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Peroxisome synthesis in the absence of preexisting peroxisomes.

South ST, Gould SJ - J. Cell Biol. (1999)

Bottom Line: We also identified human PEX16, a novel integral peroxisomal membrane protein, and found that PBD061 had inactivating mutations in the PEX16 gene.These results demonstrate that peroxisomes do not necessarily arise from division of preexisting peroxisomes.We propose that peroxisomes may form by either of two pathways: one that involves PEX11-mediated division of preexisting peroxisomes, and another that involves PEX16-mediated formation of peroxisomes in the absence of preexisting peroxisomes.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.

ABSTRACT
Zellweger syndrome and related diseases are caused by defective import of peroxisomal matrix proteins. In all previously reported Zellweger syndrome cell lines the defect could be assigned to the matrix protein import pathway since peroxisome membranes were present, and import of integral peroxisomal membrane proteins was normal. However, we report here a Zellweger syndrome patient (PBD061) with an unusual cellular phenotype, an inability to import peroxisomal membrane proteins. We also identified human PEX16, a novel integral peroxisomal membrane protein, and found that PBD061 had inactivating mutations in the PEX16 gene. Previous studies have suggested that peroxisomes arise from preexisting peroxisomes but we find that expression of PEX16 restores the formation of new peroxisomes in PBD061 cells. Peroxisome synthesis and peroxisomal membrane protein import could be detected within 2-3 h of PEX16 injection and was followed by matrix protein import. These results demonstrate that peroxisomes do not necessarily arise from division of preexisting peroxisomes. We propose that peroxisomes may form by either of two pathways: one that involves PEX11-mediated division of preexisting peroxisomes, and another that involves PEX16-mediated formation of peroxisomes in the absence of preexisting peroxisomes.

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The PMP import defect of CG9 cells applies to numerous PMPs. The CG9 cell line, PBD061 (A–C); and the CG10 cell line,  PBD094 (D–F), were transfected with vectors designed to express PMP70 myc (A and D), PEX12myc (B and E), and PMP32myc (C  and F). 2 d after transfection the cells were processed for indirect immunofluorescence using antibodies specific for the myc tag. The  peroxisomal nature of the punctate structures in PBD094 cells was confirmed by double label experiments with peroxisomal membrane  markers (data not shown). Bar, 10 μm.
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Figure 2: The PMP import defect of CG9 cells applies to numerous PMPs. The CG9 cell line, PBD061 (A–C); and the CG10 cell line, PBD094 (D–F), were transfected with vectors designed to express PMP70 myc (A and D), PEX12myc (B and E), and PMP32myc (C and F). 2 d after transfection the cells were processed for indirect immunofluorescence using antibodies specific for the myc tag. The peroxisomal nature of the punctate structures in PBD094 cells was confirmed by double label experiments with peroxisomal membrane markers (data not shown). Bar, 10 μm.

Mentions: As an independent test of this phenotype we followed the distribution of overexpressed PMPs in this cell line. Plasmids designed to express the integral peroxisomal membrane proteins PMP70myc, PEX12myc and PMP32myc (a human peroxisomal solute carrier) were transfected into PBD061 cells and a control PBD fibroblast line, PBD094, which is mutated in the PEX2 gene (Shimozawa et al., 1992). We were unable to detect peroxisomal or vesicular staining for these PMPs in PBD061 cells, even though all three proteins were efficiently targeted to peroxisome membranes in PBD094 cells (Fig. 2). Similar rates of transfection in both cell lines were confirmed by cotransfection with a cytosolic marker expression vector (data not shown). Similar results were observed in cells transfected with expression vectors designed to express other human PMPs, including PEX3myc (Kammerer et al., 1998), PEX10myc (Warren et al., 1998), PEX11αmyc (Schrader et al., 1998), PEX11βmyc (Schrader et al., 1998), and PEX13myc (Gould et al., 1996) (data not shown). Although we were unable to detect peroxisomal or vesicular structures with any of the 11 PMPs tested, it is possible that some as yet unidentified PMPs may reside in some type of peroxisome-related structure in PBD061 cells.


Peroxisome synthesis in the absence of preexisting peroxisomes.

South ST, Gould SJ - J. Cell Biol. (1999)

The PMP import defect of CG9 cells applies to numerous PMPs. The CG9 cell line, PBD061 (A–C); and the CG10 cell line,  PBD094 (D–F), were transfected with vectors designed to express PMP70 myc (A and D), PEX12myc (B and E), and PMP32myc (C  and F). 2 d after transfection the cells were processed for indirect immunofluorescence using antibodies specific for the myc tag. The  peroxisomal nature of the punctate structures in PBD094 cells was confirmed by double label experiments with peroxisomal membrane  markers (data not shown). Bar, 10 μm.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2132891&req=5

Figure 2: The PMP import defect of CG9 cells applies to numerous PMPs. The CG9 cell line, PBD061 (A–C); and the CG10 cell line, PBD094 (D–F), were transfected with vectors designed to express PMP70 myc (A and D), PEX12myc (B and E), and PMP32myc (C and F). 2 d after transfection the cells were processed for indirect immunofluorescence using antibodies specific for the myc tag. The peroxisomal nature of the punctate structures in PBD094 cells was confirmed by double label experiments with peroxisomal membrane markers (data not shown). Bar, 10 μm.
Mentions: As an independent test of this phenotype we followed the distribution of overexpressed PMPs in this cell line. Plasmids designed to express the integral peroxisomal membrane proteins PMP70myc, PEX12myc and PMP32myc (a human peroxisomal solute carrier) were transfected into PBD061 cells and a control PBD fibroblast line, PBD094, which is mutated in the PEX2 gene (Shimozawa et al., 1992). We were unable to detect peroxisomal or vesicular staining for these PMPs in PBD061 cells, even though all three proteins were efficiently targeted to peroxisome membranes in PBD094 cells (Fig. 2). Similar rates of transfection in both cell lines were confirmed by cotransfection with a cytosolic marker expression vector (data not shown). Similar results were observed in cells transfected with expression vectors designed to express other human PMPs, including PEX3myc (Kammerer et al., 1998), PEX10myc (Warren et al., 1998), PEX11αmyc (Schrader et al., 1998), PEX11βmyc (Schrader et al., 1998), and PEX13myc (Gould et al., 1996) (data not shown). Although we were unable to detect peroxisomal or vesicular structures with any of the 11 PMPs tested, it is possible that some as yet unidentified PMPs may reside in some type of peroxisome-related structure in PBD061 cells.

Bottom Line: We also identified human PEX16, a novel integral peroxisomal membrane protein, and found that PBD061 had inactivating mutations in the PEX16 gene.These results demonstrate that peroxisomes do not necessarily arise from division of preexisting peroxisomes.We propose that peroxisomes may form by either of two pathways: one that involves PEX11-mediated division of preexisting peroxisomes, and another that involves PEX16-mediated formation of peroxisomes in the absence of preexisting peroxisomes.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Chemistry, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.

ABSTRACT
Zellweger syndrome and related diseases are caused by defective import of peroxisomal matrix proteins. In all previously reported Zellweger syndrome cell lines the defect could be assigned to the matrix protein import pathway since peroxisome membranes were present, and import of integral peroxisomal membrane proteins was normal. However, we report here a Zellweger syndrome patient (PBD061) with an unusual cellular phenotype, an inability to import peroxisomal membrane proteins. We also identified human PEX16, a novel integral peroxisomal membrane protein, and found that PBD061 had inactivating mutations in the PEX16 gene. Previous studies have suggested that peroxisomes arise from preexisting peroxisomes but we find that expression of PEX16 restores the formation of new peroxisomes in PBD061 cells. Peroxisome synthesis and peroxisomal membrane protein import could be detected within 2-3 h of PEX16 injection and was followed by matrix protein import. These results demonstrate that peroxisomes do not necessarily arise from division of preexisting peroxisomes. We propose that peroxisomes may form by either of two pathways: one that involves PEX11-mediated division of preexisting peroxisomes, and another that involves PEX16-mediated formation of peroxisomes in the absence of preexisting peroxisomes.

Show MeSH
Related in: MedlinePlus