Biochemical studies of Zmpste24-deficient mice
Biochemical studies of Zmpste24-deficient mice
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DOI：
10.1074/jbc.m102908200
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发表时间：
2001-08-03
影响因子：
4.8
通讯作者：
Young, SG
中科院分区：
文献类型：
--
作者：
Leung, GK;Schmidt, WK;Young, SG
在酿酒酵母中的遗传研究鉴定了两个基因，STE 24和RCE 1，参与从以CAAX序列基序终止的异戊二烯化蛋白质切割三个羧基末端氨基酸。Ste 24 p从酵母交配信息素a-因子切割羧基末端“-AAX”，而Rce 1 p从a-因子和Ras 2 p切割-AAX。Ste 24 p还切割α-因子的氨基末端。小鼠基因组含有酵母RCE 1和STE 24的直系同源物。我们以前证明，与基因敲除实验，小鼠卷轴是必不可少的发展和卷轴是完全负责的羧基末端蛋白水解处理的小鼠Ras蛋白。在这项研究中，我们克隆了小鼠Zmpste 24，酵母STE 24的直向同源物，并表明它可以促进α-因子在酵母中表达时的生产。然后，为了评估Zmpste 24在发育中的重要性，我们产生了Zmpste 24缺陷小鼠。与Reel基因敲除小鼠不同，Zmpste 24缺陷小鼠在发育中存活，并且具有生育能力。由于尚未鉴定出哺乳动物Zmpste 24的天然底物，因此使用酵母α-因子作为替代底物来研究来自Zmpste 24缺陷型小鼠的细胞和组织的膜中的生物化学活性。我们证明，Zmpste 24缺陷的小鼠膜，如Ste 24 p缺陷的酵母膜，具有减少的CAAX蛋白水解活性，并缺乏切割α-因子前体的氨基末端的能力。因此，酵母Ste 24 p的两种酶活性在小鼠Zmpste 24中是保守的，但这些酶活性对于小鼠发育或生育力不是必需的。
Genetic studies in Saccharomyces cerevisiae identified two genes, STE24 and RCE1, involved in cleaving the three carboxyl-terminal amino acids from isoprenylated proteins that terminate with a CAAX sequence motif. Ste24p cleaves the carboxyl-terminal "-AAX" from the yeast mating pheromone a-factor, whereas Rce1p cleaves the -AAX from both a-factor and Ras2p. Ste24p also cleaves the amino terminus of a-factor. The mouse genome contains orthologues for both yeast RCE1 and STE24. We previously demonstrated, with a gene-knockout experiment, that mouse Reel is essential for development and that Reel is entirely responsible for the carboxyl-terminal proteolytic processing of the mouse Ras proteins. In this study, we cloned mouse Zmpste24, the orthologue for yeast STE24 and showed that it could promote a-factor production when expressed in yeast. Then, to assess the importance of Zmpste24 in development, we generated Zmpste24-deficient mice. Unlike the Reel knockout mice, Zmpste24-deficient mice survived development and were fertile. Since no natural substrates for mammalian Zmpste24 have been identified, yeast a-factor was used as a surrogate substrate to investigate the biochemical activities in membranes from the cells and tissues of Zmpste24-deficient mice. We demonstrate that Zmpste24-deficient mouse membranes, like Ste24p-deficient yeast membranes, have diminished CAAX proteolytic activity and lack the ability to cleave the amino terminus of the a-factor precursor. Thus, both enzymatic activities of yeast Ste24p are conserved in mouse Zmpste24, but these enzymatic activities are not essential for mouse development or for fertility.