Identification of snapin and three novel proteins (BLOS1, BLOS2, and BLOS3/reduced pigmentation) as subunits of biogenesis of lysosome-related organelles complex-1 (BLOC-1)
Identification of snapin and three novel proteins (BLOS1, BLOS2, and BLOS3/reduced pigmentation) as subunits of biogenesis of lysosome-related organelles complex-1 (BLOC-1)
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DOI：
10.1074/jbc.m402513200
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发表时间：
2004-07-02
影响因子：
4.8
通讯作者：
Dell'Angelica, EC
中科院分区：
文献类型：
--
作者：
Starcevic, M;Dell'Angelica, EC
溶酶体相关细胞器生物合成复合物-1（BLOC-1）是一种广泛表达的多亚基蛋白复合物，是内体-溶酶体系统中特殊细胞器（如黑素体和血小板致密颗粒）正常生物合成所必需的。已知该复合物含有卷曲螺旋形成蛋白、Pallidin、Muted、Cappuccino和Dysbindin。编码这些蛋白质的基因在作为赫尔曼斯基-普德拉克综合征（HPS）模型的近交小鼠品系中存在缺陷，HPS是一种以色素减退和血小板储存库缺陷为特征的遗传性疾病。此外，人类Dysbindin的突变导致HPS 7型。在这里，我们报告的复合物的另外四个亚基的鉴定。一个是Snapin，一种卷曲螺旋形成蛋白，以前被表征为突触体相关蛋白25和23的结合伴侣，并参与膜融合事件的调节。其他三个是以前未表征的蛋白质，我们将其命名为BLOC亚基1，2和3（BLOS 1，-2和-3）。使用特异性抗体检测来自人类和小鼠细胞的内源性蛋白质，我们发现Snapin，BLOS 1，BLOS 2和BLOS 3与先前已知的BLOC-1亚基在尺寸排阻色谱法上共免疫沉淀和共破碎。此外，这四种蛋白质的稳态水平在来自苍白小鼠的细胞中显着降低，这些小鼠携带Pallidin突变并显示其他BLOC-1亚基的继发性丢失。酵母双杂交分析表明，二元相互作用的网络，涉及所有以前已知的和新发现的亚基。有趣的是，HPS小鼠模型品系，减少色素沉着，在编码BLOS 3的基因中携带无义突变。从尺寸排阻色谱分析判断，减少色素沉着突变影响BLOC-1的组装严重程度低于苍白突变。编码Snapin和BLOS蛋白的人类基因突变可能是HPS新形式的基础。
Biogenesis of lysosome-related organelles complex-1 (BLOC-1) is a ubiquitously expressed multisubunit protein complex required for the normal biogenesis of specialized organelles of the endosomal-lysosomal system, such as melanosomes and platelet dense granules. The complex is known to contain the coiled-coil-forming proteins, Pallidin, Muted, Cappuccino, and Dysbindin. The genes encoding these proteins are defective in inbred mouse strains that serve as models of Hermansky-Pudlak syndrome (HPS), a genetic disorder characterized by hypopigmentation and platelet storage pool deficiency. In addition, mutation of human Dysbindin causes HPS type 7. Here, we report the identification of another four subunits of the complex. One is Snapin, a coiled-coil-forming protein previously characterized as a binding partner of synaptosomal-associated proteins 25 and 23 and implicated in the regulation of membrane fusion events. The other three are previously uncharacterized proteins, which we named BLOC subunits 1, 2, and 3 (BLOS1, -2, and -3). Using specific antibodies to detect endogenous proteins from human and mouse cells, we found that Snapin, BLOS1, BLOS2, and BLOS3 co-immunoprecipitate, and co-fractionate upon size exclusion chromatography, with previously known BLOC-1 subunits. Furthermore, steady-state levels of the four proteins are significantly reduced in cells from pallid mice, which carry a mutation in Pallidin and display secondary loss of other BLOC-1 subunits. Yeast two-hybrid analyses suggest a network of binary interactions involving all of the previously known and newly identified subunits. Interestingly, the HPS mouse model strain, reduced pigmentation, carries a nonsense mutation in the gene encoding BLOS3. As judged from size exclusion chromatographic analyses, the reduced pigmentation mutation affects BLOC-1 assembly less severely than the pallid mutation. Mutations in the human genes encoding Snapin and the BLOS proteins could underlie novel forms of HPS.