Role of the S100 Family of Proteins in Lens Physiology and Cataract

项目总结 人工晶状体在将入射光聚焦到视网膜上起着至关重要的作用，晶状体透明度会发生像差。 以及导致视力受损的机械特性。尽管我们继续努力，但我们对 调控晶状体细胞结构、内稳态、变形性、清晰度和 白内障的发病机制尚不完全清楚。我们最近发现，S100的几名成员 钙结合蛋白家族在晶状体中大量表达，并表现出不同的分布特征。 S100A6和S100A10在晶状体上皮细胞和晶状体纤维中均有表达，S100A4在晶状体上皮细胞和晶状体纤维中均有表达 分散到晶状体纤维细胞。缺乏S100A4而不是S100A6或S100A10会导致乳白色晚发 小鼠白内障模型的形成以及S100A5基因的表达上调 否则在野生型镜头中是无法检测到的。除了S100A4上的这些新数据，我们目前没有 了解S100家族蛋白质在晶状体功能中的作用。S100A4在细胞中表达， 组织特有的方式，并参与各种疾病的病理生理学。的生理功能 然而，人们对S100A4却知之甚少。S100A4的一个众所周知的分子功能是它与 非肌肉肌球蛋白II(NM II)，特别是NM II IA，以及对NM II组装、肌动蛋白聚合、 收缩特性、细胞迁移和可塑性。NM II在晶状体细胞结构中起着至关重要的作用， 机械和功能。尽管NM IIA突变与人类白内障的发展有关，但很少 已知在晶状体功能的背景下调节NM II活动。而镜片是从成熟的 S100A4裸鼠晶状体(2至6个月大)保持透明，并保持正常生长和大小，与6个月大的小鼠相同 一个月龄小鼠表现出NM IIA的组装和磷酸化，肌动蛋白的聚合，以及 水通道蛋白-5、连接蛋白-50、细胞黏附蛋白(整合素-β1、NrCAM、ZO-1)水平的变化 和α-晶体蛋白，最终导致八个月大的小鼠形成乳白色白内障。基于这些 初步和新的发现，我们假设S100A4在调节NM II组装中发挥关键作用 S100A4的缺失扰乱了NM II的组装和磷酸化，肌动蛋白聚合， 机械性能和晶状体中的渗透平衡，导致迟发性白内障的形成。为了测试这一点 假设，在三个相互关联的具体目标中，我们将研究：1)S100A4在调节中的机制作用 晶状体NM II组装和磷酸化、肌动蛋白聚合、细胞黏附和晶状体拉伸特性，2) 白内障发病机制中渗透调节和溶质载体转运机制的破坏 S100A4缺失晶状体；3)S100A4缺失小鼠晶状体纤维中S100A4表达的挽救作用(使用 转基因方法)对细胞骨架完整性和晶状体表型的影响。这些新颖且相互关联的研究是 预计将揭示S100A4和S100家族其他成员蛋白在晶状体功能中的重要性， 已知它们参与了几种疾病的病理生理学。

PROJECT SUMMARY The ocular lens plays a crucial role in focusing incident light on to the retina, with aberrations in lens transparency and mechanical properties leading to impairment of vision. Despite continuing effort, our understanding of the various molecular pathways governing lens cytoarchitecture, homeostasis, deformability, clarity, and the etiological mechanisms of cataract remains incomplete. We recently identified that several members of the S100 family of calcium binding proteins are abundantly expressed and exhibit distinct distribution profiles in the lens. While expression of S100A6 and S100A10 distributed to both the lens epithelium and fibers, S100A4 distributes discretely to lens fiber cells. Absence of S100A4 but not S100A6 or S100A10 leads to opalescent late-onset cataract formation in a mouse model, together with the robust upregulation of S100A5, the expression of which was otherwise undetectable in the wild type lens. Other than these new data on S100A4, we currently have no knowledge of the role played by the S100 family of proteins in lens function. S100A4 is expressed in a cell and tissue specific manner and is involved in the pathophysiology of various diseases. The physiological function of S100A4 however, is poorly understood. A well-understood molecular function of S100A4 is its interaction with non-muscle myosin II (NM II), especially NM IIA, and regulation of NM II assembly, actin polymerization, contractile characteristics, cell migration, and plasticity. NM II plays a crucial role in lens cytoarchitecture, mechanics, and function. Though NM IIA mutations are associated with cataract development in humans, little is known about regulation of NM II activity in the context of lens function. While lenses derived from mature S100A4 null mouse lenses (2 to 6 month old) remain clear and maintain normal growth and size, those from six month-old mice exhibit impairments in assembly and phosphorylation of NM IIA, polymerization of actin, and alterations in the levels of aquaporin-5, CLIC5, connexin-50, cell adhesive proteins (integrin-β1, NrCAM, ZO-1) and α-crystallin, culminating in opalescent cataract formation in eight month-old mice. Based on these preliminary and novel findings, we hypothesize that S100A4 plays a crucial role in regulating NM II assembly in the lens, and that the absence of S100A4 disrupts NM II assembly and phosphorylation, actin polymerization, mechanical properties, and osmotic homeostasis in the lens, leading to late onset cataract formation. To test this hypothesis, in three interrelated specific aims, we will investigate: 1) the mechanistic role of S100A4 in regulation of lens NM II assembly and phosphorylation, actin polymerization, cell adhesion, and lens tensile properties, 2) the disruption of osmoregulation and solute carrier transport mechanisms underlying cataractogenesis in S100A4 null lenses, and 3) the effects of rescuing expression of S100A4 in S100A4 null mouse lens fibers (using a transgenic approach) on cytoskeletal integrity and lens phenotype. These novel and interrelated studies are expected to uncover the significance of S100A4 and other members of the S100 family proteins in lens function, which are known to participate in the pathophysiology of several diseases.