Developmental regulation of lens gene expression

晶状体基因表达的发育调控

基本信息

批准号：
10436285
负责人：
Ales Cvekl
金额：
$ 47.01万
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：
2003
资助国家：
美国
起止时间：
2003-04-01 至 2025-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10436285
关键词：
3&apos Untranslated Regions 3-Dimensional Binding Biological Birth Blindness Cataract Cell Differentiation process Cell Nucleus Cells Chromatin Code Complex Coupled Crystallins Cytoplasm DNA DNA Binding DNA Modification Process Data Degenerative Disorder Development Embryo Event Eye Ferritin Gene Expression Gene Expression Regulation Genes Genetic Genetic Transcription Genetic Translation Genomics Goals Growth Factor Homeostasis Human Individual Knowledge Lens Fiber Lens development Life Cycle Stages Light Link Messenger RNA Modeling Molecular Mus Mutate Mutation Nuclear Nuclear Export Nucleic Acid Regulatory Sequences Outcome Study Pathway interactions Post-Transcriptional Regulation Post-Translational Protein Processing Process Proteins Proteomics RNA RNA Splicing RNA-Binding Proteins Regulation Regulatory Element Research Resolution Site Spatial Distribution Structure System Testing Tissue-Specific Gene Expression Tissues Transgenic Organisms Translations Untranslated RNA Untranslated Regions Visualization base cell type congenital cataract deletion analysis design fiber cell gamma-Crystallins genome wide association study in vivo in vivo Model insight lens lens morphogenesis novel programs promoter regenerative therapy single molecule solute transcription factor transcriptomics

项目摘要

ABSTRACT The long-term goal of this research program is to uncover the transcriptional and posttranscriptional mechanisms coordinating the regulation of tissue-specific gene expression during mammalian embryonic lens development and differentiation. Despite the simple composition of the lens that contains only two mature cell types, the full range and spectrum of mechanisms and pathways governing lens gene expression remain to be identified. A hallmark feature of lens development and cellular differentiation is the temporally and spatially regulated expression of a- and b-/g-crystallins required for lens structure, homeostasis and transparency. This application seeks to identify novel cis-acting DNA regulatory sites and novel RNA-binding proteins (RBPs) that regulate the expression of mammalian crystallins in vivo during lens differentiation. We have now linked the expression of the bB3-crystallin gene in the lens with in vivo binding of transcription factor Pax6 to its “open” promoter chromatin domain in lens enabling prediction of novel cis-acting sites and transcription factors and their posttranslational modifications (PTMs) that spatially and temporally control its expression. We have also discovered transient nuclear accumulation of spliced Crybb3 mRNAs in early lens fiber cell nuclei along with multiple spatially and temporally regulated RBPs that likely regulate bB3- and other crystallin mRNAs transport, stability, translation and decay. These general mechanisms control expression of other lens-specific genes making their discovery an essential first step towards understanding both transcriptional and posttranscriptional regulatory mechanisms underlying lens differentiation. To test our predictions, Aim 1 is designed to uncover the lens-specific cis-regulatory grammar regulating the mouse Crybb3 as a model crystallin. Aim 2 is designed to examine posttranscriptional regulation of lens gene expression by identifying novel RBPs and their sites in crystallin mRNAs particularly within their 3’-UTRs, determining function of identified differentiation-regulated RBPs, and employing a state-of-art MS2-MCP system to in vivo track individual crystallin mRNAs from their birth in the nucleus throughout their nuclear export and localization in the vast space of lens fiber cells at single molecule resolution. These results will reveal those mechanisms linking a-, b/g-crystallin mRNA regulation, identify novel and diverse RBPs that interact with crystallin mRNAs, establish molecular mechanisms of Crybb3 transcription, and provide new insights into the 3D-distribution of individual crystallin mRNAs in lens fiber cells at specific stages of lens cell differentiation. Together, these studies will generate transformative insights into the molecular and cellular mechanisms required for mammalian lens morphogenesis, transparency and refraction and provide a basis for comprehensive understanding of gene expression in more complex tissues in and outside of the eye.

抽象的该研究计划的长期目标是揭示转录和转录后哺乳动物胚胎期间组织特异性基因表达的协调调节机制尽管镜头的构成很简单，只包含两个镜头。成熟细胞类型、全套机制和控制晶状体基因晶状体发育和细胞分化的一个标志性特征仍有待确定。晶状体结构所需的a-和b-/g-晶状体蛋白的时间和空间调节表达，该应用旨在识别新的顺式作用 DNA 调控位点。以及调节哺乳动物体内晶状体蛋白表达的新型 RNA 结合蛋白 (RBP) 我们现在已经将晶状体中 bB3-晶状体蛋白基因的表达与晶状体中的表达联系起来。转录因子 Pax6 与晶状体中“开放”启动子染色质结构域的体内结合新顺式作用位点和转录因子及其翻译后修饰的预测（PTM）在空间和时间上控制其表达，我们还发现了瞬时核。剪接的 Crybb3 mRNA 在早期晶状体纤维细胞核中的积累以及多个空间和时间调节的 RBP 可能调节 bB3- 和其他晶状体蛋白 mRNA 的运输、稳定性、这些一般机制控制其他晶状体特异性基因的表达。他们的发现是朝着转录和转录后迈出的重要的第一步为了检验我们的预测，目标 1 旨在实现晶状体差异化的监管机制。揭示调节小鼠 Crybb3 作为晶体蛋白模型的晶状体特异性顺式调节语法。 2旨在通过鉴定新的RBP来检查晶状体基因表达的转录后调控及其在晶状体蛋白 mRNA 中的位点，特别是在其 3’-UTR 内，决定了已识别的功能分化调节的 RBP，并采用最先进的 MS2-MCP 系统在体内跟踪个体晶状体蛋白 mRNA 从在细胞核中诞生到其核输出和在广大细胞中的定位单分子分辨率下晶状体纤维细胞的空间这些结果将揭示那些联系机制。 a-、b/g-晶状体蛋白 mRNA 调节，识别与晶状体蛋白 mRNA 相互作用的新颖且多样的 RBP，建立 Crybb3 转录的分子机制，并为 3D 分布提供新的见解晶状体细胞分化特定阶段的晶状体纤维细胞中的各个晶状体蛋白 mRNA 一起，这些研究将对分子和细胞机制产生变革性的见解。哺乳动物晶状体的形态发生、透明度和折射，并为综合研究提供基础了解眼睛内外更复杂组织中的基因表达。