Sterol homeostasis requires regulated degradation of squalene monooxygenase by the ubiquitin ligase Doa10/Teb4.
Sterol homeostasis requires regulated degradation of squalene monooxygenase by the ubiquitin ligase Doa10/Teb4.
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DOI：
10.7554/elife.00953
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发表时间：
2013-07-23
期刊：
影响因子：
7.7
通讯作者：
Carvalho P
中科院分区：
文献类型：
--
作者：
Foresti O;Ruggiano A;Hannibal-Bach HK;Ejsing CS;Carvalho P
甾醇稳态对细胞膜的功能至关重要，需要羟甲基戊酸途径的限速酶HMGR的反馈抑制。由于HMGR在该通路的起始阶段起作用，其调控影响固醇和其他必需的甲羟戊酸衍生代谢物（如泛醌或醇）的合成。在这里，我们描述了一个新颖的，进化上保守的反馈系统，在甲羟戊酸途径的甾醇特异性步骤操作。这涉及由酵母Doa10或哺乳动物Teb4介导的角鲨烯单加氧酶的甾醇依赖性降解，Teb4是一种泛素连接酶，涉及内质网（ER）相关蛋白降解（ERAD）途径的一个分支。由于调节HMGR需要ERAD的另一个分支，因此我们的研究结果揭示了ERAD在固醇稳态中的基本作用，该途径的两个分支共同作用，在不同水平上控制甾醇生物合成，从而允许独立调节甲羟戊酸途径的多种产物。DOI: http://dx.doi.org/10.7554/eLife.00953.001所有的细胞都被一层膜包裹着，这层膜是由称为脂质的脂肪分子组成的，上面布满了蛋白质。这层膜允许细胞检测外部事件并作出反应。由于外部条件（如温度）变化很大，膜需要能够调整其性能。例如，随着温度的升高，脂质变得更加流动，因此膜通过结合称为甾醇的分子来增加其刚性来应对热应力。事实上，甾醇对膜特性有深远的影响，对调节许多细胞过程至关重要。但是高水平的甾醇是有毒的，所以必须小心控制。固醇，如酵母中的麦角甾醇或哺乳动物中的胆固醇，是在一个严格调控的多步骤过程中合成的；甾醇生产的一些早期步骤也为细胞中其他关键分子提供了共同的构建块。控制甾醇水平的一种机制是对一种酶进行有调节的破坏，这种酶在甾醇合成的早期阶段起作用。这是通过内质网相关降解（ERAD）途径的一个分支发生的，该途径也会破坏非功能性蛋白质。现在，Foresti等人发现甾醇合成也受到ERAD途径的另一个分支的调控。第二个控制点发生在生物合成过程的后期，允许细胞独立于从相同的初步化合物衍生的途径的其他产物来调节甾醇水平。在酵母中，两个ERAD分支由Hrd1和Doa10引导。这两种都是泛素连接酶——一种将泛素标签附着在其他蛋白质上的蛋白质，从而使它们被蛋白酶体（本质上是细胞中的废物处理复合体）回收利用。为了鉴定被Doa10标记的蛋白，Foresti等人比较了缺乏Doa10的菌株与野生型酵母的蛋白水平。出乎意料的是，帮助合成麦角甾醇的Erg1酶在缺乏Doa10的细胞中更为丰富。Foresti等人发现，当细胞内麦角甾醇的基本成分水平上升时，Doa10标记Erg1进行破坏。这些麦角甾醇中间体对酵母是有毒的，酵母利用蛋白质Are1/2将它们转化为危害较小的固醇酯分子。当DOA10或ARE1/2基因缺失时，这些中间产物更加丰富；引人注目的是，当这三个基因在同一菌株中被敲除时，它们变得更加普遍。相反，通过删除HRD1阻断另一个ERAD分支不会导致麦角甾醇中间体积累，也不会加剧ARE1/2敲除的影响。结合之前的研究结果，这些结果提供了ERAD途径的不同分支以不同的步骤调节麦角甾醇合成的证据。当检测到高水平的胆固醇时，在人类细胞中也观察到同样的机制。通过确定控制固醇水平的平行途径，这项工作加强了膜完整性对生命的重要性。DOI: http://dx.doi.org/10.7554/eLife.00953.002
Sterol homeostasis is essential for the function of cellular membranes and requires feedback inhibition of HMGR, a rate-limiting enzyme of the mevalonate pathway. As HMGR acts at the beginning of the pathway, its regulation affects the synthesis of sterols and of other essential mevalonate-derived metabolites, such as ubiquinone or dolichol. Here, we describe a novel, evolutionarily conserved feedback system operating at a sterol-specific step of the mevalonate pathway. This involves the sterol-dependent degradation of squalene monooxygenase mediated by the yeast Doa10 or mammalian Teb4, a ubiquitin ligase implicated in a branch of the endoplasmic reticulum (ER)-associated protein degradation (ERAD) pathway. Since the other branch of ERAD is required for HMGR regulation, our results reveal a fundamental role for ERAD in sterol homeostasis, with the two branches of this pathway acting together to control sterol biosynthesis at different levels and thereby allowing independent regulation of multiple products of the mevalonate pathway. DOI: http://dx.doi.org/10.7554/eLife.00953.001 All cells are enclosed by a membrane that is made up of fatty molecules called lipids and is studded with proteins. This membrane allows cells to detect and react to outside events. Since external conditions, such as temperature, can vary dramatically, membranes need to be able to adjust their properties. For example, lipids become more fluid as the temperature rises, so membranes respond to heat stress by incorporating molecules called sterols to increase their rigidity. In fact, sterols have profound effects on membrane properties and are essential to regulate a number of cellular processes. But high levels of sterols can become toxic, so it is essential that they are carefully controlled. Sterols, such as ergosterol in yeast or cholesterol in mammals, are synthesized in a tightly regulated multi-step process; some of the early steps in sterol production also make common building blocks for other key molecules in the cell. A mechanism to control sterol levels is the regulated destruction of an enzyme that carries out an early step of their synthesis. This occurs via one branch of the ER-associated degradation (ERAD) pathway, which also destroys non-functional proteins. Now, Foresti et al. have found that sterol synthesis is also regulated by another branch of the ERAD pathway. This second control point, which occurs later in the biosynthetic process, allows cells to regulate sterol levels independent of the other products of the pathway that are derived from the same preliminary compounds. In yeast, the two ERAD branches are directed by Hrd1 and Doa10. These are both ubiquitin ligases—proteins that attach a tag called ubiquitin to other proteins, thus labeling them for recycling by the proteasome (essentially a waste-disposal complex in the cell). To identify the proteins that are tagged by Doa10, Foresti et al. compared protein levels in strains lacking Doa10 with those in wild type yeast. Unexpectedly, the enzyme Erg1, which helps to synthesize ergosterol, was more abundant in cells lacking Doa10. Foresti et al. found that Doa10 tagged Erg1 for destruction when levels of the building blocks of ergosterol rose inside the cell. These ergosterol intermediates are toxic to yeast, which converts them into less harmful molecules known as sterol esters using the proteins Are1/2. When the DOA10 or ARE1/2 genes were deleted, these intermediates were more abundant; strikingly, they became even more prevalent when all three genes were knocked out in the same strain. In contrast, blocking the other ERAD branch by deleting HRD1 did not cause ergosterol intermediates to accumulate, nor did it exacerbate the effects of ARE1/2 knockout. When combined with previous findings, these results provide evidence that the different branches of the ERAD pathway regulate ergosterol synthesis at distinct steps. The same mechanism is observed in human cells when high levels of cholesterol are detected. By identifying parallel routes to control sterol levels, this work reinforces the importance of membrane integrity to life. DOI: http://dx.doi.org/10.7554/eLife.00953.002