Functional Analysis of Optineurin in Glaucoma

Optineurin 在青光眼中的功能分析

• 批准号: 7061198
• 负责人: Mansoor Sarfarazi
• 金额: $ 31.86万
• 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/7061198
• 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，从出生到生命的晚期。原发性开角型青光眼（POAG）是最常见的形式，在40岁以上的白色人群中的患病率约为1%。POAG的几个基因位点已被确定，但到目前为止，只有Myocilin基因突变的报告，在青少年发病和某些其他成人发病的情况下。最近，我们发现了一个基因，主要涉及一个亚组的成人发病POAG，通常被称为正常眼压性青光眼（NTG）。我们命名为Optineurin（OPTN）的基因在我们的遗传NTG家族中有16.7%发生突变，定位于10 p14上的GLC 1 E位点，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蛋白研究显示与高尔基体共定位，分泌到房水中，并在许多眼和非眼组织中表达。小鼠和猴基因的克隆显示出与人OPTN非常相似的mRNA和蛋白表达模式。其他研究者也表明OPTN与Ad E3-14.7K、Huntingtin、TFIIIA、RAB 8和其他2种未知激酶相互作用。尽管现有证据表明OPTN通过其与其他蛋白质的相互作用可能利用TNF-α或Fas-配体途径介导凋亡、炎症或血管收缩，但迄今为止，这些还没有明确的迹象表明OPTN突变是如何通过其与其他蛋白质的相互作用来介导凋亡、炎症或血管收缩的。 导致NTG或POAG。因此，作为理解该基因及其蛋白产物的眼部功能的第一步，我们提出以下内容。1)-筛选大量青光眼患者的OPTN突变，以建立基因型/表型相关性; 2）-使用OPTN作为“诱饵”，通过酵母双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在正常和青光眼眼中的分布，并确定其潜在的差异组织表达模式;和5）-使用原位杂交研究小鼠胚胎中Optn的发育表达模式。在这项研究的结论，预计我们的研究将提供必要的科学信息，有一天可能有助于设计创新和未来的药物干预这组视神经病变。