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

该研究的长期目标是从生理和机制两个层面上确定植物小分子热激蛋白（sHsp）的功能。sHsps是一组普遍存在的应激蛋白，据信其充当分子伴侣以防止其他蛋白质的不可逆变性/聚集。sHsp分子伴侣的功能对于植物在许多不同种类的胁迫下的存活以及植物生长和发育的特定阶段是潜在的关键。植物的特征在于属于sHsp家族的蛋白质的不寻常的复杂性;植物表达多种胞质sHsp和靶向细胞内细胞器的特异性sHsp同种型。sHsp家族在植物中的进化扩展表明，sHsp在许多情况下随着时间的推移提供了选择优势。拟南芥sHsp家族的完整定义，沿着我们对sHsp结构和分子伴侣作用的研究，为研究sHsp功能提供了一个新的起点。利用分子遗传学和生物化学方法相结合，在拟南芥中，我们将定义特定的sHsps在胁迫耐受性，生长和发育中的生理作用，并确定特定的sHsp底物。拟议的研究有三个具体目标。目的1：确定sHsp功能缺失和获得植物的生长和胁迫耐受表型。RNA干扰转基因和/或T-DNA插入突变体的基因代表类I/II胞质sHsps，以及植物组成型表达类I和/或II sHsps，将表征生长和胁迫耐受性。目的2：鉴定sHsp的潜在底物。我们将测试sHsps与广泛的蛋白质底物结合以及每种sHsps类别与独特底物相互作用的假设。目的3：确定sHsp功能丧失突变体的其他生化表型。通过这些实验，最终有可能测试长期存在的关于sHsps对植物胁迫耐受性和发育的重要性的假设，并发现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.