Targeting of proConA to the plant vacuole depends on its nine amino-acid C-terminal propeptide.
Concanavalin A (ConA) is a well characterized and extensively used lectin accumulated in the protein bodies of jack bean cotyledons. ConA is synthesized as an inactive precursor proConA. The maturation of inactive proConA into biologically active ConA is a complex process including the removal of an internal glycopeptide and a C-terminal propeptide (CTPP), followed by a head-to-tail ligation of the two largest polypeptides. The cDNA encoding proConA was cloned and expressed in tobacco BY-2 cells. ProConA was slowly transported to the vacuole where its maturation into ConA was similar to that in jack bean cotyledons, apart from an incomplete final ligation. To investigate the role of the nine amino acid CTPP, a truncated form lacking the propeptide (proConADelta9) was expressed in BY-2 cells. In contrast to proConA, proConADelta9 was rapidly chased out of the endoplasmic reticulum (ER) and secreted into the culture medium. The CTPP was then fused to the C-terminal end of a secreted form of green fluorescent protein (secGFP). When expressed in tobacco BY-2 cells and leaf protoplasts, the chimaeric protein was located in the vacuole whereas secGFP was located in the culture medium and in the vacuole. Altogether, our results show we have isolated a new C-terminal vacuolar sorting determinant.
Saint-Jore-Dupas
Claude
C
CNRS UMR 6037, IFRMP 23, GDR 2590, Université de Rouen, UFR des Sciences, Bâtiment Extension Biologie, Mont-Saint-Aignan, France.
Claude
Saint-Jore-Dupas
SJ
Gilbert
Marie-Agnès
MA
Marie-Agnès
Gilbert
G
Ramis
Catalina
C
Catalina
Ramis
R
Paris
Nadine
N
Nadine
Paris
P
Kiefer-Meyer
Marie-Christine
MC
Marie-Christine
Kiefer-Meyer
KM
Neuhaus
Jean-Marc
JM
Jean-Marc
Neuhaus
N
Faye
Loïc
L
Loïc
Faye
F
Gomord
Véronique
V
Véronique
Gomord
G
D001483:Base Sequence; D002460:Cell Line; D003208:Concanavalin A / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers; D018076:DNA, Complementary; D016521:Electrophoresis, Gel, Pulsed-Field; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000378:metabolism*; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism*; D014026:Nicotiana / Q000166:cytology / Q000378:metabolism*; D014617:Vacuoles / Q000378:metabolism*
D017931:DNA Primers; D018076:DNA, Complementary; D009842:Oligopeptides; D011498:Protein Precursors; D003208:Concanavalin A