Massive endocytosis triggered by surface membrane palmitoylation under mitochondrial control in BHK fibroblasts.
Massive endocytosis triggered by surface membrane palmitoylation under mitochondrial control in BHK fibroblasts.
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作者：
Hilgemann DW;Fine M;Linder ME;Jennings BC;Lin MJ
大钙瞬变引起大量内吞作用（MEND）在BHK成纤维细胞的非经典机制。我们目前的证据表明，MEND依赖于线粒体通透性转换孔（PTP）的开放，随后辅酶A（CoA）的释放，酰基CoA的合成，和膜蛋白棕榈酰化。MEND通过抑制线粒体Ca摄取或PTP开口、消耗脂肪酸、阻断酰基CoA合成、代谢CoA或抑制棕榈酰化来阻断。它是由线粒体去极化或促进PTP开放触发的。在线粒体MEND阻断后，MEND通过细胞质酰基CoA或CoA恢复。MEND被质膜酰基转移酶DHHC 5的siRNA敲低阻断。当酰基CoA丰富时，瞬时H2O2氧化应激或PKC激活启动MEND，但H2O2的立即存在可防止MEND。PTP抑制剂NIM811显著增加正常生长细胞中的质膜。因此，MEND途径可能有助于依赖于线粒体应激信号传导的组成性以及病理性质膜周转。http://dx.doi.org/10.7554/eLife.01293.001细胞利用一种称为内吞作用的过程来吸收蛋白质和其他分子。内吞作用有许多形式，但它们通常涉及感兴趣的分子被塞入细胞膜中形成的芽中。然后将这个芽掐掉，将分子留在细胞内的囊泡中。一般来说，内吞作用是由“笼”蛋白如网格蛋白触发的，但其他形式也是可能的。这些“非经典”形式的内吞作用涉及病原体的内化和细胞对创伤的反应等多种过程，有时它们涉及大部分细胞膜被夹断。已经观察到几种不同形式的“大量内吞作用”，但与经典形式的内吞作用相比，它们仍然是谜。现在，Hilgemann等人报道了一种新的大规模内吞作用途径，该途径是由钙离子突然流入婴儿肾脏（BHK）成纤维细胞引发的。在这个过程中，高达70%的细胞膜可以被夹断，特别是脂质和蛋白质以比平均细胞膜更有序的模式排列的膜区域。在大量的内吞作用之后，BHK细胞需要大约30分钟来替换在内吞作用期间丢失的区域。Hilgemann等人发现钙离子通过线粒体发挥其影响力，线粒体是大多数细胞的主要能量来源。与所有其他细胞器不同，线粒体被两层同心膜包围。钙离子的流入导致线粒体内膜中的孔（称为渗透性转换孔）打开，使得脂肪酸代谢所需的小分子辅酶A释放到细胞的细胞质中。随后是辅酶A与脂肪酸的缩合以及脂肪酸与表面膜蛋白的附着。这些脂肪酸的附着（一个被称为棕榈酰化的过程）明显地促进了细胞表面有序区域的聚结，并作为膜囊泡被夹入细胞质。一个关键的未回答的问题是，除了胞吞作用之外，线粒体释放辅酶A是否还调节生化过程。DOI：http://dx.doi.org/10.7554/eLife.01293.002网站
Large Ca transients cause massive endocytosis (MEND) in BHK fibroblasts by nonclassical mechanisms. We present evidence that MEND depends on mitochondrial permeability transition pore (PTP) openings, followed by coenzyme A (CoA) release, acyl CoA synthesis, and membrane protein palmitoylation. MEND is blocked by inhibiting mitochondrial Ca uptake or PTP openings, depleting fatty acids, blocking acyl CoA synthesis, metabolizing CoA, or inhibiting palmitoylation. It is triggered by depolarizing mitochondria or promoting PTP openings. After mitochondrial MEND blockade, MEND is restored by cytoplasmic acyl CoA or CoA. MEND is blocked by siRNA knockdown of the plasmalemmal acyl transferase, DHHC5. When acyl CoA is abundant, transient H2O2 oxidative stress or PKC activation initiates MEND, but the immediate presence of H2O2 prevents MEND. The PTP inhibitor, NIM811, significantly increases plasmalemma in normally growing cells. Thus, the MEND pathway may contribute to constitutive as well as pathological plasmalemma turnover in dependence on mitochondrial stress signaling. DOI: http://dx.doi.org/10.7554/eLife.01293.001 Cells use a process called endocytosis to absorb proteins and other molecules. There are many forms of endocytosis, but they usually involve the molecule of interest becoming tucked into a bud that forms in the cell membrane. This bud is then pinched off to leave the molecule inside a vesicle that is inside the cell. In general endocytosis is triggered by ‘caging’ proteins such as clathrin, but other forms are also possible. These “non-classical” forms of endocytosis are involved in processes as diverse as the internalization of pathogens and the response of cells to wounding, and sometimes they involve large fractions of the cell membrane being pinched off. Several different forms of “massive endocytosis” have been observed, but they have remained enigmatic in comparison to the classical forms of endocytosis. Now Hilgemann et al. report a new pathway for massive endocytosis that is triggered by a sudden influx of calcium ions into Baby Hamster Kidney (BHK) fibroblasts. Up to 70% of the cell membrane can be pinched off during this process, especially areas of the membrane in which lipids and proteins are arranged in a more ordered pattern than in the average cell membrane. After massive endocytosis, it takes BHK cells about 30 minutes to replace the regions that were lost during the endocytosis. Hilgemann et al. find that calcium ions exert their influence via mitochondria, which are the primary source of energy for most cells. In contrast to all other cell organelles, the mitochondria are surrounded by two concentric membranes. The influx of calcium ions causes pores in the inner membrane of the mitochondria, called permeability transition pores, to open so that coenzyme A, a small molecule that is required for fatty acid metabolism, is released into the cytoplasm of the cell. This is followed by the condensation of coenzyme A with a fatty acid and the attachment of fatty acids to the surface membrane proteins. The attachment of these fatty acids (a process known as palmitoylation) evidently promotes ordered regions of the cell surface to coalesce and be pinched off into the cytoplasm as membrane vesicles. A key unanswered question is whether the release of coenzyme A by mitochondria regulates biochemical processes in addition to endocytosis. DOI: http://dx.doi.org/10.7554/eLife.01293.002