Stress Granule-Defective Mutants Deregulate Stress Responsive Transcripts

To reduce expression of gene products not required under stress conditions, eukaryotic cells form large and complex cytoplasmic aggregates of RNA and proteins (stress granules; SGs), where transcripts are kept translationally inert. The overall composition of SGs, as well as their assembly requirements and regulation through stress-activated signaling pathways remain largely unknown.
We have performed a genome-wide screen of S. cerevisiae gene deletion mutants for defects in SG formation upon glucose starvation stress. The screen revealed numerous genes not previously implicated in SG formation. Most mutants with strong phenotypes are equally SG defective when challenged with other stresses, but a considerable fraction is stress-specific. Proteins associated with SG defects are enriched in low-complexity regions, indicating that multiple weak macromolecule interactions are responsible for the structural integrity of SGs. Certain SG-defective mutants, but not all, display an enhanced heat-induced mutation rate. We found several mutations affecting the Ran GTPase, regulating nucleocytoplasmic transport of RNA and proteins, to confer SG defects. Unexpectedly, we found stress-regulated transcripts to reach more extreme levels in mutants unable to form SGs: stress-induced mRNAs accumulate to higher levels than in the wild-type, whereas stress-repressed mRNAs are reduced further in such mutants.
Our findings are consistent with the view that, not only are SGs being regulated by stress signaling pathways, but SGs also modulate the extent of stress responses. We speculate that nucleocytoplasmic shuttling of RNA-binding proteins is required for gene expression regulation during stress, and that SGs modulate this traffic. The absence of SGs thus leads the cell to excessive, and potentially deleterious, reactions to stress.

为降低应激条件下非必需基因产物的表达，真核细胞可形成由RNA与蛋白质构成的大型复杂细胞质聚集体——应激颗粒（stress granules, SGs），其中滞留的转录本处于翻译沉默状态。目前，关于SGs的整体组成、组装所需条件，以及其通过应激激活信号通路实现调控的机制，学界仍知之甚少。

本研究针对酿酒酵母（S. cerevisiae）的基因缺失突变体开展全基因组筛选，以鉴定葡萄糖饥饿应激下SG形成存在缺陷的菌株。筛选结果揭示了诸多此前未被关联至SG形成过程的基因。多数表现出强表型的突变体，在遭受其他类型应激时同样存在SG形成缺陷，但亦有相当比例的突变体仅在特定应激下表现出缺陷表型。与SG形成缺陷相关的蛋白质普遍富集低复杂度区域，提示多重弱大分子相互作用是维持SG结构完整性的核心基础。部分（而非全部）SG缺陷突变体呈现出热诱导突变率升高的特征。本研究还发现若干影响Ran GTP酶（Ran GTPase）的突变可导致SG形成缺陷，该酶负责调控RNA与蛋白质的核质运输过程。值得注意的是，我们观察到无法形成SG的突变体中，应激调控转录本的表达水平出现极端偏移：应激诱导的信使RNA（mRNA）积累量显著高于野生型菌株，而应激抑制的mRNA在这类突变体中的丰度则进一步降低。

本研究结果支持如下学术观点：SG不仅受应激信号通路调控，同时也可调节应激反应的强度。我们推测，应激过程中的基因表达调控需要RNA结合蛋白的核质穿梭，而SG可对这一运输过程进行调控。因此，SG的缺失会导致细胞对应激产生过度且潜在有害的反应。