Calcium-activated protease in cataract formation

描述（由申请人提供）：该基金的重点是提供钙激活蛋白酶（钙蛋白酶）在透镜中作用的详细机制。 正在测试的总体假设是，钙蛋白酶的蛋白水解是必不可少的有限的截断，截短的β-晶体蛋白的更紧密的包装，并在正常的透镜纤维分化和成熟过程中发生的细胞骨架元素的重排。 在白内障形成过程中，由于大量钙离子的涌入，钙蛋白酶的过度激活产生快速的蛋白水解和光散射，因为截短的晶体蛋白不再正确有序。 在最近的资助周期中的研究产生了几个与这一假设相关的意想不到的发现：晶状体特异性钙蛋白酶Lp 82（肌肉p94的剪接变体，而不是m-钙蛋白酶）的活性与年轻啮齿动物的白内障形成密切相关。 此外，发现钙蛋白酶10是人透镜中普遍存在的第三种钙蛋白酶，并且钙蛋白酶10在大鼠的糖尿病性白内障中显著增加。 Lp 82、Lp 85（透镜中的低丰度剪接变体）和钙蛋白酶10可在适当的透镜成熟或病理性白内障形成所必需的独特位点截短特异性透镜蛋白或切割蛋白。 为了检验这些想法，将追求三个具体目标： 1.确定Lp 82的生化性质和底物。 2.确定p94变体是否承担啮齿动物Lp 82在人类晶状体中的作用，并负责人类晶状体中无法解释的蛋白水解。 3.明确钙蛋白酶10在人透镜中的作用。 这些研究将为使用蛋白酶抑制剂预防白内障提供合理的依据。

DESCRIPTION (provided by applicant): This grant is focused on providing a detailed mechanism for the role of calcium-activated proteases (calpains) in lens. The overall hypothesis being tested is that proteolysis by calpains is essential for limited truncation, tighter packing of truncated beta-crystallins, and rearrangement of the cytoskeletal elements occurring during normal lens fiber differentiation and maturation. During cataract formation, over-activation of calpains by massive influx of calcium produces rapid proteolysis and light scattering because the truncated crystallins are no longer properly ordered. Studies during the latest grant cycle produced several unexpected findings relevant to this hypothesis: Activity of lens-specific calpain Lp82, a splice variant of muscle p94, and not m-calpain, was most strongly associated with cataract formation in young rodents. Further, calpain 10 was discovered as a third ubiquitous calpain in human lens, and calpain 10 was markedly increased in diabetic cataracts in rats. Lp82, Lp85 (a low-abundance splice variant in lens), and calpain 10 may truncate specific lens proteins or cleave proteins at unique sites necessary for proper lens maturation or pathologic cataract formation. To test these ideas, three specific aims will be pursued: 1. Determine the biochemical properties and substrates of Lp82. 2. Determine if p94 variants assume the role of rodent Lp82 in human lenses and are responsible for unexplained proteolysis in human lenses. 3. Define the roles of calpain 10 in human lens. These studies will provide a rational basis for the use of protease inhibitors to prevent cataracts.