BIOGENESIS OF PEROXISOME WITH YEAST MUTANTS AND CLONED GENES

齐薇格综合征引起的主要缺陷与过氧化物酶体的组装受损有关。到目前为止，该综合征的病因被分为9个基因互补组。对于他们中的任何一个，基因产物的功能都没有被理解。本联合研究分离了酵母过氧化物酶体组装所必需的基因，研究了该细胞器的生物发生过程。分离出过氧化物酶体组装缺陷的酿酒Sacchromyces cerevisiae突变体，这些突变体能够在乙酸上生长，但在油酸上生长，并且在细胞质中错误定位了邻近基质酶。突变体有18个互补基团（pas1-pas18）。PAS1基因编码一个117 kda疏水蛋白。预测的序列包含两个假定的atp结合位点，并揭示了与参与各种细胞过程的蛋白质的结构关系：细胞周期控制（Cdc48p），囊泡介导的蛋白质运输（Sec18p）和HIV基因表达（TBP-1）。PAS2基因的产物是泛素结合蛋白家族的第10个成员。pas2突变体细胞积累了含有pas3p的膜状结构（见下文）。这些结构可能是所谓的齐薇格成纤维细胞的幽灵。Pas3p是由PAS3基因编码的一个50.6 kda的蛋白，当大部分分子暴露在细胞质中时，它应该通过其氨基末端锚定在过氧化物酶体膜上。分离了一种脂肪酸同化酵母——麦芽糖假丝酵母（Candida maltosa）的PAS1同源物。缺乏PAS3野生型等位基因的细胞在过氧化物酶的分布上表现出与pas1突变体相同的表型，并且对活性氧的有效繁殖剂白蜡素敏感。

The primary defect arising from Zellweger syndrome is linked to impaired assembly of peroxisomes. The causes of the syndrome are thus far classified into 9 genetic complementation groups. For none of them, the function of the gene product has been understood. In this joint Research, yeast genes essential for the assembly of peroxisomes were isolated to study the process of the biogenesis of this organelle.Sacchromyces cerevisiae mutants defective in peroxisome assembly were isolated that were able to grow on acetate but on oleic acid, and mislocated proxisomal matrix enzymes in the cytosol. There are 18 complementation groups of mutants (pas1-pas18). The PAS1 gene encoded a hydrophobic 117-kDa protein. The predicted sequence contained two putative ATP-binding sites and revealed a structural relationship to proteins involved in various cellular processes : control of cell cycle (Cdc48p), vesicle-mediated protein transport(Sec18p), and gene expression of HIV (TBP-1). The product of the PAS2 gene was the 10th member of the ubiquitin-conjugating protein family. The pas2 mutant cells accumulated the membranous structures that contained pas3p (see below). These structures could be the so-called ghosts of Zellweger fibroblasts. Pas3p, a 50.6-kDa protein encoded by the PAS3 gene, was supposed to be anchored in the peroxisomal membrane by its amino-terminus while the bulk of the molecule was exposed to cytosol.The PAS1 homologue of a fatty-acid-assimilating yeast, Candida maltosa, was isolated. The cells lacking the wild-type alleles of PAS3 showed the same phenotype as the pas1 mutant in respect of the distribution of peroxisomal enzymes, and became sensitive to plumbagin, which is a potent propagator of reactive oxygen.