Phosphatidylinositol-4-kinase type II α is a component of adaptor protein-3-derived vesicles
Phosphatidylinositol-4-kinase type II α is a component of adaptor protein-3-derived vesicles
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DOI：
10.1091/mbc.e05-01-0020
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发表时间：
2005-08-01
影响因子：
3.3
通讯作者：
Faundez, V
中科院分区：
文献类型：
--
作者：
Salazar, G;Craige, B;Faundez, V
富含衔接蛋白（AP）-3货物锌转运蛋白3的囊泡中的膜组分从PC 12细胞中产生，并用于通过质谱法鉴定这些细胞器的新组分。蛋白质中突出表示的馏分包括AP-3亚基，突触囊泡蛋白，和溶酶体蛋白已知被排序在AP-3依赖的方式或与AP-3的遗传相互作用。在该级分中富集的蛋白质是磷脂酰肌醇-4-激酶II型α（P14 KII α）。生物化学，药理学和形态学分析支持AP-3阳性细胞器中存在P14 KII α。此外，在AP-3缺陷摩卡突变小鼠的细胞中，P14 KII α的亚细胞定位发生了改变。通常存在于核周和外周细胞器中的P14 KII α在AP-3缺陷的摩卡成纤维细胞的外周膜中显著降低。此外，是在AP-3依赖的方式排序的其他蛋白质的情况下，P14 KII α含量强烈减少摩卡海马苔藓纤维的神经末梢。通过P14 KII α敲低实验进一步探索AP-3和P14 KII α之间的功能关系。P14 KII α的细胞含量的减少强烈降低了在PC 12细胞中观察到的AP-3的点状分布。这些结果表明P14 KIIa存在于AP-3细胞器上，在那里它调节AP-3功能。
A membrane fraction enriched in vesicles containing the adaptor protein (AP) -3 cargo zinc transporter 3 was generated from PC12 cells and was used to identify new components of these organelles by mass spectrometry. Proteins prominently represented in the fraction included AP-3 subunits, synaptic vesicle proteins, and lysosomal proteins known to be sorted in an AP-3-dependent way or to interact genetically with AP-3. A protein enriched in this fraction was phosphatidylinositol-4-kinase type II alpha (P14KII alpha). Biochemical, pharmacological, and morphological analyses supported the presence of P14KII alpha in AP-3-positive organelles. Furthermore, the subcellular localization of P14KII alpha was altered in cells from AP-3-deficient mocha mutant mice. The P14KII alpha normally present both in perinuclear and peripheral organelles was substantially decreased in the peripheral membranes of AP-3-deficient mocha fibroblasts. In addition, as is the case for other proteins sorted in an AP-3-dependent way, P14KII alpha content was strongly reduced in nerve terminals of mocha hippocampal mossy fibers. The functional relationship between AP-3 and P14KII alpha was further explored by P14KII alpha knockdown experiments. Reduction of the cellular content of P14KII alpha strongly decreased the punctate distribution of AP-3 observed in PC12 cells. These results indicate that P14KIIa is present on AP-3 organelles where it regulates AP-3 function.