Transport of chimeric proteins that contain a carboxy-terminal targeting signal into plant microbodies.
Malate synthase is a glyoxysome-specific enzyme. The carboxy-terminal tripeptide of the enzyme is Ser-Arg-Leu (SRL), which is known to function as a peroxisomal targeting signal in mammalian cells. To analyze the function of the carboxy-terminal amino acids of pumpkin malate synthase in plant cells, a chimeric gene was constructed that encoded a fusion protein which consisted of beta-glucuronidase and the carboxyl terminus of the enzyme. The fusion protein was expressed and accumulated in transgenic Arabidopsis that had been transformed with the chimeric gene. Immunocytochemical analysis of the transgenic plants revealed that the carboxy-terminal five amino acids of pumpkin malate synthase were sufficient for transport of the fusion protein into glyoxysomes in etiolated cotyledons, into leaf peroxisomes in green cotyledons and in mature leaves, and into unspecialized microbodies in roots, although the fusion protein was no longer transported into microbodies when SRL at the carboxyl terminus was deleted. Transport of proteins into glyoxysomes and leaf peroxisomes was also observed when the carboxy-terminal amino acids of the fusion protein were changed from SRL to SKL, SRM, ARL or PRL. The results suggest that tripeptides with S, A or P at the -3 position, K or R at the -2 position, and L or M at the carboxyl terminal position can function as a targeting signal for three kinds of plant microbody.
Hayashi
M
M
Department of Cell Biology, National Institute for Basic Biology, Okazaki, Japan.
Aoki
M
M
Kato
A
A
Kondo
M
M
Nishimura
M
M
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D001483:Base Sequence; D001693:Biological Transport, Active; D018744:DNA, Plant / Q000235:genetics; D008292:Malate Synthase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008830:Microbodies / Q000378:metabolism; D016253:Microscopy, Immunoelectron; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D030821:Plants, Genetically Modified; D021382:Protein Sorting Signals / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014675:Vegetables / Q000201:enzymology / Q000235:genetics
D018744:DNA, Plant; D010940:Plant Proteins; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D008292:Malate Synthase