Unconventional Myosin - Organelle Interactions

小鼠“稀释”非常规肌球蛋白（Myo5a）位点为研究非常规肌球蛋白家族密切相关成员的分子运动-货物相互作用和重叠功能提供了一个杰出的遗传系统。“稀释”系统的几个方面有助于强有力地回答这些问题：“稀释”位点和“灰白”、“铅灰色”和“稀释抑制”位点上的多个突变等位基因的可用性；Mercer最近对“稀释”家族的第二个哺乳动物成员myr 6 （Myo5b）的描述；操纵老鼠基因组的能力；原代黑素细胞培养系统的可用性为“稀释”肌球蛋白功能提供了视觉检测。这些研究将通过提供对机动-货物相互作用的见解和测试非常规肌球蛋白家族的功能冗余来影响细胞器运输领域。这些研究有以下目标：1、利用细胞生物学、生物化学和遗传学方法，确定“稀释”的功能，并鉴定与“稀释”在原发性黑素细胞和黑素瘤细胞系中的相互作用的蛋白质。2、在培养细胞和转基因小鼠中，利用与“稀释”肌球蛋白（myosin- va）及其近亲myr6 （myosin- vb）的部分嵌合cDNA构建物，鉴定“稀释”肌球蛋白尾部赋予货物结合特异性的区域。

The murine "dilute" unconventional myosin (Myo5a) locus provides an outstanding genetic system to study molecular motor-cargo interactions and overlapping functions of closely related members of unconventional myosin families. Several aspects of the "dilute" system contribute to a powerful approach to answering these questions: the availability of multiple mutant alleles at the "dilute" locus and the "ashen," "leaden," and "dilute suppressor" loci; the recent characterization by Mercer of a second mammalian member of the "dilute" family, myr 6 (Myo5b); the ability to manipulate the genome of the mouse; and the availability of a primary melanocyte cell culture system that provides a visual assay for "dilute" myosin function. The studies will impact the organelle transport field by providing insights into motor-cargo interactions and testing for functional redundancy within an unconventional myosin family.The studies have the following objectives:1, Determine the functions of "dilute" and identify proteins that interact with "dilute" in both primary melanocyte and melanoma cell lines, using cell biological, biochemical, and genetic methods.2, Identify regions of the "dilute" myosin tail that confer cargo-binding specificity using chimeric cDNA constructs with portions of the "dilute" myosin (myosin-Va) and its close relative, myr 6 (myosin-Vb), in both cultured cells and in transgenic mice.