BIOGENESIS OF MELANOSOMES

拟议研究的长期目标是了解 正常和病理性皮肤色素沉着的分子基础 states. 四年期间的总体目标是 要求的是描绘黑素细胞的基本机制， 创造了它的专门的细胞器，黑素体， 了解黑素体蛋白的功能。 着力 针对四个具体目标： 1. 黑素体-溶酶体关系。 第一阶段黑素小体 显示了黑素体和 细胞器的内体/溶酶体谱系。 细胞和生化 技术将被用来解决这个细胞器的性质， 理解“黑素溶酶体”疾病的关键，如Chediak- 东 黑素体是否是一种“特化的”溶酶体， 阶段I黑素体是黑素体的分歧点， 将确定细胞器的溶酶体谱系。 2. 酪氨酸酶相关蛋白-1的作用描述。 TRP-1是 早期黑素体区室的标记物。 在这方面的突变 蛋白质影响黑素体的结构和质量和数量 制造黑色素。 小鼠毛色遗传学、细胞生物学和生物化学 技术将被用来检查突变的后果， TRP-1的亚细胞分布及其在黑化中的作用。 的 肤色的种族差异是由于 在TRP-1的数量，分布和功能将被测试。 3. 酪氨酸酶相关蛋白-2的作用描述。 酶 多巴色素互变异构酶与第二种酪氨酸酶相关的 在slaty位点编码的蛋白质。 DHICA，反应产物 是黑色素的主要成分。 合成 和TRP-2的亚细胞分布和“黑素生成”的性质， “这是一个复杂的问题，它将被审查。 石板变异会 被用来深入了解第一个后酪氨酸酶步骤， 黑素生成 4. 色素沉着的药理学调节。 视黄酸和 双乙酰胺抑制培养黑色素瘤色素沉着 细胞，但只有视黄酸会导致黑素体的紊乱。 将用这些试剂处理黑素细胞，并观察它们对黑素细胞的作用。 蛋白质的表达、亚细胞分布和功能 兴趣分析，以了解每种药物如何影响 色素沉着在生化和细胞水平。

The long term goal of the proposed research is an understanding of the molecular basis of cutaneous pigmentation in normal and pathologic states. The overall goal during the four years for which support is requested is to delineate the basic mechanisms by which a melanocyte creates its specialized organelle, the melanosome, and to better understand the function of melanosomal proteins. Efforts will be directed at four specific aims: 1. Melanosomal-lysosomal relationship. The Stage I melanosome demonstrates characteristics of both the melanosomal and endosomal/lysosomal lineage of organelles. Cellular and biochemical techniques will be used to address the nature of this organelle which is the key to understanding "melanolysosomal" disorders such as the Chediak- Higashi. Whether the melanosome is a "specialized" lysosome or whether the Stage I melanosome is a point of divergence for the melanosomal and lysosomal lineage of organelles will be determined. 2. Delineation of the role of Tyrosinase-related protein-1. TRP-1 is a marker for the early melanosomal compartment. Mutations in this protein affect melanosomal structures and the quality and quantity of melanin made. Mouse coat color genetics, cell biologic and biochemical techniques will be employed to examine the consequences of mutations in TRP-1 on its subcellular distribution and its role in melanization. The hypothesis that racial differences in skin color are due to differences in the amount, distribution and function of TRP-1 will be tested. 3. Delineation of the role of Tyrosinase-related protein-2. The enzyme dopachrome tautomerase is identical to a second tyrosinase-related protein encoded at the slaty locus. DHICA, the product of the reaction catalyzed by this enzyme, is a major component of melanin. The synthesis and subcellular distribution of TRP-2 and the nature of the "melanogenic complex" of which it is a part will be examined. The slaty mutation will be employed to gain insight into the first post-tyrosinase steps in melanogenesis. 4. Pharmacologic regulation of pigmentation. Both retinoic acid and hexamethylene bisacetamide inhibit pigmentation in cultured melanoma cells, but only retinoic acid causes a derangement of melanosomes. Melanocytes will be treated with these agents and their effects on the expression, subcellular distribution and function of the proteins of interest analyzed to understand how each drug works to affect pigmentation at the biochemical and cellular level.