Analysis of yeast mutants defective in a signal transduction pathway from the plasma membrane to the nucleus

我们之前报道了分泌缺陷导致核糖体蛋白和rRNA基因特异性和显著的转录抑制。使用各种sec突变体的进一步检查表明，从新生肽预先插入内质网到参与质膜形成的分泌途径中的任何地方都存在缺陷，从而阻止核糖体成分的继续合成。我们提出存在从质膜到细胞核的信号转导途径。本研究的目的是阐明该信号的分子机制。生成了一个质粒，其中HIS3基因由核糖体蛋白基因的启动子驱动。将该质粒转化为酵母温度敏感的sec突变体sly1（his3）。我们分离了几个似乎在信号通路上有缺陷的突变体。其中一个突变体rrs1-1是冷敏感的，rrs1-1突变极大地降低了由分泌缺陷引起的rRNA和核糖体蛋白基因的转录抑制。RRS1是一种在真核生物中保守的含有203个氨基酸残基的核蛋白的重要基因。序列分析表明，rrs1-1在密码子114内只有一个核苷酸差异，导致一个停止密码子。截断的rrs1-1蛋白似乎与野生型蛋白相似。通过将RRS1置于GAL1启动子的控制下，构建了一个条件RRS1零突变体。Rrs1缺失导致前rna加工和核糖体亚基组装缺陷。这些结果表明，Rrs1对核糖体的生物发生至关重要，并且在分泌缺陷引起的信号传导中也起着重要作用。

We previously reported that a secretory defect caused specific and significant transcriptional repression of both ribosomal protein and rRNA genes. Futher examination using various sec mutants showed that a defect anywhere in the secretory pathway, from a step prior insertion of the nascent peptide into the endoplasmic reticulum to step involved in the formation of the plasma membrane, prevents the continued synthesis of the components of the ribosome. We propose the existence of a signal transduction pathway from the plasma membrane to the nucleus. The purpose of this research is to elucidate the molecular machanism of the signaling.A plasmid was generated in which the HIS3 gene was driven by the promoter of a ribosomal protein gene. This plasmid was transformed into a yeast temperature sensitive sec mutant, sly1(his3). We isolated several mutants which appeared to be defective in the signaling pathway. One of the mutants, rrs1-1, was cold sensitive and the rrs1-1 mutation greatly reduced transcriptional repression of both rRNA and ribosomal protein genes that is caused by secretory defect. RRS1 is a novel, essential gene encoding a nuclear protein of 203 amino acid residues that is conserved in eukaryotes. Sequence analysis showed that rrs1-1 had only one nucleotide difference, within codon 114, resulting in a stop coden. The truncated rrs1-1 protein appeared to have similar to the wild -type protein. A conditional rrs1-null mutant was constructed by placing RRS1 under the control of the GAL1 promoter. Rrs1 depletion caused defects in processing of pre-RNA and assembly of ribosomal subunits. These results suggest that Rrs1 is essential for ribosome biogenesis and also has an important role in the signaling caused by a secretory defect.