Sakai Y, Marshall P A, Saiganji A, Takabe K, Saiki H, Kato N, Goodman J M
Department of Agricultural Chemistry, Faculty of Agriculture, Kyoto University, Japan.
J Bacteriol. 1995 Dec;177(23):6773-81. doi: 10.1128/jb.177.23.6773-6781.1995.
The mechanism of peroxisome proliferation is poorly understood. Candida boidinii is a methylotrophic yeast that undergoes rapid and massive peroxisome proliferation and serves as a good model system for this process. Pmp30A and Pmp30B (formerly designated Pmp31 and Pmp32, respectively) are two closely related proteins in a polyploid strain of this yeast that are strongly induced by diverse peroxisome proliferators such as methanol, oleate, and D-alanine. The function of these proteins is not understood. To study this issue, we used a recently described haploid strain (S2) of C. boidinii that can be manipulated genetically. We now report that strain S2 contains a single PMP30 gene very similar in sequence (greater than 93% identity at the DNA level) to PMP30A and PMP30B. When PMP30 was disrupted, cell growth on methanol was greatly inhibited, and cells grown in both methanol and oleate had fewer, larger, and more spherical peroxisomes than wild-type cells. A similar phenotype was recently described for Saccharomyces cerevisiae cultured on oleate in which PMP27, which encodes a protein of related sequence that is important for peroxisome proliferation, was disrupted. To determine whether Pmp27 is a functional homolog of Pmp30, gentle complementation was performed. PMP30A was expressed in the PMP27 disruptant of S. cerevisiae, and PMP27 was expressed in the PMP30 disruptant of C. boidinii S2. Complementation, in terms of both cell growth and organelle size, shape, and number, was successful in both directions, although reversion to a wild-type phenotype was only partial for the PMP30 disruptant. We conclude that these proteins are functional homologs and that both Pmp30 and Pmp27 have a direct role in proliferation and organelle size rather than a role in a specific peroxisomal metabolic pathway of substrate utilization.
过氧化物酶体增殖的机制目前还知之甚少。博伊丁假丝酵母是一种甲基营养型酵母,它会经历快速且大量的过氧化物酶体增殖,是研究这一过程的良好模型系统。Pmp30A和Pmp30B(以前分别命名为Pmp31和Pmp32)是这种酵母多倍体菌株中两个密切相关的蛋白质,它们会被多种过氧化物酶体增殖剂(如甲醇、油酸和D-丙氨酸)强烈诱导。这些蛋白质的功能尚不清楚。为了研究这个问题,我们使用了最近描述的一种可进行基因操作的博伊丁假丝酵母单倍体菌株(S2)。我们现在报告,菌株S2含有一个单一的PMP30基因,其序列与PMP30A和PMP30B非常相似(在DNA水平上同一性大于93%)。当PMP30被破坏时,细胞在甲醇上的生长受到极大抑制,在甲醇和油酸中生长的细胞比野生型细胞具有更少、更大且更呈球形的过氧化物酶体。最近在油酸上培养的酿酒酵母中也描述了类似的表型,其中编码对过氧化物酶体增殖很重要的相关序列蛋白质的PMP27被破坏。为了确定Pmp27是否是Pmp30的功能同源物,进行了温和互补实验。PMP30A在酿酒酵母的PMP27破坏株中表达,PMP27在博伊丁假丝酵母S2的PMP30破坏株中表达。在细胞生长以及细胞器大小、形状和数量方面,两个方向的互补都取得了成功,尽管对于PMP30破坏株,恢复到野生型表型只是部分成功。我们得出结论,这些蛋白质是功能同源物,并且Pmp30和Pmp27在增殖和细胞器大小方面都具有直接作用,而不是在底物利用的特定过氧化物酶体代谢途径中起作用。