MYOSIN I BETA AND INNER EAR STRUCTURE/FUNCTION

描述：毛细胞使用一种适应机制来确保其 转导通道在其最佳敏感点运行。 蛋白质分子同一性的测定 适应之所以重要，有几个原因：第一，这种认同 应该有助于我们尝试发现转导的其他成员 仪器。其次，因为毛细胞是如此稀缺的材料来源 对负责基因的研究和鉴定将允许我们表达 其丰富的蛋白质可用于生化和生物物理研究。最后， 人类耳聋是否应该源于编码适应的突变基因 马达或相关蛋白，未来的基因替代疗法采用 可以想象，功能基因可以纠正这种缺陷。最好的候选人 毛细胞的适应马达是肌球蛋白IB，一种非传统的肌球蛋白 在立体纤毛端发现的同工酶。因为毛细胞是如此的 稀有，因为发束移位直接打开转导 渠道，证明肌球蛋白IB是适应马达使用生化 方法即使不是不可能获得，也是极其困难的。为了测试 无论肌球蛋白IB是否真的是适应马达，我们将取而代之的是消除 从小鼠的基因组中提取它的基因，并研究其对听觉和 随之产生的前庭功能。因为肌球蛋白IB的完全敲除 基因可能有广泛的影响，可能会扰乱我们评估其 在特定的适应作用下，我们也会在转基因小鼠中表达突变 肌球蛋白基因应该对适应有可预测的影响。这 该提案利用了三个实验室的优势：美世 实验室，在老鼠遗传学和分子生物学方面有丰富的经验 克隆，以及吉莱斯皮和科里实验室，在 两种动物适应的生化分析和生理评价 两栖动物和哺乳动物的毛细胞。这里提出的实验将 直接测试肌球蛋白IB是否真的是毛细胞的适应马达。

DESCRIPTION: Hair cells use an adaptation mechanism to ensure that their transduction channels operate at their point of optimal sensitivity. Determination of the molecular identity of proteins responsible for adaptation is important for several reasons: First, such identification should assist us in attempts to discover other members of the transduction apparatus. Second, because hair cells are such a scarce source of material to study, identification of the responsible gene will permit us to express its protein in abundance for biochemical and biophysical studies. Finally, should human deafnesses derive from mutated genes encoding the adaptation motor or associated proteins, future gene-replacement therapies employing a functional gene could conceivably correct such deficits. The best candidate for the hair cell's adaptation motor is myosin IB, an unconventional myosin isozyme that is found at stereociliary tips. Because hair cells are so scarce and because hair-bundle displacements directly open transduction channels, proof that myosin IB is the adaptation motor using biochemical methods is extremely difficult, if not impossible, to obtain. To test whether myosin IB is indeed the adaptation motor, we will instead eliminate its gene from the mouse's genome, and study the consequences on auditory and vestibular function that ensue. Because a complete knockout of myosin IB's gene may have broad effects that may confound our ability to assess its specific role in adaptation, we will also express in transgenic mice mutant myosin genes that should have predictable effects on adaptation. This proposal draws on the strengths of three laboratories: the Mercer laboratory, with extensive experience in mouse genetics and molecular cloning, and the Gillespie and Corey laboratories, with strengths in biochemical analysis and physiological assessment of adaptation in both amphibian and mammalian hair cells. The experiments proposed here will directly test whether myosin IB is indeed the hair cell's adaptation motor.