Unraveling the mechanism by which Rps26-deficient ribosomes form to support the stress response

项目摘要/摘要 最近核糖体亚群的组成不同，缺乏单独的核糖体蛋白(RP)， 或包含特定的修改引起了人们的极大兴趣。就RP内容而言，多项研究 已经表明，细胞中存在缺乏特定RPS的核糖体，包括Rps26缺乏的核糖体。 虽然在大多数情况下，它们的生理相关性仍不清楚，但卡布斯坦实验室最近已经 证明了缺乏Rps26的核糖体是在高盐和pH胁迫下特异性地产生的，从而使 编码Hog1和Rim101途径所需蛋白的mRNAs的优先翻译 对这些压力的反应。在含有Rps26的和缺陷的之间的这种mRNA特异性的变化 核糖体产生于Rps26对Kozak序列的-4位置的识别，从而支持 富含Rps26的核糖体对翻译好的mRNAs的翻译，以及对 在Hog1和Rim101途径中由Rps26缺失的核糖体形成的特异性mRNAs 压力。 目前尚不清楚的是Rps26缺乏的核糖体是如何在高盐和高pH胁迫下形成的。我的 初步结果表明，缺乏Rps26的核糖体是由先前存在的Rps26释放而产生的 核糖体。因此，我将进一步更详细地剖析导致Rps26产生的机制- 缺乏核糖体，探索Rps26特异性伴侣TSR2在运送和提取中的作用 Rps26来自核糖体(Aim1)，并测试哪些翻译后修饰调节这一途径(AIM2)。 接下来，我将测试Rps26是否被(重新)整合到Rps26缺失的核糖体中，以允许快速切换 不需要重新构建新的核糖体的成本，或者这些核糖体是否 降级(Aim3)。 总之，这些实验将阐明缺乏Rps26的核糖体是如何在压力下形成的。除了进一步 在压力诱导特定核糖体群体产生的这一新范例上展开，结果 也将对与Rps26缺乏有关的疾病的发展产生影响，例如钻石- 布莱克凡贫血。因为10%-15%的病例的病因尚不清楚，且与RP- 缺乏，导致Rps释放的通路过度激活可能是 负责一部分病例，类似于Rps26-伴侣缺乏引起的病例子集 TSR2.此外，缺乏单个RPS的核糖体，包括Rps26，是在癌细胞中产生的，其中 它们与糟糕的结果有关。因此，这项工作还将有助于描绘癌细胞如何调节 颠覆翻译目的的翻译机器。

Project Summary/Abstract Recently ribosome subpopulations that differ in their composition, lacking individual ribosomal proteins (RPs), or containing specific modifications have garnered a lot of interest. In the case of RP content, multiple studies have shown that ribosomes lacking specific RPs are present in cells, including ribosomes deficient in Rps26. While their physiological relevance remains unclear in most cases, the Karbstein lab has recently demonstrated that ribosomes lacking Rps26 are produced specifically under high salt and pH stress to enable the preferential translation of mRNAs encoding proteins from the Hog1 and Rim101 pathways that are required for the response to these stresses. This change in mRNA specificity between Rps26-containing and deficient ribosomes arises from the recognition of the -4 position of the Kozak sequence by Rps26, thereby supporting the translation of well-translated mRNAs by Rps26-containing ribosomes in rich medium, and the translation of specific mRNAs in the Hog1 and Rim101 pathways by Rps26-deficient ribosomes that are formed under these stresses. What remains unknown is how Rps26-deficient ribosomes form under high salt and high pH stress. My preliminary results suggest Rps26-deficient ribosomes are produced by release of Rps26 from pre-existing ribosomes. Therefore, I will further dissect in more detail the mechanism that leads to the production of Rps26- deficient ribosomes, exploring the role of the Rps26-specific chaperone Tsr2 in delivering to and extracting Rps26 from ribosomes (Aim1), and testing which posttranslational modifications regulate this pathway (Aim2). Next, I will test if Rps26 is (re-) incorporated into Rps26-deficient ribosomes, to allow for a rapid switch in mRNA-specificity without the costs of re-building new ribosomes, or whether instead these ribosomes are degraded (Aim3). Together, these experiments will clarify how Rps26-deficient ribosomes form under stress. In addition to further expanding on this novel paradigm of stress-induced production of a specific ribosome population, the results will also have implications for the development of diseases linked to Rps26-deficiency, such as Diamond- Blackfan anemia. Because the etiology of 10-15% of all cases remains unknown, and is not linked to RP- deficiency, it is possible that overactivation of pathways leading to the release of RPs such as Rps26 might be responsible for a subset of cases, similar to the subset of cases caused by deficiency of the Rps26-chaperone Tsr2. Furthermore, ribosomes lacking individual RPs, including Rps26, are produced in cancer cells, where they are associated with poor outcomes. Thus, this work will also help delineate how cancer cells modulate the translational machinery to subvert translation to its purposes.