Kozlowski K, Walter M A
Department of Ophthalmology, University of Alberta, Edmonton, Alberta T6G 2H7, Canada.
Hum Mol Genet. 2000 Sep 1;9(14):2131-9. doi: 10.1093/hmg/9.14.2131.
The autosomal dominant disorders iris hypolasia (IH), iridogoniodysgenesis syndrome (IGDS) and Axenfeld-Rieger syndrome (ARS) are characterized by maldevelopment of the anterior segment of the eye associated with an increased risk of early-onset glaucoma. IH, IGDS and ARS are allelic disorders, as all three can result from mutations of the transcription factor PITX2. IH is the mildest of the three, whereas ARS exhibits the most severe ocular malformations. We hypothesize that varying amounts of residual PITX2 activity could underlie the severity of these phenotypes. Missense mutations of the PITX2 homeodomain identified in IH (Arg46Trp), IGDS (Arg31His) and ARS patients (Leu16Gln; Thr30Pro; Arg53Pro) were introduced into recombinant PITX2 cDNA by site-directed mutagenesis. PITX2 mutant proteins expressed in COS-7 cells were determined to be stable and localized to the nucleus; however, the Arg53Pro ARS mutant also displayed cytoplasmic staining. Our findings are consistent with the possibility of a novel nuclear localization signal (NLS) within helix 3 of the PITX2 homeodomain, homologous to the NLS of the related transcription factor PDX-1. Analysis of the five mutant PITX2 proteins by DNA-binding shifts and transactivation studies demonstrated reduced activity of the IH and IGDS mutant PITX2 proteins, with the IH mutant retaining the most activity in both studies, whereas the ARS mutant PITX2 proteins proved to be non-functional. In addition to providing insight into the etiological mechanism of IH, IGDS and ARS, these results are consistent with the hypothesis that mutant PITX2 proteins that retain partial function result in milder anterior segment aberrations.
常染色体显性疾病虹膜发育不全（IH）、虹膜角膜内皮发育不良综合征（IGDS）和Axenfeld-Rieger综合征（ARS）的特征是眼前节发育异常，并伴有早发性青光眼风险增加。IH、IGDS和ARS是等位基因疾病，因为这三种疾病都可能由转录因子PITX2的突变引起。IH是这三种疾病中最轻微的，而ARS表现出最严重的眼部畸形。我们推测，不同水平的残余PITX2活性可能是这些表型严重程度的基础。通过定点诱变将在IH（Arg46Trp）、IGDS（Arg31His）和ARS患者（Leu16Gln；Thr30Pro；Arg53Pro）中鉴定出的PITX2同源异型域错义突变引入重组PITX2 cDNA。在COS-7细胞中表达的PITX2突变蛋白被确定为稳定的，并定位于细胞核；然而，Arg53Pro ARS突变体也显示出细胞质染色。我们的发现与PITX2同源异型域螺旋3内存在一个新的核定位信号（NLS）的可能性一致，该信号与相关转录因子PDX-1的NLS同源。通过DNA结合迁移和反式激活研究对五种突变PITX2蛋白进行分析，结果表明IH和IGDS突变PITX2蛋白的活性降低，在两项研究中IH突变体保留的活性最高，而ARS突变PITX2蛋白被证明无功能。这些结果除了有助于深入了解IH、IGDS和ARS的病因机制外，还与以下假设一致：保留部分功能的突变PITX2蛋白会导致较轻的眼前节畸变。
