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改变与人体器官功能衰竭之间的相互关系。这一提议背后的理由是，黑素体和溶酶体之间有明显的相似之处。这项研究描述了黑素体和溶酶体的生物合成之间的相互关系及其在人类中的功能作用。提出这一建议的基本原理是，黑素体和溶酶体在生物合成过程中存在显著的相似性，即这两个颗粒存在于酸性隔室中，并共享许多共同的生物合成过程，如跨高尔基网络(TGN)的囊泡运输。黑素小体由糖蛋白组成，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含量最高。我们以前的研究表明：(1)I期黑素小体起源于内体颗粒，如早期和晚期的e…(2)黑素小体与阳离子非依赖性甘露糖6磷酸受体(CI-M6PR)相关，并受ADP核糖化因子(ARF)调节；(3)Rab7(小GTP结合蛋白)在TYRP1从TGN到早期黑素糖体的运输中起关键作用。在这个项目中，我们希望利用转基因小鼠系统来研究黑素体内酸性内体和溶酶体颗粒生物合成过程中的囊泡运输及其生物学作用。我们发现(1)抑制Rab7功能导致蛋白酶体降解导致TYRP1优先从黑素细胞中消除，(2)内源和外源TYRP1与适配素1(AP1)共免疫共沉淀，(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，以及(4)FLAG标记的wt TYRP1与AP1、MGPR和GGA3共定位。我们还成功地建立了显性/阴性Rab7转基因小鼠，但尚未成功鉴定这些小鼠的视觉色素缺陷。较少

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