Early Functions of Tbx2 and Tbx3 in inner ear development

Tbx2 和 Tbx3 在内耳发育中的早期功能

• 批准号: 10273131
• 负责人: Hansoo Song
• 金额: $ 4.17万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-16 至 2022-07-15
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10273131
• 关键词: Acceleration; Alleles; Anterior; Apoptosis; Auditory; Avidin; Biological Assay; Candidate Disease Gene; Cell Proliferation; Cells; ChIP-seq; Cochlea; Complex; DNA Binding Domain; Data; Defect; Development; Dorsal; Ectopic Expression; Embryo; Failure; Family; Future; Gene Expression Profile; Genes; Genetic; Goals; Hearing; Histologic; Histology; Homeobox; Homologous Gene; In Situ Hybridization; Individual; Knock-out; Knockout Mice; Labyrinth; LoxP-flanked allele; Mammals; Molecular; Morphogenesis; Mus; Mutant Strains Mice; Neurons; Organ; Organogenesis; Otic Vesicle; Outcome; Paint; Pathway interactions; Pattern; Phenotype; Phosphoric Monoester Hydrolases; Population; Positioning Attribute; Process; Quantitative Reverse Transcriptase PCR; Semicircular canal structure; Sensory; Short Interspersed Nucleotide Elements; Signal Transduction; Simple Epithelium; Specific qualifier value; Structure; Structure of thyroid parafollicular cell; Testing; Tissues; Transcription Coactivator; base; bone morphogenetic protein 4; conditional knockout; design; developmental disease; experimental study; inner ear development; interest; member; mutant; novel; precursor cell; spatiotemporal; transcription factor; Acceleration; Alleles; Anterior; Apoptosis; Auditory; Avidin; Biological Assay; Candidate Disease Gene; Cell Proliferation; Cells; ChIP-seq; Cochlea; Complex; DNA Binding Domain; Data; Defect; Development; Dorsal; Ectopic Expression; Embryo; Failure; Family; Future; Gene Expression Profile; Genes; Genetic; Goals; Hearing; Histologic; Histology; Homeobox; Homologous Gene; In Situ Hybridization; Individual; Knock-out; Knockout Mice; Labyrinth; LoxP-flanked allele; Mammals; Molecular; Morphogenesis; Mus; Mutant Strains Mice; Neurons; Organ; Organogenesis; Otic Vesicle; Outcome; Paint; Pathway interactions; Pattern; Phenotype; Phosphoric Monoester Hydrolases; Population; Positioning Attribute; Process; Quantitative Reverse Transcriptase PCR; Semicircular canal structure; Sensory; Short Interspersed Nucleotide Elements; Signal Transduction; Simple Epithelium; Specific qualifier value; Structure; Structure of thyroid parafollicular cell; Testing; Tissues; Transcription Coactivator; base; bone morphogenetic protein 4; conditional knockout; design; developmental disease; experimental study; inner ear development; interest; member; mutant; novel; precursor cell; spatiotemporal; transcription factor

Abstract: The mature inner ear is responsible for the vestibular and auditory senses in mammals. A simple otic vesicle (OV) gives rise to all mature inner ear structures. However, much about the genetic controls regulating cell fate decisions required for morphogenesis of the complex inner ear structures are unknown. There are three T-box transcription factor genes, Tbx1, Tbx2 and Tbx3 expressed in the OV. With the use of Pax2-Cre/+, I inactivated Tbx2 and/or Tbx3 floxed alleles and I uncovered cochlear and saccular defects in Tbx2 conditional knockout (Tbx2cKO) embryos, semicircular canal defects in Tbx3 conditional knockout (Tbx3cKO) embryos and complete failure of the OV to undergo morphogenesis in Tbx2/3cKO double mutant embryos. From these preliminary results, I suggest that Tbx2 and Tbx3 have unique and shared functions in inner ear development. I have generated preliminary RNAscope in situ hybridization data using NeuroD1 (Neurogenic Differentiation 1) that marks neuronal precursors, Bmp4 (Bone Morphogenetic Protein 4) that marks sensory precursor cells and Otx2 (Orthodenticle Homeobox 2) that marks non-sensory precursor cells. We identified an ectopic domain of NeuroD1 expression in the posterior OV, an expansion of the Bmp4 expression in the posterior domain and reduction of Otx2 expression in the ventral domain in Tbx2/3cKO embryos. Our lab previously found that inactivation of Tbx1 results in failed morphogenesis of the OV with expansion of the neurogenic domain to the posterior OV, as in Tbx2/3cKO embryos. Based upon these expression changes, we hypothesize that Tbx2 and Tbx3 have early shared functions in restricting sensory and neuronal cell fates and promoting non-sensory cell fates. In Aim 1, I will carefully determine the spatiotemporal expression patterns of Tbx2 and Tbx3 and will further assess phenotypes of individual and combined Tbx2 and Tbx3 mutant embryos to gain a better understanding of when and where inner ear defects arise with respect to expression of the two genes. In Aim 2, I will take a candidate gene approach to determine the relative position as to where Tbx1, Tbx2 and Tbx3 fit into the genetic hierarchy of cell fate decisions to form the neuronal, sensory or non-sensory fates. I will also distinguish between changes in cell fate and changes in cell populations. Taken together, this project will elucidate the early shared functions of Tbx2 and Tbx3, and compare that with established Tbx1, Eya1 (EYA Transcriptional Coactivator And Phosphatase 1), and Six1 (Sine Oculis Homeobox Homolog 1) pathways, all required for morphogenesis of the inner ear.

抽象的： 成熟的内耳负责哺乳动物的前庭和听觉感觉。一个简单的耳囊 （OV）产生了所有成熟的内耳结构。但是，关于调节细胞命运的遗传控制的很多 复杂的内耳结构形态发生所需的决策尚不清楚。有三个T-box 在OV中表达的转录因子基因TBX1，TBX2和TBX3。使用PAX2-CRE/+的使用，我灭活了 tbx2和/或tbx3 floxed等位基因，我在TBX2中发现了耳蜗和寒冷的缺陷条件敲除 （TBX2CKO）胚胎，TBX3条件敲除（TBX3CKO）胚胎中的半圆形管缺陷并完成 OV失败在TBX2/3CKO双突变胚胎中发生形态发生。从这些初步 结果，我建议TBX2和TBX3在内耳发育中具有独特的和共享的功能。我有 使用NeuroD1（神经源分化1）生成初步的rnascope原位杂交数据 标记神经元前体，BMP4（骨形态发生蛋白4），标记感觉前体细胞和OTX2 （正源性同源2）标记非感官前体细胞。我们确定了一个异位域 NeuroD1在后OV中的表达，后域中BMP4表达的扩展和 在TBX2/3CKO胚胎中的腹侧结构域中的OTX2表达还原。我们的实验室以前发现 TBX1的失活导致OV的形态发生失败，而神经源性结构域则扩大到 后OV，如TBX2/3CKO胚胎。基于这些表达变化，我们假设TBX2和 TBX3在限制感觉和神经元细胞命运并促进非感官细胞方面具有早期共享功能 命运。在AIM 1中，我将仔细确定TBX2和TBX3的时空表达模式，并将进一步 评估个体和联合TBX2和TBX3突变体胚胎的表型，以获得更好的理解 内耳缺陷的何时及其在两个基因的表达中产生。在AIM 2中，我会接受 候选基因方法来确定tbx1，tbx2和tbx3适合遗传的相对位置 细胞命运决定的层次结构，形成神经元，感觉或非感官命运。我还将区分 细胞命运的变化和细胞种群的变化。综上所述，该项目将阐明早期共享 TBX2和TBX3的功能，并将其与已建立的TBX1，EYA1（EYA转录共激活因子 和磷酸酶1）和六1（正弦Oculis同型同源物1）途径，所有这些都需要形态发生 内耳。