Pax6 as a key regulator of lens development

Pax6 作为晶状体发育的关键调节因子

基本信息

批准号：
8630418
负责人：
Ales Cvekl
金额：
$ 54.46万
依托单位：
ALBERT EINSTEIN COLLEGE OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-01-01 至 2017-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8630418
关键词：
Apert-Crouzon syndrome Autistic Disorder Binding Sites Biological Assay Blindness CDKN1C gene Cataract Cell Culture Techniques Cell Cycle Cell Cycle Regulation Cell Differentiation process Cells Chromatin Cognition Disorders Craniofacial Abnormalities Crystallins Cyclin-Dependent Kinase Inhibitor DNA Binding DNA Sequence Data Deoxyribonucleases Development Distal Dysplasia ETV1 gene Embryo Enhancers Epilepsy Event Eye Abnormalities FGFR2 gene Failure Fibroblast Growth Factor Fibroblasts Fluorescence Foundations Gene Expression Gene Targeting Genes Genetic Genetic Transcription Goals Growth Human Hypoparathyroidism In Vitro Individual Investigation JUN gene Kidney Lens Fiber Light Link Malignant - descriptor Malignant Neoplasms Mediating Mental Retardation Molecular Molecular Profiling Mus Mutagenesis Mutation Non-Insulin-Dependent Diabetes Mellitus Nuclear Pathway interactions Process Proteins RNA Regulation Regulator Genes Reporter Reporter Genes Reporting Retinoblastoma Protein Role Signal Pathway Signal Transduction Site Site-Directed Mutagenesis Stem cells Syndrome Time Transgenic Mice Transgenic Organisms Up-Regulation Validation Vesicle Work base bone chromatin immunoprecipitation deafness enhanced green fluorescent protein fiber cell inhibitor/antagonist lens lens induction lens morphogenesis lens transparency mutant nervous system disorder novel precursor cell programs public health relevance relating to nervous system research study transcription factor

项目摘要

The long-term goal of this program is to elucidate the molecular mechanisms of mammalian lens development through studies of DNA-binding transcription factor Pax6. Previous studies have shown that Pax6 is essential for establishing lens progenitor cells and regulation of crystallin gene expression. However, additional roles of Pax6 in lens morphogenesis remain to be determined. Genetic studies have shown that Pax6 regulates cell cycle exit of lens precursor cells. The external regulation of lens fiber cell differentiation is mediated by BMP and FGF signaling, through transcription factors Gata3 and Prox1. Using chromatin immunoprecipitation in combination with DNA sequencing (ChIP-seq), and RNA expression profiling in Pax6 mutant lenses, we have now identified a group of of genes directly regulated by Pax6 including Prox1, FGFR2 and Etv1/ER81. Expression of Prox1 is upregulated in the posterior part of the lens vesicle and Prox1 regulates expression of Cdkn1b/p27 and Cdkn1c/p57, two proteins required cell cycle exit of lens precursor cells. FGFR2 and Etv1/ER81 are components of FGF signaling. Gata3 expression is restricted to the posterior part of lens vesicle, and is upstream of Cdkn1b/p27 and Cdkn1c/p57. These findings suggest that the Pax6-dependent cell cycle exit includes FGFR2, Etv1/ER81, Prox1. BMP signaling regulates expression of Gata3 in Pax6-independent manner. Gata3 and Prox1 jointly regulate expression of Cdkn1b/p27 and Cdkn1c/p57. In order to carry out this long-term goal, the following specific aims are proposed: (1) To define Pax6-dependent gene regulatory networks governing expression of Prox1, FGFR2, and Etv1/ER81, and to elucidate FGF-dependent up-regulation of Prox1 in the embryonic lens. (2) To establish molecular basis of Gata3 expression in lens cells via BMP and FGF signaling. (3) To demonstrate that expression of Cdkn1b/p27 and Cdkn1c/p57 is regulated in lens by a combination of Gata3 and Prox1 at the level of transcription. These Aims will be achived through the identification and characterization of distal enhancres in Prox1, FGFR2, Etv1/Er81 and Gata3 genes using transgenic gene reporter and cell culture studies, identification of binding sites of these factors in lens chromatin and in vitro, and identification BMP- and FGF-dependent enhancers in Gata3, and FGF- responsive enhancres in Prox1 gene, respectively.

该程序的长期目标是阐明哺乳动物镜头的分子机制通过研究DNA结合转录因子PAX6的开发。先前的研究表明 PAX6对于建立晶状体祖细胞和结晶蛋白基因表达的调节至关重要。但是，PAX6在晶状体形态发生中的其他作用仍有待确定。遗传研究已有表明PAX6调节透镜前体细胞的细胞周期出口。透镜纤维电池的外部调节分化是由BMP和FGF信号通过转录因子GATA3和Prox1介导的。使用染色质免疫沉淀与DNA测序（CHIP-SEQ）和RNA结合使用 PAX6突变透镜中的表达分析，我们现在已经确定了一组直接调节的基因由PAX6包括Prox1，FGFR2和ETV1/ER81。 Prox1的表达在后部上调透镜囊泡和Prox1的调节CDKN1B/P27和CDKN1C/p57的表达，需要两种蛋白透镜前体细胞的细胞周期出口。 FGFR2和ETV1/ER81是FGF信号的组成部分。 GATA3 表达仅限于晶状体囊泡的后部，并且是CDKN1B/P27的上游 CDKN1C/p57。这些发现表明，PAX6依赖性细胞周期出口包括FGFR2，ETV1/ER81， Prox1。 BMP信号传导以非依赖性的方式调节GATA3的表达。 GATA3和PROX1 共同调节CDKN1B/P27和CDKN1C/P57的表达。为了实现这个长期目标，提出了以下特定目标：（1）定义依赖PAX6的基因调节网络 Prox1，FGFR2和ETV1/ER81的表达，并阐明Prox1的FGF依赖性上调胚胎镜头。（2）通过BMP和FGF建立晶状体细胞中GATA3表达的分子基础信号。（3）证明CDKN1B/P27和CDKN1C/p57的表达在A中受到A的调节 gata3和prox1在转录水平上的组合。这些目标将通过使用Prox1，FGFR2，ETV1/ER81和GATA3基因的远端增强剂的识别和表征转基因基因报告基因和细胞培养研究，识别这些因子的结合位点染色质和体外，以及GATA3中的BMP-和FGF依赖性增强子，以及FGF- Prox1基因中的响应性增强。