Existence of catalase-less peroxisomes in Sf21 insect cells.
Catalase activity, a peroxisomal marker enzyme, was not detectable in any of the subcellular fractions of Spodoptera frugiperda (Sf) 21 insect cells, although marker enzymes in other organelles were distributed in the fractions in a manner similar to that seen in mammalian cells. When a green fluorescent protein fused with peroxisome targeting signal 1 at the C-terminal (GFP-SKL) was expressed in Sf21 cells, punctate fluorescent dots were observed in the cytoplasm. The fraction where GFP-SKL was concentrated exhibited long-chain and very-long-chain fatty acid beta-oxidation activities in the presence of KCN and the density of this fraction was slightly higher than that of mitochondria. Immunoelectron microscopy studies with anti-SKL antibody demonstrated that Sf21 cells have immunoreactive peroxisome-like organelles which are structurally distinct from mitochondria, endoplasmic reticulum, and lysosomes. In contrast to peroxisomal matrix proteins, adrenoleukodystrophy protein, a peroxisomal membrane protein, was not located to peroxisomes. This suggests that the targeting signal for PMP in insect cells is distinct from that in mammalian cells. These results demonstrate that Sf21 insect cells have unique catalase-less peroxisomes capable of beta-oxidation of fatty acids.
Kurisu
Mikinori
M
Department of Biological Chemistry, Faculty of Pharmaceutical Sciences, Toyama Medical and Pharmaceutical University, 2630 Sugitani, 930-0194, Toyama, Japan.
Morita
Masashi
M
Kashiwayama
Yoshinori
Y
Yokota
Sadaki
S
Hayashi
Hiroshi
H
Sakai
Yasuyoshi
Y
Ohkuma
Shoji
S
Nishimura
Mikio
M
Imanaka
Tsuneo
T
D000074063:ATP Binding Cassette Transporter, Subfamily D, Member 1; D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D002374:Catalase / Q000378:metabolism*; D002460:Cell Line; D004274:DNA, Recombinant / Q000235:genetics; D005227:Fatty Acids / Q000378:metabolism; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D016253:Microscopy, Immunoelectron; D010084:Oxidation-Reduction; D000074435:Peroxisome-Targeting Signal 1 Receptor; D020675:Peroxisomes / Q000378:metabolism* / Q000648:ultrastructure; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D018411:Spodoptera; D013347:Subcellular Fractions / Q000378:metabolism
C115572:ABCD1 protein, human; D000074063:ATP Binding Cassette Transporter, Subfamily D, Member 1; D018528:ATP-Binding Cassette Transporters; D004274:DNA, Recombinant; D005227:Fatty Acids; D008164:Luminescent Proteins; D000074435:Peroxisome-Targeting Signal 1 Receptor; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D002374:Catalase