The ESCRT-III pathway facilitates cardiomyocyte release of cBIN1-containing microparticles.
The ESCRT-III pathway facilitates cardiomyocyte release of cBIN1-containing microparticles.
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ESCRT-III途径促进了含CBIN1的微粒的心肌细胞的释放。
DOI：
10.1371/journal.pbio.2002354
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发表时间：
2017-08
期刊：
影响因子：
9.8
通讯作者：
Hong T
中科院分区：
文献类型：
--
作者：
Xu B;Fu Y;Liu Y;Agvanian S;Wirka RC;Baum R;Zhou K;Shaw RM;Hong T
微粒（MP）是源自细胞表面质膜的细胞间通讯囊泡，但尚不清楚它们是否源自心室肌。在心室心肌细胞中，膜变形蛋白心脏桥接积分器 1（cBIN1 或 BIN1+13+17）会产生横管（t 管）膜微折叠，从而促进离子通道运输并调节局部离子浓度。微折叠产生的微结构域不断重组，适应压力来调节钙信号传导装置。我们探讨了 cBIN1 微折叠从心肌细胞外部释放的可能性。利用免疫金标记的电子显微镜成像，我们在小鼠血浆中发现cBIN1存在于大小约200 nm的膜囊泡中，这与MP的大小一致。在心脏特异性杂合 Bin1 缺失的小鼠中，流式细胞术发现血浆中的 cBIN1-MP 减少了 47%，支持心脏起源。心脏释放也通过在分离的成年小鼠心肌细胞的纯群体的培养基中检测到 cBIN1-MP 来证明。在人血浆中，渗透压休克增加了酶联免疫吸附测定 (ELISA) 检测到的 cBIN1 含量，而患有心力衰竭（一种心肌 cBIN1 减少的疾病）的人中 cBIN1 水平降低，这两者都支持人类心脏 MP 中 cBIN1 的释放。在探索 MP 释放的假定机制时，我们发现运输所需的膜裂变复合物内体分选复合物 (ESCRT)-III 亚基带电多泡体蛋白 4B (CHMP4B) 与 cBIN1 共定位并共免疫沉淀，肌动蛋白稳定增强了这种相互作用。在 cBIN1 过度表达的 HeLa 细胞中，CHMP4B 的敲除减少了 cBIN1-MP 的释放。使用截短突变体，我们确定 cBIN1 中的 N 端 BAR (N-BAR) 结构域是 CHMP4B 结合和 MP 释放所必需的。这项研究将 BAR 蛋白超家族与哺乳动物心室细胞 MP 生物发生的 ESCRT 通路联系起来，确定了细胞质 cBIN1 释放到血液中的通路元件。微粒是由细胞表面膜产生并释放到外部以与其他细胞通讯的小囊泡。我们现在证明，形成心脏主要泵室的心室肌细胞在小鼠和人类体内都释放微粒。心室微粒由心脏桥接积分蛋白 1 (cBIN1) 组织的表面膜微区产生，cBIN1 是一种膜变形蛋白，已被证明在人类心力衰竭期间会减少。在这里，我们鉴定了血液中含有 cBIN1 的微粒，这些微粒在缺乏心脏 cBIN1 表达的突变小鼠中减少。此外，导致微粒释放的过程涉及招募 CHMP4B 蛋白来剪断 cBIN1 膜。在人类中，cBIN1 存在于血液和微粒内。渗透压休克后，人体微粒破裂，从而可以通过酶联免疫吸附测定 (ELISA) 对血浆中的 cBIN1 进行定量。心力衰竭患者中测得的 cBIN1 水平大大降低。因此，我们引入了一种新的基于血液的诊断工具的生物学原理，该工具可以评估人类患者的心肌健康状况并识别衰竭的心脏。
Microparticles (MPs) are cell–cell communication vesicles derived from the cell surface plasma membrane, although they are not known to originate from cardiac ventricular muscle. In ventricular cardiomyocytes, the membrane deformation protein cardiac bridging integrator 1 (cBIN1 or BIN1+13+17) creates transverse-tubule (t-tubule) membrane microfolds, which facilitate ion channel trafficking and modulate local ionic concentrations. The microfold-generated microdomains continuously reorganize, adapting in response to stress to modulate the calcium signaling apparatus. We explored the possibility that cBIN1-microfolds are externally released from cardiomyocytes. Using electron microscopy imaging with immunogold labeling, we found in mouse plasma that cBIN1 exists in membrane vesicles about 200 nm in size, which is consistent with the size of MPs. In mice with cardiac-specific heterozygous Bin1 deletion, flow cytometry identified 47% less cBIN1-MPs in plasma, supporting cardiac origin. Cardiac release was also evidenced by the detection of cBIN1-MPs in medium bathing a pure population of isolated adult mouse cardiomyocytes. In human plasma, osmotic shock increased cBIN1 detection by enzyme-linked immunosorbent assay (ELISA), and cBIN1 level decreased in humans with heart failure, a condition with reduced cardiac muscle cBIN1, both of which support cBIN1 release in MPs from human hearts. Exploring putative mechanisms of MP release, we found that the membrane fission complex endosomal sorting complexes required for transport (ESCRT)-III subunit charged multivesicular body protein 4B (CHMP4B) colocalizes and coimmunoprecipitates with cBIN1, an interaction enhanced by actin stabilization. In HeLa cells with cBIN1 overexpression, knockdown of CHMP4B reduced the release of cBIN1-MPs. Using truncation mutants, we identified that the N-terminal BAR (N-BAR) domain in cBIN1 is required for CHMP4B binding and MP release. This study links the BAR protein superfamily to the ESCRT pathway for MP biogenesis in mammalian cardiac ventricular cells, identifying elements of a pathway by which cytoplasmic cBIN1 is released into blood. Microparticles are small vesicles generated from the cell surface membrane and externally released for communication with other cells. We now show that heart ventricular muscle cells, which form the main pumping chambers of the heart, release microparticles in both mouse and human. Ventricular microparticles arise from surface membrane microdomains organized by cardiac bridging integrator 1 (cBIN1), a membrane deformation protein that has been shown to be reduced during human heart failure. Here we identify microparticles containing cBIN1 in blood, which were reduced in mutant mice lacking heart cBIN1 expression. Furthermore, the process leading to microparticle release involves the recruitment of CHMP4B protein to snip the cBIN1 membrane. In humans, cBIN1 is present in blood and within microparticles. Upon osmotic shock, human microparticles burst, allowing for the quantification of cBIN1 in plasma by enzyme-linked immunosorbent assay (ELISA). The measured cBIN1 level was greatly reduced in patients with heart failure. Thus, we introduce the biology for a new blood-based diagnostic tool that can assess cardiac muscle health and identify failing heart in human patients.
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DOI：
10.1038/nrm2937
发表时间：
2010-08
期刊：
Nature reviews. Molecular cell biology
影响因子：
--
作者：
通讯作者：
--
影响因子：
82.9
作者：
通讯作者：
--
影响因子：
10.4
作者：
Antoniak, S.;Boltzen, U.;Rauch, U.
通讯作者：
Rauch, U.
影响因子：
4
作者：
Leach, RN;Desai, JC;Orchard, CH
通讯作者：
Orchard, CH
影响因子：
56.9
作者：
Guizetti, Julien;Schermelleh, Lothar;Gerlich, Daniel W.
通讯作者：
Gerlich, Daniel W.