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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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Different fates for different PMPs in PBD061 cells. Total  cellular protein was extracted from  the CG9 cell line, PBD061, and the  CG10 cell line, PBD094. Equal  amounts of each lysate (by protein)  were separated by SDS-PAGE, and  blotted with antibodies specific for  the PMPs PMP70 (A), P70R (B);  and PEX14 (C). In addition, total  cellular protein was extracted from  PBD061 and PBD094 cells that had  been transfected with the PEX12myc  expression vector. Equal amounts  of each lysate (by protein) were  separated by SDS-PAGE, and blotted with antibodies specific for the  myc epotope tag (D).
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Figure 3: Different fates for different PMPs in PBD061 cells. Total cellular protein was extracted from the CG9 cell line, PBD061, and the CG10 cell line, PBD094. Equal amounts of each lysate (by protein) were separated by SDS-PAGE, and blotted with antibodies specific for the PMPs PMP70 (A), P70R (B); and PEX14 (C). In addition, total cellular protein was extracted from PBD061 and PBD094 cells that had been transfected with the PEX12myc expression vector. Equal amounts of each lysate (by protein) were separated by SDS-PAGE, and blotted with antibodies specific for the myc epotope tag (D).

Mentions: To better understand the fates of PMPs in PBD061 cells, total cellular protein was extracted from the CG9 cell line, PBD061, and a CG10 cell line, PBD094, as well as from PBD061, and PBD094 cells that had been transfected with the PEX12myc expression vector. The abundance of the PMPs (PMP70, P70R, PEX14, and PEX12myc) were then determined by immunoblot with antibodies specific for each protein (Fig. 3). As evident from this experiment, levels of full-length PMP70 and P70R in PBD061 cells were below the limit of detection in PBD061 cells but were readily detected in PBD094 cells, indicating that these proteins might be degraded in PBD061 cells. This hypothesis is supported by the presence of anti–P70R-reactive products of lower molecular mass in PBD061 cells, but not in PBD094 cells. However, this was not the fate of all PMPs, since levels of PEX14 were similar in PBD061 cells and PBD094 cells, as were levels of PEX12myc. Thus, the fate of PMPs in a cell line defective in PMP import varies from protein to protein, just as the fate of peroxisomal matrix proteins varies in cells with specific defects in matrix protein import (Lazarow and Fujiki, 1985; Santos et al., 1988a, 1988b).


Peroxisome synthesis in the absence of preexisting peroxisomes.

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

Different fates for different PMPs in PBD061 cells. Total  cellular protein was extracted from  the CG9 cell line, PBD061, and the  CG10 cell line, PBD094. Equal  amounts of each lysate (by protein)  were separated by SDS-PAGE, and  blotted with antibodies specific for  the PMPs PMP70 (A), P70R (B);  and PEX14 (C). In addition, total  cellular protein was extracted from  PBD061 and PBD094 cells that had  been transfected with the PEX12myc  expression vector. Equal amounts  of each lysate (by protein) were  separated by SDS-PAGE, and blotted with antibodies specific for the  myc epotope tag (D).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2132891&req=5

Figure 3: Different fates for different PMPs in PBD061 cells. Total cellular protein was extracted from the CG9 cell line, PBD061, and the CG10 cell line, PBD094. Equal amounts of each lysate (by protein) were separated by SDS-PAGE, and blotted with antibodies specific for the PMPs PMP70 (A), P70R (B); and PEX14 (C). In addition, total cellular protein was extracted from PBD061 and PBD094 cells that had been transfected with the PEX12myc expression vector. Equal amounts of each lysate (by protein) were separated by SDS-PAGE, and blotted with antibodies specific for the myc epotope tag (D).
Mentions: To better understand the fates of PMPs in PBD061 cells, total cellular protein was extracted from the CG9 cell line, PBD061, and a CG10 cell line, PBD094, as well as from PBD061, and PBD094 cells that had been transfected with the PEX12myc expression vector. The abundance of the PMPs (PMP70, P70R, PEX14, and PEX12myc) were then determined by immunoblot with antibodies specific for each protein (Fig. 3). As evident from this experiment, levels of full-length PMP70 and P70R in PBD061 cells were below the limit of detection in PBD061 cells but were readily detected in PBD094 cells, indicating that these proteins might be degraded in PBD061 cells. This hypothesis is supported by the presence of anti–P70R-reactive products of lower molecular mass in PBD061 cells, but not in PBD094 cells. However, this was not the fate of all PMPs, since levels of PEX14 were similar in PBD061 cells and PBD094 cells, as were levels of PEX12myc. Thus, the fate of PMPs in a cell line defective in PMP import varies from protein to protein, just as the fate of peroxisomal matrix proteins varies in cells with specific defects in matrix protein import (Lazarow and Fujiki, 1985; Santos et al., 1988a, 1988b).

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