An unconventional dileucine-based motif and a novel cytosolic motif are required for the lysosomal and melanosomal targeting of OA1
An unconventional dileucine-based motif and a novel cytosolic motif are required for the lysosomal and melanosomal targeting of OA1
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DOI：
10.1242/jcs.02930
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发表时间：
2006-05-15
影响因子：
4
通讯作者：
Schiaffino, Maria Vittoria
中科院分区：
文献类型：
--
作者：
Piccirillo, Rosanna;Palmisano, Ilaria;Schiaffino, Maria Vittoria
1型眼白化病基因的蛋白产物OA1是一种色素细胞特异性膜糖蛋白，具有g蛋白偶联受体的特征，但仅局限于晚期核内体、溶酶体和黑素小体。为了剖析负责OA1细胞内定位的信号，我们生成了由OA1的细胞质结构域融合到LAMP1的管腔和跨膜结构域组成的嵌合蛋白；此外，我们还生成了全长OA1的错义和缺失突变体。使用这种方法，我们在黑素细胞和非黑素细胞中确定了两种独立的分选信号，这两种信号对于细胞内保留以及溶酶体和黑素体定位都是必要和充分的。这些分选信号是位于第三细胞质环内的非常规二亮氨酸基序和位于c端尾部的以色氨酸-谷氨酸双偶体为特征的新型基序。这两个基序必须发生突变才能促进OA1的质膜定位，这表明它们可以独立驱动OA1的细胞内靶向。此外，这两种基序在非黑素细胞中作为溶酶体分选信号的作用相似，但在黑素细胞中似乎具有不同的特异性。我们的研究结果表明，OA1含有多种非常规信号，负责其溶酶体和黑素体的定位，并揭示了多体蛋白分选到专门分泌细胞器的调节的显着和意想不到的复杂性。
The protein product of the gene responsible for ocular albinism type 1, named OA1, is a pigment-cell-specific membrane glycoprotein, displaying features of G-protein-coupled receptors, yet exclusively localized to late endosomes, lysosomes and melanosomes. To dissect the signals responsible for the intracellular localization of OA1, we generated chimeric proteins consisting of the cytosolic domains of OA1 fused to the lumenal and transmembrane domains of LAMP1; in addition, we generated missense and deletion mutants of full-length OA1. Using this approach, we identified two separate sorting signals that are both necessary and sufficient for intracellular retention, as well as lysosomal and melanosomal localization, in melanocytic and non-melanocytic cells. These sorting signals are an unconventional dileucine motif within the third cytosolic loop and a novel motif, characterized by a tryptophan-glutamic acid doublet, within the C-terminal tail. Both motifs must be mutated to promote the plasma membrane localization of OA1, suggesting that they can independently drive its intracellular targeting. In addition, both motifs act similarly as lysosomal sorting signals in non-melanocytic cells, but appear to carry different specificities in melanocytic cells. Our findings indicate that OA1 contains multiple unconventional signals responsible for its lysosomal and melanosomal localization, and reveal a remarkable and unforeseen complexity in the regulation of polytopic protein sorting to specialized secretory organelles.