Formation of peroxisomes from peroxisomal ghosts in a peroxisome-deficient mammalian cell mutant upon complementation by protein microinjection.
Most mammalian cell strains genetically deficient in peroxisome biogenesis have abnormal membrane structures called ghosts, containing integral peroxisomal membrane protein, PMP70, but lacking the peroxisomal matrix proteins. Upon genetic complementation, these mutants regain the ability of peroxisome biogenesis. It is postulated that, in this process, the ghosts act as the precursors of peroxisomes, but there has been no evidence to support this. In the present study, we investigated this issue by protein microinjection to a mutant Chinese hamster ovary cell line defective of PEX5, encoding a peroxisome-targeting signal receptor. When recombinant Pex5p and green fluorescent protein (GFP) carrying a peroxisome-targeting signal were co-injected into the mutant cells, the GFP fluorescence gathered over time to particulate structures where PMP70 was co-localized. This process was dependent on both Pex5p and the targeting signal, and, most importantly, occurred even in the presence of cycloheximide, a protein synthesis inhibitor. These findings suggest that the ghosts act as acceptors of matrix proteins in the peroxisome recovery process at least in the PEX5 mutant, and support the view that peroxisomes can grow by incorporating newly synthesized matrix proteins.
Yamasaki
M
M
Department of Life Science, Faculty of Science, Himeji Institute of Technology, 3-2-1 Koto, Kamigori, Hyogo 678-1297, Japan.
Hashiguchi
N
N
Fujiwara
C
C
Imanaka
T
T
Tsukamoto
T
T
Osumi
T
T
D018528:ATP-Binding Cassette Transporters; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D005816:Genetic Complementation Test; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D008845:Microinjections; D000074435:Peroxisome-Targeting Signal 1 Receptor; D020675:Peroxisomes / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure*; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection
C063305:ABCD3 protein, human; D018528:ATP-Binding Cassette Transporters; D008164:Luminescent Proteins; D008565:Membrane Proteins; C000614264:PEX5 protein, human; D000074435:Peroxisome-Targeting Signal 1 Receptor; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins