Modifications of Small Heat Shock Proteins in the Lens

α-晶体蛋白和热休克蛋白-27(Hsp27)是在晶状体中发现的两种小的热休克蛋白(HSPs)，被认为可以保护晶状体蛋白质免受氧化和紫外线等应激的影响。除了作为伴侣蛋白发挥作用外，它们还是一种多功能的抗凋亡蛋白。在晶状体中，甲基乙二醛(MGO)是糖酵解过程中的代谢副产物，其浓度至少是血浆中的20倍。糖尿病患者的晶状体比非糖尿病患者的晶状体含有更高的MGO浓度。氧化镁很容易与蛋白质中的赖氨酸、精氨酸和半胱氨酸残基反应，形成稳定的加合物，其中几个已经在人类晶状体中被鉴定出来。我们最近对MGO修饰的研究提供了令人惊讶的结果，即MGO修饰增强了sHsps的伴侣功能。我们最新的研究表明，白内障晶状体中含有高水平的磷酸化HSP27(PHSP27)，并且PHSP27对MGO的修饰非常敏感。我们还发现，氧化镁修饰的α-晶状体蛋白是一种比天然未修饰蛋白更好的抗凋亡蛋白。这些有趣的观察促使我们进一步研究MGO诱导的sHsps结构变化，并确定这种变化是如何改变其功能的。我们有四个目标。在目标1中，我们将详细研究sHSP暴露在MGO下所造成的结构变化。我们将重点关注精氨酸残留物，因为它们是最容易被MGO修饰的。在目标2中，我们将确定MGO诱导的修饰如何改变sHSPs的抗凋亡功能。在目标3中，我们将研究氧化镁诱导的修饰对sHSPs与其他晶状体蛋白相互作用的影响。最后，在目标4中，我们将确定在细胞压力增加时，sHSPs的MGO修饰是否会影响它们向细胞核的运输。这些研究将加深我们对sHsps与代谢途径之间相互作用的理解，它们将有助于我们确定sHsps在晶状体上皮细胞凋亡中的作用以及在白内障形成中的意义。

Alpha-crystallin and heat shock protein-27 (Hsp27), the two small heat shock proteins (Hsps) found in the lens are thought to protect lens proteins from the stress, such as, oxidation and ultra violet light. In addition to functioning as chaperone proteins, they are versatile anti-apoptotic proteins. In the lens methylglyoxal (MGO) is produced as a metabolic by-product during glycolysis and its concentration is at least 20 times higher than in plasma. Lenses from diabetics have far greater MGO concentrations than lenses from nondiabetic individuals. MGO readily reacts with lysine, arginine and cysteine residues in proteins to form stable adducts, several of which have been identified in the human lens. Our recent studies on MGO modifications provided the surprising results that MGO-modification enhances the chaperone function of sHsps. Our most recent studies demonstrate that cataractous lenses contain high levels of phosphorylated Hsp27 (pHsp27) and pHsp27 is highly susceptible for modification by MGO. We also found MGO-modified alpha-crystallin is a better anti-apoptotic protein than the native unmodified protein. These intriguing observations prompted us to examine further on the MGO-induced structural alterations in sHsps, and to determine how such alterations change their functions. We have four aims. In aim 1, we will examine in detail the structural changes resulting from exposure of sHsps to MGO. We will focus on arginine residues, because these are the most vulnerable to modification by MGO. In aim 2, we will determine how MGO-induced modifications alter the anti-apoptotic functions of sHsps. In aim 3, we will study the impact of MGO-induced modifications on the interaction of sHsps with other lens proteins. Finally, in aim 4, we will determine whether MGO-modification of sHsps influence their transportation into the nucleus during increased cellular stress. These studies will improve our understanding of the interplay between sHsps and metabolic pathways, and they will help us to define the role of sHsps in apoptosis of lens epithelial cells and the implications for cataract formation.