Fusion of small peroxisomal vesicles in vitro reconstructs an early step in the in vivo multistep peroxisome assembly pathway of Yarrowia lipolytica.
We have identified and purified six subforms of peroxisomes, designated P1 to P6, from the yeast, Yarrowia lipolytica. An analysis of trafficking of peroxisomal proteins in vivo suggests the existence of a multistep peroxisome assembly pathway in Y. lipolytica. This pathway operates by conversion of peroxisomal subforms in the direction P1, P2-->P3-->P4-->P5-->P6 and involves the import of various peroxisomal proteins into distinct vesicular intermediates. We have also reconstituted in vitro the fusion of the earliest intermediates in the pathway, small peroxisomal vesicles P1 and P2. Their fusion leads to the formation of a larger and more dense peroxisomal vesicle, P3. Fusion of P1 and P2 in vitro requires cytosol and ATP hydrolysis and is inhibited by antibodies to two membrane-associated ATPases of the AAA family, Pex1p and Pex6p. We provide evidence that the fusion in vitro of P1 and P2 peroxisomes reconstructs an actual early step in the peroxisome assembly pathway operating in vivo in Y. lipolytica.
Titorenko
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Department of Cell Biology, University of Alberta, Edmonton, Alberta T6G 2H7, Canada.
Chan
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Rachubinski
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D000074183:ATPases Associated with Diverse Cellular Activities; D000251:Adenosine Triphosphatases / Q000378:metabolism; D001483:Base Sequence; D002352:Carrier Proteins / Q000378:metabolism; D018076:DNA, Complementary; D007425:Intracellular Membranes / Q000502:physiology*; D008561:Membrane Fusion / Q000502:physiology*; D008969:Molecular Sequence Data; D052066:N-Ethylmaleimide-Sensitive Proteins; D020675:Peroxisomes / Q000502:physiology*; D004718:Saccharomycetales / Q000378:metabolism / Q000502:physiology*; D033921:Vesicular Transport Proteins
D002352:Carrier Proteins; D018076:DNA, Complementary; D033921:Vesicular Transport Proteins; D000251:Adenosine Triphosphatases; D000074183:ATPases Associated with Diverse Cellular Activities; C096998:Pex6 protein, rat; D052066:N-Ethylmaleimide-Sensitive Proteins; C498737:Nsf protein, rat