Nonoptimal Codon Usage Is Critical for Protein Structure and Function of the Master General Amino Acid Control Regulator CPC-1.
Nonoptimal Codon Usage Is Critical for Protein Structure and Function of the Master General Amino Acid Control Regulator CPC-1.
复制标题
非最佳密码子使用对于主要通用氨基酸控制调节器 CPC-1 的蛋白质结构和功能至关重要
作者：
Lyu X;Liu Y
一般的氨基酸控制反应是生物体适应氨基酸饥饿条件的关键。偏爱使用某些同义密码子是所有基因组的普遍特征。此前，同义密码子的改变被认为是沉默的突变。在这项研究中，我们发现与基因组中的其他基因相比，脉孢子菌CPC-1基因具有不寻常的密码子使用情况。我们发现，在不改变其氨基酸序列的情况下对CPC-1基因进行密码子优化会导致CPC-1表达增加，蛋白质降解率改变，并由于蛋白质结构的改变而削弱蛋白质的功能。综上所述，这些结果揭示了同义密码子使用在调节CPC-1表达和功能中的关键作用，并建立了密码子使用在蛋白质结构中重要性的遗传学例子。摘要在氨基酸饥饿条件下，真核生物激活一种普遍的氨基酸控制反应。在粗糙脉孢菌中，交叉通路控制蛋白1(CPC-1)是酿酒酵母bZIP转录因子Gcn4的同源基因，是一般氨基酸控制反应的主要调节因子。密码子使用偏向是真核生物基因组的普遍特征，对基因表达的调控至关重要。虽然密码子的使用也与蛋白质结构和功能的调节有关，但支持这一结论的遗传证据非常有限。在这里，我们表明，脉孢霉CPC-1有一个非最佳的NNU丰富的密码子使用谱，与基因组中强烈的NNC密码子偏好形成对比。虽然用同义的NNC密码子替换CPC-1的NNU密码子提高了CPC-1在脉孢霉中的表达，但它改变了CPC-1的降解速率，并取消了氨基酸饥饿诱导的蛋白质稳定。密码子操纵的CPC-1蛋白对有限的蛋白酶消化也表现出不同的敏感性。此外，CPC-1在挽救CPC-1缺失突变体的细胞生长和激活其靶基因表达方面的功能因同义密码子的改变而受损。综上所述，这些结果揭示了密码子使用在调节CPC-1表达和功能中的关键作用，并建立了密码子使用在蛋白质折叠中重要性的遗传学例子。重要性一般的氨基酸控制反应对于生物体适应氨基酸饥饿条件是至关重要的。偏爱使用某些同义密码子是所有基因组的普遍特征。此前，同义密码子的改变被认为是沉默的突变。在这项研究中，我们发现与基因组中的其他基因相比，脉孢子菌CPC-1基因具有不寻常的密码子使用情况。我们发现，在不改变其氨基酸序列的情况下对CPC-1基因进行密码子优化会导致CPC-1表达增加，蛋白质降解率改变，并由于蛋白质结构的改变而削弱蛋白质的功能。综上所述，这些结果揭示了同义密码子使用在调节CPC-1表达和功能中的关键作用，并建立了密码子使用在蛋白质结构中重要性的遗传学例子。
The general amino acid control response is critical for adaptation of organisms to amino acid starvation conditions. The preference to use certain synonymous codons is a universal feature of all genomes. Synonymous codon changes were previously thought to be silent mutations. In this study, we showed that the Neurospora cpc-1 gene has an unusual codon usage profile compared to other genes in the genome. We found that codon optimization of the cpc-1 gene without changing its amino acid sequence resulted in elevated CPC-1 expression, an altered protein degradation rate, and impaired protein functions due to changes in protein structure. Together, these results reveal the critical role of synonymous codon usage in regulation of CPC-1 expression and function and establish a genetic example of the importance of codon usage in protein structure. ABSTRACT Under amino acid starvation conditions, eukaryotic organisms activate a general amino acid control response. In Neurospora crassa, Cross Pathway Control Protein 1 (CPC-1), the ortholog of the Saccharomyces cerevisiae bZIP transcription factor GCN4, functions as the master regulator of the general amino acid control response. Codon usage biases are a universal feature of eukaryotic genomes and are critical for regulation of gene expression. Although codon usage has also been implicated in the regulation of protein structure and function, genetic evidence supporting this conclusion is very limited. Here, we show that Neurospora cpc-1 has a nonoptimal NNU-rich codon usage profile that contrasts with the strong NNC codon preference in the genome. Although substitution of the cpc-1 NNU codons with synonymous NNC codons elevated CPC-1 expression in Neurospora, it altered the CPC-1 degradation rate and abolished its amino acid starvation-induced protein stabilization. The codon-manipulated CPC-1 protein also exhibited different sensitivity to limited protease digestion. Furthermore, CPC-1 functions in rescuing the cell growth of the cpc-1 deletion mutant and activation of the expression of its target genes were impaired by the synonymous codon changes. Together, these results reveal the critical role of codon usage in regulation of CPC-1 expression and function and establish a genetic example of the importance of codon usage in protein folding. IMPORTANCE The general amino acid control response is critical for adaptation of organisms to amino acid starvation conditions. The preference to use certain synonymous codons is a universal feature of all genomes. Synonymous codon changes were previously thought to be silent mutations. In this study, we showed that the Neurospora cpc-1 gene has an unusual codon usage profile compared to other genes in the genome. We found that codon optimization of the cpc-1 gene without changing its amino acid sequence resulted in elevated CPC-1 expression, an altered protein degradation rate, and impaired protein functions due to changes in protein structure. Together, these results reveal the critical role of synonymous codon usage in regulation of CPC-1 expression and function and establish a genetic example of the importance of codon usage in protein structure.