A vertebrate model of viral and hereditary microcephaly

病毒和遗传性小头畸形的脊椎动物模型

基本信息

批准号：
10576107
负责人：
Priya Shirish Shah
金额：
$ 15.99万
依托单位：
UNIVERSITY OF CALIFORNIA AT DAVIS
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-09-07 至 2024-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10576107
关键词：
Affect Anatomy Animal Model Biological Assay Biological Models Brain Brain Diseases Brain region CRISPR/Cas technology Cell Polarity Chemical Exposure Chemicals Defect Development Disease Dissection Drosophila genus Embryo Embryonic Development Etiology Eye Foundations Future Gene Mutation Gene Transfer Techniques Genes Goals Growth Head Human Immunofluorescence Microscopy Infection Inherited Intellectual functioning disability Knowledge Lead Length Measurement Measures Microcephaly Modeling Molecular Morphology Mutate Mutation Nonstructural Protein Pathogenicity Pathway interactions Phenotype Proteins Publishing Specificity Structure Study models Syndrome System Testing Transgenic Organisms Variant Vertebrates Viral Virus Virus Diseases Work Zebrafish Zika Virus alcohol exposure behavioral study brain size cDNA Expression cell type experimental study fly genome editing high throughput screening human disease in utero in vivo inhibitor nervous system disorder paralogous gene pathogen screening stem cells tool

项目摘要

PROJECT SUMMARY/ABSTRACT Microcephaly, in which head and brain development are severely inhibited, can result in extreme intellectual disability. Microcephaly has many different causes, including gene mutation, pathogen infection and chemical exposure. While the causes of microcephaly may be broad, there may be shared molecular mechanisms involved. Recently published results show how Zika virus non-structural protein 4A (NS4A) inhibits brain development in fruit flies by inhibiting the same ANKLE2 protein whose function is disrupted by gene mutation in a hereditary form of microcephaly. This suggests that there may be key similarities in the molecular mechanisms contributing to a viral and hereditary form of microcephaly. Studying these molecular similarities in vertebrate models is essential to understanding disease in humans since vertebrates have different brain structure and genes that regulate brain development. However, the tools to study this, namely a strong vertebrate model system, do not exist. The primary goal of the proposed project is to fill these major gaps in knowledge by developing a vertebrate model of viral and hereditary microcephaly acting through the ANKLE2 pathway. For the hereditary model, ankle2 will be mutated using CRISPR/Cas9 genome editing. For the viral model, Zika virus NS4A protein will be expressed using Tol2 transgenesis. Defects in development, including various externally measured morphology metrics, brain size, structure and cellular defects in proliferation and survival will be assayed using anatomical measurement and immunofluorescence microscopy. When completed, this new model system will lay the foundation for a molecular and cellular level understanding of ANKLE2 function during vertebrate brain development and how it is disrupted by gene mutation and viral infection. In the long-term, it will enable high- throughput screening of chemical sensitizers and inhibitors of microcephaly, allow the exploration vertebrate- specific mechanisms of brain development in vivo, and behavioral studies.

项目概要/摘要小头畸形，头部和大脑发育受到严重抑制，可导致智力极度低下残疾。小头畸形有许多不同的原因，包括基因突变、病原体感染和化学物质感染。接触。虽然小头畸形的原因可能很广泛，但可能存在共同的分子机制涉及。最近发表的结果显示寨卡病毒非结构蛋白 4A (NS4A) 如何抑制大脑通过抑制相同的 ANKLE2 蛋白（其功能因基因突变而被破坏）来促进果蝇的发育遗传性小头畸形。这表明分子结构可能存在关键相似之处导致病毒性和遗传性小头畸形的机制。研究这些分子相似性脊椎动物模型对于了解人类疾病至关重要，因为脊椎动物具有不同的大脑调节大脑发育的结构和基因。然而，研究这个的工具，即强大的脊椎动物模型系统，不存在。拟议项目的主要目标是通过开发脊椎动物来填补这些主要知识空白通过 ANKLE2 途径发挥作用的病毒和遗传性小头畸形模型。对于遗传模型来说，脚踝2将通过CRISPR/Cas9基因组编辑进行突变。对于病毒模型，寨卡病毒 NS4A 蛋白将是使用 Tol2 转基因表达。发育缺陷，包括各种外部测量的形态将使用解剖学来分析指标、大脑大小、结构以及增殖和存活的细胞缺陷测量和免疫荧光显微镜。完成后，这个新模型系统将奠定为在脊椎动物大脑中分子和细胞水平理解 ANKLE2 功能奠定基础发育以及它如何被基因突变和病毒感染破坏。从长远来看，它将实现高小头畸形化学敏化剂和抑制剂的通量筛选，允许探索脊椎动物体内大脑发育的具体机制和行为研究。