Failure of vesicular transport and development of multi-organ defects through acidic melanosomal and lysosomal granules

本研究描述了黑素小体生物合成的改变与人体器官功能衰竭之间的相互关系。这一建议背后的基本原理是黑素体和溶酶体之间有明显的相似之处。本研究描述了黑素体和溶酶体的生物合成及其在人体中的功能作用之间的相互关系。这一建议背后的基本原理是黑素小体和溶酶体在生物合成过程中存在明显的相似性，即这两种颗粒存在于酸性隔间中，并且具有许多共同的生物合成过程，例如反高尔基网络（TGN）的囊泡运输。黑素体由糖蛋白组成，糖蛋白由酪氨酸酶、酪氨酸酶相关蛋白（TYRPs）和结构蛋白gp100组成。在这些黑素体蛋白中，TYRP1是最丰富的一种。我们的前期研究表明(1)I期黑素小体来源于早期和晚期的内体颗粒，(2)黑素小体室室与阳离子非依赖性甘糖6磷酸受体（CI-M6PR）相关，并受adp核糖基化因子（ARF）的调节，(3)Rab7（小gtp结合蛋白）在TYRP1从TGN转运到早期黑素小体中起关键作用。在本项目中，我们希望利用转基因小鼠系统表征黑素小体酸性内体、溶酶体颗粒生物合成过程中的囊泡运输及其在人体内的生物学作用。我们发现(1)Rab7功能的抑制导致黑色素细胞因蛋白酶体降解而优先消除TYRP1，(2)内源性和外源性TYRP1与适应蛋白1（AP1）共免疫沉淀，(3)TYRP1和AP1之间的相互作用存在于tgn衍生的囊泡运输的早期阶段，(4)flag标记的wt TYRP1与AP1， MGPR和GGA3共定位。我们也成功建立了Rab7显性/阴性转基因小鼠，但尚未成功鉴定这些小鼠的视觉色素沉着缺陷。少

This study characterizes the interrelationship between alteration of melanosome biosynthesis and functional failure of organs in man. The rationale behind this proposal is that there are marked similarities between melanosomes and lysosomes. This study characterizes the interrelationship between the biosynthesis of melanosomes and lysosomes and their functional roles in man. The rationale behind this proposal is that there exists a marked similarity in the biosynthesis processes between melanosomes and lysosomes, i.e., the two granules are present in acidic compartments and share many common biosynthesis processes such as vesicular transport from trans Golgi network (TGN). Melanosomes are made of glycoproteins, which consist of tyrosinase, tyrosinase-related proteins (TYRPs) and structural gp100 protein. Among these melanosomal proteins TYRP1 is the most abundant one. Our previous studies indicated (1) that the stage I melanosomes derive from endosomal granules such as early and late e … More ndosomes, (2) that melanosomal compartments are associated with cation independent-mannose 6 phosphate receptor (CI-M6PR) and regulated by ADP-ribosylation factor (ARF), and (3) that Rab7 (small GTP-binding protein) plays a key role in transport of TYRP1 from TGN to early melanosomes. In this project we wish to characterize the vesicular transport in the biosynthesis processes of acidic endosomal, lysosomal granules of melanosomes and their biological roles in man by utilizing transgenic mouse system.We found (1) that the inhibition of Rab7 function resulted in preferential TYRP1 elimination from melanocytes due to proteasomal degradation, (2) that endogenous and exogenous TYRP1 is co-immunoprecipitated with adaptin 1(AP1), (3) that the interaction between TYRP1 and AP1 is present in early stage of the TGN-derived vesicular transport, and (4) that Flag-tagged wt TYRP1 is co-localized with AP1, MGPR and GGA3. We also succeeded in establishing dominant/negative Rab7 transgenic mice, but are not yet successful in identifying visual pigmentation defects in these mice. Less