A new approach for the AAA peroxin research

AAA蛋白家族的两种过氧化物素Pexlp和Pex6p分别与过氧化物酶体生物发生障碍(PBDS)有关，如互补基团1(CG1)和CG4的Zellweger综合征。这些过氧化物酶被Pex26p募集到过氧化物体膜上，推测可能参与了PTS1基质蛋白的可溶性再循环受体Pex5p从过氧体中的输出。此外，我们最近报道，Pexlp部分存在于细胞质中，但没有形成Pexlp-Pex6p-Pex26p复合体。然而，Pex5p循环系统的分子机制还不是很清楚。因此，我们研究了AAA过氧化物素在过氧化体膜和胞浆中的作用。免疫共沉淀和下拉实验表明，Pex26p能够与Pex5p相关的Pex14p结合。Pex26p-Pex14p复合体被Pexlp和Pex6p以依赖于ATP的方式解离。对HEK293细胞胞浆组分和重组蛋白的Blue-Native凝胶分析表明，Pex1p主要以同源三聚体形式存在。在对CHO-K1细胞中Pex5p寡聚体结构的分析中，胞浆中的Pex5p既有二聚体也有单体形式，而Pex1p缺失的ZP107细胞胞浆中的Pex5p主要以单体形式存在。综上所述，Pexlp-Pex6p复合体可能调节Pex26p和Pex14p的相互作用，可能从Pex14p中释放Pex5p。此外，Pexlp复合体的同源三聚体可能参与了胞质Pex5p从单体向二聚体的转化。我们讨论了AAA过氧化物素在过氧化体蛋白输入和Pex5p循环中的顺序作用。

Two peroxins, Pexlp and Pex6p, of the AAA protein family are responsible for peroxisome biogenesis disorders (PBDs) such as Zellweger syndrome of complementation group 1 (CG1) and CG4, respectively. These peroxins were recruited to peroxisomal membrane by Pex26p and suggested to be involved in the export from peroxisomes of Pex5p, the soluble recycling receptor for PTS1 matrix proteins. Furthermore, we recently reported that Pexlp was present partly in cytosol without forming a Pexlp-Pex6p-Pex26p complex. However, the molecular mechanism of the recycling system for Pex5p is not well understood. Therefore we investigated the role of AAA peroxins on peroxisomal membrane as well as in the cytosol. Co-immunoprecipitation and pull-down assays showed that Pex26p is competent to bind to Pex14p associated with Pex5p. The Pex26p-Pex14p complex was dissociated by Pexlp and Pex6p in an ATP-dependent manner. Blue-Native gel analysis of the cytosolic fraction from HEK293 cells and recombinant protein showed that Pex1p is mostly in a homo-trimer. In the assays for Pex5p oligomer structure in CHO-K1 cells, cytosolic Pex5p showed both dimer and monomer forms, although cytosolic Pex5p from Pex1p-deficient ZP107 cells was predominantly in a monomer form. Taken together, Pexlp-Pex6p complexes are likely to modulate the interaction of Pex26p and Pex14p, presumably releasing Pex5p from Pex14p. Furthermore, homo-trimer of Pexlp complexes were suggested to be involved in the conversion from a monomer to a dimer of cytosolic Pex5p. We discussed the sequential role of AAA peroxins in peroxisomal protein import and Pex5p recycling.