MMP-2 is localized to the mitochondria-associated membrane of the heart
MMP-2 is localized to the mitochondria-associated membrane of the heart
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DOI：
10.1152/ajpheart.00909.2013
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发表时间：
2014-03-01
影响因子：
4.8
通讯作者：
Schulz, Richard
中科院分区：
文献类型：
--
作者：
Hughes, Bryan G.;Fan, Xiaohu;Schulz, Richard
基质金属蛋白酶-2 （MMP-2）在细胞外基质重塑的背景下得到了广泛的研究，但它也定位于细胞内，可以被促氧化剂激活以水解特定的细胞间靶点。虽然有报道称线粒体中存在MMP-2，这是细胞氧化应激的一个关键来源，但这些研究没有考虑到线粒体相关膜（MAM）的存在，MAM是内质网（ER）的一个亚结构域。我们假设MMP-2位于MAM中，因此研究了其在线粒体和MAM之间的亚细胞分布。免疫金电镜显示MMP-2定位于小鼠心脏切片的线粒体中。相比之下，在HL-1心肌细胞中表达的MMP-2: HaloTag融合蛋白的免疫荧光分析显示ER样分布，与线粒体标记物MitoTracker红色相比，ER标记物（蛋白二硫异构酶）具有更大的共定位。虽然MMP-2蛋白和酶活性存在于粗粒线体中，但一旦它们被分离成纯化的线粒体和MAM， MMP-2主要与后者相关。因此，尽管线粒体可能含有最低水平的MMP-2，但先前确定为“线粒体”的大部分MMP-2实际上与MAM有关。我们还发现钙网蛋白，一种er和mam -驻地Ca2+处理蛋白和伴侣，可以在体外被MMP-2蛋白水解。因此，mam定位的MMP-2可能通过钙网蛋白水解影响er -线粒体Ca2+信号传导，从而潜在地影响线粒体功能。
Matrix metalloproteinase-2 (MMP-2) has been extensively studied in the context of extracellular matrix remodeling but is also localized within cells and can be activated by prooxidants to proteolyze specific intercellular targets. Although there are reports of MMP-2 in mitochondria, a critical source of cellular oxidative stress, these studies did not take into account the presence within their preparations of the mitochondria-associated membrane (MAM), a subdomain of the endoplasmic reticulum (ER). We hypothesized that MMP-2 is situated in the MAM and therefore investigated its subcellular distribution between mitochondria and the MAM. Immunogold electron microscopy revealed MMP-2 localized in mitochondria of heart sections from mice. In contrast, immunofluorescence analysis of an MMP-2: HaloTag fusion protein expressed in HL-1 cardiomyocytes showed an ER-like distribution, with greater colocalization with an ER marker (protein disulfide isomerase) relative to the mitochondrial marker, MitoTracker red. Although MMP-2 protein and enzymatic activity were present in crude mitochondrial fractions, once these were separated into purified mitochondria and MAM, MMP-2 was principally associated with the latter. Thus, although mitochondria may contain minimal levels of MMP-2, the majority of MMP-2 previously identified as "mitochondrial" is in fact associated with the MAM. We also found that calreticulin, an ER-and MAM-resident Ca2+ handling protein and chaperone, could be proteolyzed by MMP-2 in vitro. MAM-localized MMP-2 could therefore potentially impact mitochondrial function by affecting ER-mitochondrial Ca2+ signaling via its proteolysis of calreticulin.