Functional Analysis of Optineurin in Glaucoma

Optineurin 在青光眼中的功能分析

• 批准号: 6888027
• 负责人: Mansoor Sarfarazi
• 金额: $ 32.63万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-05-01 至 2008-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6888027
• 关键词: clinical research; developmental genetics; developmental neurobiology; disease /disorder etiology; gene expression; gene mutation; genetic screening; genetic susceptibility; genotype; glaucoma; human subject; immunocytochemistry; in situ hybridization; laboratory mouse; messenger RNA; patient oriented research; phenotype; protein localization; protein protein interaction; protein quantitation /detection; protein structure function; transmission electron microscopy; yeast two hybrid system

DESCRIPTION (provided by applicant): Glaucoma is an optic neuropathy that affects over 67 million people worldwide. This condition has broad clinical subtypes ranging from birth to very late in life. The most common form, Primary Open Angle Glaucoma (POAG) has a prevalence of about 1% of white population over 40 years of age. Several genetic loci have been identified for POAG but so far only mutations in Myocilin gene are reported in juvenile-onset and certain other adult-onset cases. Recently, we identified a gene that is primarily involved in a subgroup of Adult-Onset POAG, commonly known as Normal Tension Glaucoma (NTG). This gene that we named Optineurin (OPTN) is mutated in 16.7% of our hereditary NTG families, maps to the GLC1E locus on 10p14, has 13 coding exons and encodes for a 577-amino acids protein (~66-kDa). Our OPTN protein studies showed co-localization with Golgi, secretion into aqueous humor and expression in many ocular and non-ocular tissues. Cloning of both mouse and monkey genes showed very similar patterns of mRNA and protein expression to human OPTN. It has also been shown by other investigators that OPTN interacts with Ad E3-14.7K, Huntingtin, TFIIIA, RAB8 and 2 other unknown kinases. Although existing evidence suggest that OPTN through its interaction with other proteins may be utilizing TNF-a or Fas-Ligand pathways to mediate apoptosis, inflammation or vasoconstriction but, as yet, these is no clear indication on how OPTN mutations lead to either NTG or POAG. Therefore, as an initial step towards understanding the ocular function of this gene and its protein products we propose the followings. 1)-Screen a large number of glaucoma patients for OPTN mutations in order to establish a genotype/phenotype correlation; 2)-Use OPTN as "bait" and search for new interacting proteins by Yeast Two Hybrid System and to identify motifs that are important for OPTN function; 3)-Determine ultracellular localization of OPTN in normal and glaucomatous eyes by immunogold labeling; 4)-Study OPTN distribution in normal and glaucomatous eyes by immunohistochemistry and to determine its potential differential tissue expression patterns; and 5)-Use In Situ hybridization to study developmental expression patterns of Optn in mouse embryos. At the conclusion of this investigation, it is anticipated that our study will provide essential scientific information that one day may contribute to the design of innovative and futuristic drug intervention for this group of optic neuropathies.

描述（由申请人提供）：青光眼是一种视神经病变，影响全球超过 6700 万人。这种疾病有广泛的临床亚型，从出生到晚年。最常见的形式是原发性开角型青光眼 (POAG)，40 岁以上白人的患病率约为 1%。已鉴定出 POAG 的几个遗传位点，但迄今为止仅报道了青少年发病和某些其他成人发病病例中 Myocilin 基因的突变。最近，我们发现了一个主要与成人发病的 POAG 亚组有关的基因，通常称为正常眼压性青光眼 (NTG)。这个我们命名为 Optineurin (OPTN) 的基因在我们遗传性 NTG 家族中的 16.7% 中发生突变，映射到 10p14 上的 GLC1E 基因座，具有 13 个编码外显子，编码 577 个氨基酸的蛋白质 (~66-kDa)。我们的 OPTN 蛋白研究表明，它与高尔基体共定位，分泌到房水中，并在许多眼部和非眼部组织中表达。小鼠和猴子基因的克隆显示出与人类 OPTN 非常相似的 mRNA 和蛋白质表达模式。其他研究人员还表明，OPTN 与 Ad E3-14.7K、亨廷顿蛋白、TFIIIA、RAB8 和其他 2 种未知激酶相互作用。尽管现有证据表明 OPTN 通过与其他蛋白质的相互作用可能利用 TNF-a 或 Fas-Ligand 途径介导细胞凋亡、炎症或血管收缩，但到目前为止，还没有明确的迹象表明 OPTN 突变是如何发生的 导致 NTG 或 POAG。因此，作为了解该基因及其蛋白质产物的眼部功能的第一步，我们提出以下建议。 1)-对大量青光眼患者进行OPTN突变筛查，以建立基因型/表型相关性； 2)-以OPTN为“诱饵”，通过酵母二杂交系统寻找新的相互作用蛋白，并鉴定对OPTN功能重要的基序； 3)-通过免疫金标记确定OPTN在正常眼和青光眼眼中的超细胞定位； 4)-通过免疫组织化学研究OPTN在正常眼和青光眼眼中的分布，并确定其潜在的差异组织表达模式； 5)-使用原位杂交研究小鼠胚胎中 Optn 的发育表达模式。在这项调查结束时，预计我们的研究将提供重要的科学信息，有一天可能有助于设计针对这组视神经病的创新和未来药物干预措施。