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

我们以前报道，分泌缺陷引起的核糖体蛋白和rRNA基因的特异性和显着的转录抑制。使用各种sec突变体的进一步检查表明，分泌途径中任何地方的缺陷，从新生肽插入内质网之前的步骤到涉及质膜形成的步骤，都会阻止核糖体组分的继续合成。我们提出存在从质膜到细胞核的信号转导途径。本研究构建了一个由核糖体蛋白基因启动子驱动的HIS 3基因表达质粒，并对HIS 3基因的表达进行了初步研究。将该质粒转化到酵母温度敏感sec突变体sly 1（his 3）中。我们分离出几个突变体，似乎是有缺陷的信号通路。其中一个突变体rrs 1 -1是冷敏感的，并且rrs 1 -1突变大大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。RRS 1是一个新的、必需的基因，编码203个氨基酸残基的核蛋白，在真核生物中是保守的。序列分析表明rrs 1 -1在密码子114内只有一个核苷酸的差异，导致一个终止科登。截短的rrs 1 -1蛋白与野生型蛋白相似。通过将RRS 1置于GAL 1启动子的控制下，构建了条件性rRS 1无效突变体。Rrs 1缺失导致前体RNA加工和核糖体亚基组装缺陷。这些结果表明，Rrs 1是必不可少的核糖体生物合成，也有一个重要的作用，在分泌缺陷引起的信号。

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.