Function of the Small Hsps in Stress Tolerance, Growth and Development

这项研究的长期目标是从生理和机制水平上确定植物小热休克蛋白(SHSP)的功能。SHSPs是一组普遍存在的应激蛋白，被认为是分子伴侣，可以防止其他蛋白质的不可逆变性/聚集。SHSP分子伴侣功能对植物在多种逆境下的生存和植物生长发育的特定阶段具有潜在的关键作用。植物具有不寻常的复杂性，属于SHSP家族的蛋白质；植物既表达多种胞质sHSP，也表达针对细胞内细胞器的特定SHSP亚型。SHSP家族在植物中的进化扩展表明，随着时间的推移，sHSP在许多环境中提供了一种选择性优势。拟南芥SHSP家族的完整定义，以及我们在体外对SHSP结构和伴侣作用的研究建立的生化框架，为研究SHSP的功能提供了一个新的起点。利用分子遗传学和生物化学相结合的方法，我们将确定特定的sHSP在逆境耐受性、生长和发育中的生理作用，并鉴定特定的SHSP底物。这项拟议的研究有三个具体目标。目的1：测定SHSP功能丧失和功能获得植物的生长和抗逆性表型。代表I类和Amp；II类胞质sHsps基因的RNA干扰转基因和/或T-DNA插入突变体，以及组成表达I类和/或II类sHsps的植物，将以生长和抗逆性为特征。目的2：寻找潜在的SHSP底物。我们将检验这样的假设，即sHSP结合了广泛的蛋白质底物，并且每个SHSP类别都与独特的底物相互作用。目的3：确定SHSP功能缺失突变体的其他生化表型。通过这些实验，最终将有可能检验关于sHSPs对植物抗逆性和发育的重要性的长期假说，并揭示这些独特的伴侣蛋白的新功能。

The long term goal of the proposed research is to define plant small heat shock protein (sHsp) function at both physiological and mechanistic levels. sHsps are a ubiquitous group of stress proteins that are believed to act as molecular chaperones to prevent irreversible denaturation/aggregation of other proteins. sHsp chaperone function is potentially critical to plant survival of many different kinds of stress and to specific stages of plant growth and development. Plants are characterized by an unusual complexity of proteins belonging to the sHsp family; plants express both multiple cytosolic sHsps and specific sHsp isoforms targeted to intracellular organelles. The evolutionary expansion of the sHsp family in plants indicates that sHsps have provided a selective advantage over time in many contexts. The complete definition of the sHsp family from Arabidopsis, along with the biochemical framework from our studies of sHsp structure and chaperone action in vitro, provide a new starting point for investigations of sHsp function. Using a combination of molecular genetic and biochemical approaches in Arabidopsis thaliana we will define the physiological role of specific sHsps in stress tolerance, growth and development, and identify specific sHsp substrates. The proposed research has three specific aims. Aim 1: To determine the growth and stress tolerance phenotypes of sHsp loss- and gain-of-function plants. RNA interference transgenics and/or T-DNA insertional mutants for genes representing the class I & II cytosolic sHsps, as well as plants constitutively expressing class I and/or II sHsps, will be characterized for growth and stress tolerance. Aim 2: To identify potential sHsp substrates. We will test the hypothesis that sHsps bind a wide range of protein substrates and that each sHsp class interacts with unique substrates. Aim 3: To define other biochemical phenotypes of sHsp loss-of-function mutants. With these experiments it will finally be possible to test long-standing hypotheses about the importance of sHsps to plant stress tolerance and development and to uncover new functions for these unique chaperones.