ALPHA-CRYSTALLIN FUNCTION IN LENS BIOLOGY

描述（由申请人提供）：白内障形成是视力丧失的最常见原因，占全球失明病例的51%。在美国，白内障手术每年花费医疗保险系统约50亿美元。流行病学研究表明，人类白内障的发病机制涉及遗传、环境和其他疾病相关的危险因素。特别是，50%的儿童白内障病例有遗传基础。透镜晶体蛋白家族占透镜蛋白的90%，在透镜的透明度中起关键作用。研究已经确定了编码α-、β-和γ-晶体蛋白的基因中的点突变，其导致在出生时或在早期形成遗传性人类白内障。对遗传性白内障形成的功能研究可以为年龄相关性白内障的病因学研究提供重要信息。α-晶状体蛋白是两种多肽α A-和α β-晶状体蛋白的聚集体，其在透镜上皮细胞和纤维细胞中表达。在α A-和α β-晶体蛋白基因中携带单点突变的人类患者发展为遗传性白内障。为了了解遗传性白内障的病因，我们使用基于胚胎干细胞的技术来产生表达α-晶体蛋白中含有α A-R49 C或α B-R120 G突变的蛋白质的基因敲入小鼠。这两个突变与人类常染色体显性遗传性白内障有关。我们还在研究一种同时缺乏α A-和α β-晶体蛋白的基因敲除小鼠。这些小鼠模型在出生后早期就发生白内障，是了解疾病过程的重要工具。我们的第一个目标是验证α A-晶状体蛋白突变或缺失导致出生后早期发育中的透镜中组蛋白和代谢酶表达上调的假设。第二个目标是检验突变的α A-或α β-晶体破坏正常的透镜蛋白质稳态，通过自噬导致异常蛋白质丢失的假设。为了实现这些目标，我们将使用互补的生物化学，细胞生物学和遗传学方法，这些方法是定量的，客观的，不受观察者偏见的影响。我们的研究结果将为透镜发育和白内障形成的分子基础提供新的见解，并促进延缓或预防白内障的策略的发展。

DESCRIPTION (provided by applicant): Cataract formation is the most common cause of vision loss, accounting for 51% of cases of blindness worldwide. In the US, cataract operations cost the Medicare system approximately $5 billion annually. Epidemiologic studies show that the pathogenesis of human cataracts involves genetic, environmental, and other disease-associated risk factors. In particular, 50% of childhood cataract cases have a genetic basis. The lens crystallins protein family accounts for 90% of lens proteins and plays a key role in lens transparency. Research has identified point mutations in the genes encoding α-, ß-, and γ-crystallins, which lead to hereditary human cataract formation either at birth or at an early age. Functional studies on hereditary cataract formation could provide important information about the etiology of age-related cataracts. α-crystallin is an aggregate of two polypeptides, αA- and αß-crystallin, that are expressed in lens epithelial and fiber cells. Human patients harboring single point mutations in αA- and αß-crystallin genes develop hereditary cataracts. To understand disease etiology in hereditary cataracts, we have used embryonic stem cell-based technologies to generate knock-in mice expressing proteins containing either the αA-R49C or αB-R120G mutation in α-crystallins. These two mutations are associated with human autosomal dominant hereditary cataracts. We are also studying a knockout mouse lacking both αA- and αß-crystallin. These mouse models develop cataracts at an early postnatal age and are important tools for understanding the disease process. Our first aim will test the hypothesis that αA-crystallin mutation or deletion causes upregulation of histone and metabolic enzyme expression in the developing lens at an early postnatal age. The second aim will test the hypothesis that mutant αA- or αß-crystalline disrupts normal lens protein homeostasis, leading to abnormal protein loss through autophagy. To address these aims, we will use complementary biochemical, cell biological and genetic approaches which are quantitative, objective and not subject to observer bias. The results of our studies will provide new insights into the molecular basis of lens development and cataract formation and promote the development of strategies to delay or prevent cataracts.