Investigating the Roles of Epithelial Splice Regulatory Proteins in Inner Ear Development

研究上皮剪接调节蛋白在内耳发育中的作用

• 批准号: 9293902
• 负责人: ALEX ROHACEK
• 金额: $ 2.69万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2018-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9293902
• 关键词: Address; Adult; Affect; Alternative Splicing; Apical; Auditory; Auditory Brainstem Responses; Cadherins; Cell Cycle; Cell Differentiation process; Cell Maturation; Cells; Cleft Palate; Cochlea; Cochlear duct; Congenital Abnormality; Coupled; Defect; Development; Developmental Delay Disorders; Duct (organ) structure; Embryo; Enterobacteria phage P1 Cre recombinase; Epithelial; Epithelium; Event; Exhibits; Exons; Family; Fibroblast Growth Factor Receptors; General Population; Genes; Goals; Hair Cells; Head; Hearing; Hearing Impaired Persons; Human; Incidence; Individual; Infant; Kinetics; Knock-out; Knockout Mice; Labyrinth; Length; Life; Link; Maintenance; Mammals; Measures; Messenger RNA; Morphology; Mus; Mutant Strains Mice; Mutation; Myosin ATPase; Nature; Neonatal; Organ; Organ of Corti; Otic Vesicle; Paint; Pathogenicity; Pediatric Hospitals; Phase; Phenotype; Philadelphia; Process; Protein Isoforms; Proteins; RNA; RNA Splicing; RNA-Binding Proteins; Role; Sensorineural Hearing Loss; Sensory; Sensory Hair; Series; Space Perception; Specific qualifier value; Specificity; Structure; Suggestion; Supporting Cell; Supporting Cell of Organ of Corti; Temporal bone structure; Testing; Time; Tissues; Transcript; Transducers; Tube; United States; Vestibule; cell fate specification; cell type; cranium; deafness; ear development; epithelial to mesenchymal transition; exome sequencing; experimental study; gene function; genetic regulatory protein; hearing impairment; improved; inner ear development; interest; loss of function mutation; mutant; mutant mouse model; next generation sequencing; novel; organ growth; postnatal; progenitor; public health relevance; sound; spatiotemporal; transcriptome sequencing

 DESCRIPTION (provided by applicant): The mammalian inner ear is a dual function organ comprised of the vestibular apparatus that senses spatial orientation and the cochlear duct which functions in sound transduction. The development of the cochlea is of particular interest due to the high incidence of hearing loss that occurs in the general population. The cochlear duct arises from a ventral outgrowth of the inner ear anlage, the otic vesicle. During its development the cochlea extends into a coiled tube that will contain the sensory hair and support cells of the Organ of Corti that are the transducers of sound. The function of the Organ of Corti is tightly coupled to the proper timing of events that occur during its formation. Sensory cell precursors must be specified from the otic epithelium, undergo cell-cycle exit and then differentiate into hair and support cells. The precise timing of these events is crucial for the proper development of the Organ of Corti, the cochlea as a whole, and, subsequently, the sense of hearing. While many of the genes responsible for each of these developmental events have been discovered the factors controlling their timing have yet to be identified. Recently, our collaborator, Dr. Ian Krantz (Children's Hospital of Philadelphia) identified mutations in the Epithelial Splicing Regulatory Protein 1 gene in several individuals with profound sensorineural deafness. In an effort to functionally link the deafness phenotype with mutations in Esrp1 we have begun to analyze mouse mutants in Esrp1 and/or Esrp2 for inner ear phenotypes. My preliminary results indicate that Esrp1 mutants exhibit a truncated cochlear duct and a significant delay in the development of auditory hair and support cells. The goal of this proposal is to determine the mechanism by which Esrp1 regulates the timing of sensory development in the inner ear. To accomplish this objective, I will systematically analyze the inner ears of Esrp mutants for alterations in the specification and cell-cycle exit of sensory progenitors, as well as the differentiation and maturation of cochlear hair and support cells (Aim1a). I will also perform RNAseq experiments on cochlear ducts from Esrp1-/- and control embryos to identify aberrantly spliced mRNA transcripts and potential targets of Esrp1 (Aim 1b). Finally, given the neonatal lethality of Esrp1-/- mutants from a cleft palate defect, I will generate a conditional knockout of Esrp1 in the inner ear to interrogate its requirement for hearing in adult mice (Aim 2). These experiments will uncover unique roles of Esrp genes in inner ear development and highlight the mechanism by which pathogenic mutations in Esrp1 result in deafness.

 描述（由申请人提供）：哺乳动物内耳是一种双功能器官，包括感知空间方位的前庭器和在声音传导中起作用的耳蜗管。由于听力损失在普通人群中的发生率很高，因此耳蜗的发育特别令人感兴趣。耳蜗管起源于内耳原基的腹侧生长，即耳泡。在其发展过程中，耳蜗延伸成一个螺旋管，其中包含感觉毛和支持细胞的器官的科尔蒂是换能器的声音。Corti器官的功能与其形成过程中发生的事件的适当时间紧密相关。感官 细胞前体必须从耳上皮细胞中分化出来，经历细胞周期退出，然后分化成毛发和支持细胞。这些事件的精确时间对于Corti器官，整个耳蜗以及随后的听觉的正常发育至关重要。虽然已经发现了许多与这些发育事件有关的基因，但控制其时间的因素尚未确定。最近，我们的合作者Ian Krantz博士（费城儿童医院）在几个患有深度感音神经性耳聋的个体中发现了上皮剪接调节蛋白1基因的突变。为了将耳聋表型与Esrp 1突变在功能上联系起来，我们已经开始分析小鼠Esrp 1和/或Esrp 2的内耳表型突变体。我的初步结果表明，Esrp 1突变体表现出截短的耳蜗导管和显着延迟的发展听觉毛和支持细胞。该提案的目标是确定Esrp 1调节内耳感觉发育时间的机制。为了实现这一目标，我将系统地分析Esrp突变体内耳的感觉祖细胞的特化和细胞周期退出的改变，以及 耳蜗毛和支持细胞（Aim 1a）的分化和成熟。我还将对来自Esrp 1-/-和对照胚胎的耳蜗导管进行RNAseq实验，以鉴定异常剪接的mRNA转录本和Esrp 1的潜在靶点（Aim 1b）。最后，考虑到腭裂缺陷的Esrp 1-/-突变体的新生儿致死性，我将产生一个条件性敲除， esrp 1在内耳询问其对成年小鼠听力的需求（目的2）。这些实验将揭示Esrp基因在内耳发育中的独特作用，并强调Esrp 1致病性突变导致耳聋的机制。