Control of a Plant Translational Regulatory Protein Through Nutrient Signaling

Gallie 烟草花叶病毒RNA的5 '前导序列（称为omega）作为翻译增强子发挥功能。 一个102 kDa的蛋白质，以前确定为一个特定的欧米茄RNA结合蛋白的序列分析，揭示了同源性的HSP 101家族和其在酵母中的表达补充了由HSP 104基因的缺失引起的耐热性缺陷。 在表达烟草HSP 101的酵母中观察到ω-luc而不是luc mRNA的翻译增加高达20倍，而在没有HSP 101的情况下ω未能增强翻译。因此，HSP 101和ω组成了一个双组分的翻译调节机制，可以在酵母中重现。 酵母翻译突变体中HSP 101功能的分析表明，起始因子eIF 3和eIF 4G是HSP 101介导的增强所必需的。HSP 101的RNA结合和翻译调节活性在呼吸细胞中是无活性的，在氨基酸饥饿后，或在后期指数/静止生长期间，而其耐热功能保持不受影响。这导致了一种模型，其中HSP 101作为翻译调节因子的功能需要激活其RNA结合活性，而RNA结合活性又受营养信号传导控制。 本项目的第一个目标是通过缺失和突变分析来鉴定HSP 101作为RNA结合蛋白的功能。第二个目标将确定其RNA结合活性如何受到营养物质可用性的调节。 本研究旨在了解热休克诱导蛋白（HSP 101）如何调节烟草花叶病毒蛋白的合成。由于负责这种调节的两种组分，称为omega的RNA序列和烟草HSP 101蛋白，在酵母中表达时足以概括这种调节，因此本项目将研究HSP 101在酵母中结合RNA和调节翻译的作用。该项目将进一步研究通过营养可用性对酵母中HSP 101功能的调节。因此，这项研究将增加我们对赋予植物调节蛋白质产生能力的机制的了解。

Gallie The 5'-leader (called omega) of tobacco mosaic viral RNA functions as a translational enhancer. Sequence analysis of a 102 kDa protein, previously identified as a specific omega RNA-binding protein, revealed homology to the HSP101 family and its expression in yeast complemented a thermotolerance defect caused by a deletion of the HSP104 gene. Up to a 20-fold increase in the translation of omega-luc but not luc mRNA was observed in yeast expressing the tobacco HSP101 whereas omega failed to enhance translation in the absence of HSP101. Therefore, HSP101 and omega comprise a two-component translational regulatory mechanism that can be recapitulated in yeast. Analysis of HSP101 function in yeast translation mutants suggested that the initiation factors eIF3 and eIF4G were required for the HSP101-mediated enhancement. The RNA-binding and translational regulatory activities of HSP101 were inactive in respiring cells, following amino acid starvation, or during late-exponential/stationary growth, whereas its thermotolerance function remained unaffected. This has led to a model in which the function of HSP101 as a translational regulator requires activation of its RNA-binding activity which in turn is controlled by nutrient signaling. The first goal of this project is to investigate how HSP101 functions as an RNA-binding protein by identifying the RNA-binding site through deletion and mutation analysis. The second goal will determine how its RNA-binding activity is regulated by nutrient availability. This research seeks to understand how a protein that is induced in response to heat shock (HSP101) regulates the synthesis of tobacco mosaic virus proteins. Since the two components that are responsible for this regulation, an RNA sequence called omega and the tobacco HSP101 protein, are sufficient to recapitulate this regulation when expressed in yeast, this project will examine the role of HSP101 in binding to RNA and regulating translation in yeast. The project will further examine the regulation of HSP101 function in yeast by nutrient availability. Thus, this research will increase our understanding of a mechanism that confers on plants the ability to regulate protein production.