Proteins; D008565:Membrane Proteins; C499192:PEX14 protein, Arabidopsis; D000074437:Peroxisomal Targeting Signal 2 Receptor; D000074435:Peroxisome-Targeting Signal 1 Receptor; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins