A novel aspartic proteinase is targeted to the secretory pathway and to the vacuole in the moss Physcomitrella patens.
In seed plants aspartic proteases (APs) are known to reside in storage vacuoles. Targeting to this compartment is provoked by a secretory signal peptide and the plant-specific insert (PSI). In order to study secretory and vacuolar targeting in a seedless plant, the moss Physcomitrella patens, we isolated a cDNA encoding PpAP1, a novel aspartic proteinase. Sequence alignment with other members of the family of plant APs (EC 3.4.23) revealed a high overall identity and the Pfam motifs for aspartic proteinase and PSI were clearly recognised. In phylogenetic analysis PpAP1 was placed at a very basal position outside of the bigger clusters. Protoplasts transiently expressing the PpAP1 signal peptide fused to GFP showed fluorescence in a well-developed ER-Golgi network. A C-terminal fusion of GFP to the entire PpAP1 protein showed vacuolar fluorescence in transiently transfected protoplasts. Therefore, the vacuole is apparently the in-vivo target for PpAP1. In this study the three-dimensional peculiarity of the endomembrane continuum of ER and Golgi was visualised in a seedless plant for the first time. Above all the functionality of the secretory and the vacuolar targeting signals make them become useful tools for biotechnological approaches.
Schaaf
Andreas
A
Plant Biotechnology, University of Freiburg, Freiburg, Germany.
Reski
Ralf
R
Decker
Eva L
EL
D000595:Amino Acid Sequence; D016282:Aspartic Acid Endopeptidases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019068:Bryopsida / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007425:Intracellular Membranes / Q000378:metabolism; D008969:Molecular Sequence Data; D010802:Phylogeny; D021382:Protein Sorting Signals / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D022142:Secretory Vesicles / Q000378:metabolism*; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein; D014617:Vacuoles / Q000378:metabolism*
D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D016282:Aspartic Acid Endopeptidases