Pink1 Kinase and Its Membrane Potential (Δψ)-dependent Cleavage Product Both Localize to Outer Mitochondrial Membrane by Unique Targeting Mode
Pink1 Kinase and Its Membrane Potential (Δψ)-dependent Cleavage Product Both Localize to Outer Mitochondrial Membrane by Unique Targeting Mode
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DOI：
10.1074/jbc.m112.365700
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发表时间：
2012-06-29
影响因子：
4.8
通讯作者：
Voos, Wolfgang
中科院分区：
文献类型：
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作者：
Becker, Dorothea;Richter, Judith;Voos, Wolfgang
帕金森病相关激酶Pink 1靶向线粒体，在线粒体中，它被认为通过促进功能障碍线粒体的选择性自噬去除来调节线粒体质量控制。然而，Pink 1的靶向模式及其亚线粒体定位仍然没有最终解决。本研究的目的是通过一种高灵敏度的体外分析方法来剖析Pink 1的线粒体输入途径。Pink 1序列的突变分析表明，其N末端作为一个真正的基质定位序列，介导的初始膜电位（Δ psi）依赖性靶向的Pink 1前体的线粒体内膜，但它是pink 1进口或加工。信号序列下游的疏水片段阻碍了Pink 1穿过线粒体内膜的完全易位。此外，蛋白质的C-末端促进Pink 1在外膜的保留。因此，由Pink 1序列表征的多个靶向信号导致全长蛋白质及其主要的Delta psi依赖性切割产物最终定位于线粒体外膜的胞质表面。全长Pink 1和删除结构类似的天然Pink 1加工产品被发现组装成膜电位敏感的高分子量蛋白质复合物在线粒体表面，并显示出类似的细胞保护作用时，在体内表达，表明这两个物种的功能相关。
The Parkinson disease-associated kinase Pink1 is targeted to mitochondria where it is thought to regulate mitochondrial quality control by promoting the selective autophagic removal of dysfunctional mitochondria. Nevertheless, the targeting mode of Pink1 and its submitochondrial localization are still not conclusively resolved. The aim of this study was to dissect the mitochondrial import pathway of Pink1 by use of a highly sensitive in vitro assay. Mutational analysis of the Pink1 sequence revealed that its N terminus acts as a genuine matrix localization sequence that mediates the initial membrane potential (Delta psi)-dependent targeting of the Pink1 precursor to the inner mitochondrial membrane, but it is dispensable for Pink1 import or processing. A hydrophobic segment downstream of the signal sequence impeded complete translocation of Pink1 across the mitochondrial inner membrane. Additionally, the C-terminal end of the protein promoted the retention of Pink1 at the outer membrane. Thus, multiple targeting signals featured by the Pink1 sequence result in the final localization of both the full-length protein and its major Delta psi-dependent cleavage product to the cytosolic face of the outer mitochondrial membrane. Full-length Pink1 and deletion constructs resembling the natural Pink1 processing product were found to assemble into membrane potential-sensitive high molecular weight protein complexes at the mitochondrial surface and displayed similar cytoprotective effects when expressed in vivo, indicating that both species are functionally relevant.