Myosin 15 Genetics, Pathology and Therapeutic Potential

遗传性内耳疾病很普遍，并对生活质量有重大影响。遗传性内耳疾病目前还没有可用的临床治疗方法。小鼠是了解遗传性内耳疾病和开发治疗措施的理想哺乳动物模型。小鼠模型促进了人类遗传性疾病基础基因的发现，使研究这些基因在内耳发育和功能中的作用成为可能，并有望成为开发遗传性内耳疾病治疗方法的模型。这项拨款申请建立在我们的发现基础上，即非传统肌球蛋白基因Myo15的突变导致了患有DFNB3的人类和两个具有深度隐性耳聋的自发小鼠突变体Shaker 2和Shaker 2J的严重先天性耳聋。我们建议使用这两种DFNB3小鼠模型来建立这种大肌球蛋白的结构和功能特性，在子宫和出生后早期生活中的表达要求，以及受影响个体的病理发育基础。随着这些目标的实现，我们将更接近儿童先天性耳聋的基因治疗的最终目标，并扩大我们对基本听力过程的分子理解。在第一个目标中，我们将严格测试通过BAC转基因互补而实现的Shaker 2小鼠耳聋表型矫正的长期功能和结构结果。第二个目的是检验一种假说，即肌球蛋白XV缺失的毛细胞在发育过程中不能从基底膜上分离出来，从而导致细胞病理。第三个目的是研究Myo15功能的丧失是否可以被其他肌球蛋白的功能部分补偿。第四个目的是测试不寻常的MYOXV蛋白的N端三分之一是否对转基因小鼠的功能重要。最后，将利用Myo15转基因在出生后的Shaker 2小鼠中的诱导性表达来评估在新生儿或幼儿中进行DFNB3基因治疗的可行性。我们的调查团队在跨学科合作方面取得了成就，这带来了耳鼻喉科和形态学方面的专业知识以及分子和发育遗传学方面的专业知识。我们的跨学科方法对于充分利用动物模型至关重要，这些动物模型将成为开发和测试遗传性内耳疾病治疗方法的基础。

Hereditary inner ear disease is prevalent and has significant implications for quality of life. There is currently no available clinical cure for hereditary inner ear disease. The mouse serves as an ideal mammalian model for understanding genetic inner ear disease and for developing therapeutic measures. Mouse models have facilitated the discovery of genes that underlie hereditary disease in humans, have made it possible to study the role of these genes in inner ear development and function, and hold great promise as models for developing treatments for hereditary inner ear disease. This grant application builds on our discovery that mutations in the unconventional myosin gene, Myo15, are responsible for profound congenital deafness in humans with DFNB3 and in two spontaneous mouse mutants with profound recessive deafness, shaker 2 and shaker 2J. We propose to use these two mouse models of DFNB3 to establish the structure and functional properties of this large myosin, the requirement for expression in utero and early postnatal life, and the developmental basis for pathology in affected individuals. As each of these goals are accomplished we will move closer toward the ultimate objective of gene therapy for congenital deafness in children and expand our molecular understanding of the basic hearing process. In the first aim, we will rigorously test the long-term functional and structural outcome of the phenotypic correction of deafness in shaker 2 mice that we accomplished by complementation with a BAC transgene. The second aim tests the hypothesis that MYOSIN XV-deficient hair cells fail to detach from the basement membrane during development, leading to cellular pathology. The third aim examines whether loss of Myo15 function is partially compensated by the function of other myosins. The fourth aim tests whether the unusual N-terminal third of the MYOXV protein is important for function using transgenic mice. Finally the feasibility of gene therapy for DFNB3 in newborns or young children will be assessed in the last aim using inducible expression of Myo15 transgenes in young, postnatal shaker 2 mice. Our investigative team has a track record for accomplishments resulting from cross disciplinary collaboration, which has brought expertise in otolaryngology and morphology together with expertise in molecular and developmental genetics. Our interdisciplinary approach is essential to fully exploit the animal models that will be the basis for developing and testing therapy for hereditary inner ear disease.