Localization and topogenesis studies of cytoplasmic and vacuolar homologs of the Galanthus nivalis agglutinin.
The Galanthus nivalis agglutinin (GNA) is synthesized as a preproprotein. To corroborate the role of the different targeting peptides in the topogenesis of GNA and related proteins, different constructs were made whereby both the complete original GNA gene and different truncated sequences were coupled to the enhanced green fluorescent protein (EGFP). In addition, a GNA ortholog from rice that lacks the signal peptide and C-terminal propeptide sequence was fused to EGFP. These fusion constructs were expressed in tobacco BY-2 cells and their localization analyzed by confocal fluorescence microscopy. We observed that the processed preproprotein of GNA was directed towards the vacuolar compartment, whereas both the truncated forms of GNA corresponding to the mature lectin polypeptide and the rice ortholog of GNA were located in the nucleus and the cytoplasm. It can be concluded, therefore, that removal of the C-terminal propeptide and the signal peptide is sufficient to change the subcellular targeting of a normally vacuolar protein to the nuclear/cytoplasmic compartment of the BY-2 cells. These findings support the proposed hypothesis that cytoplasmic/nuclear GNA-like proteins and their vacuolar homologs are evolutionarily related and that the classical GNA-related lectins might have evolved from cytoplasmic orthologs through an evolutionary event involving the insertion of a signal peptide and a C-terminal propeptide.
Fouquaert
Elke
E
Laboratory of Biochemistry and Glycobiology, Department of Molecular Biotechnology, Ghent University, Coupure Links 653, B-9000 Gent, Belgium.
Hanton
Sally L
SL
Brandizzi
Federica
F
Peumans
Willy J
WJ
Van Damme
Els J M
EJ
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000235:genetics / Q000254:growth & development; D002460:Cell Line; D002467:Cell Nucleus; D003593:Cytoplasm / Q000378:metabolism*; D027801:Galanthus / Q000737:chemistry / Q000166:cytology* / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D008969:Molecular Sequence Data; D012275:Oryza / Q000235:genetics; D010802:Phylogeny; D018515:Plant Leaves; D037121:Plant Lectins / Q000737:chemistry / Q000378:metabolism*; D021381:Protein Transport; D014026:Nicotiana / Q000166:cytology / Q000378:metabolism; D014617:Vacuoles / Q000378:metabolism*
D037121:Plant Lectins