Pharmacological Inhibition of Dynamin II Reduces Constitutive Protein Secretion from Primary Human Macrophages
Pharmacological Inhibition of Dynamin II Reduces Constitutive Protein Secretion from Primary Human Macrophages
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DOI：
10.1371/journal.pone.0111186
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发表时间：
2014-10-27
期刊：
影响因子：
3.7
通讯作者：
Kritharides, Leonard
中科院分区：
文献类型：
--
作者：
Kockx, Maaike;Karunakaran, Denuja;Kritharides, Leonard
动力蛋白是一种裂变蛋白，介导胞内和胞外的膜事件，是药物治疗的靶点。这些研究探讨了Dynamin II是否调节构成蛋白的分泌，并首次表明，对Dynamin的药理抑制减少了载脂蛋白E(ApoE)和其他几种从原代人巨噬细胞结构性分泌的蛋白质的分泌。靶向膜上的动力素募集的抑制剂(MiTMAbs)或直接靶向GTPase结构域的抑制剂(Dygo或Dyole系列)，呈剂量和时间依赖性地减少apoE的分泌。针对所有动力蛋白II亚型的siRNA寡核苷酸证实了动力蛋白II参与了载脂蛋白E的分泌。分泌的抑制不是通过对mRNA或蛋白质合成的影响来实现的。2D-凝胶电泳法显示载脂蛋白E在高尔基体中被加工和糖基化后发生抑制，活细胞成像显示分泌受抑与高尔基体后含有apoE-GFP的囊泡运动减少有关。这种作用并不局限于巨噬细胞，也不是通过抑制剂对微管的影响来调节的。抑制Dynamin还改变了其他蛋白质的结构性分泌，减少了纤维连接蛋白、基质金属蛋白酶9、几丁质酶-3样蛋白1和溶菌酶的分泌，但意外地增加了炎症介质亲环素A的分泌。我们得出结论，Dynamin II的药物抑制剂作为一种类别效应调节巨噬细胞apoE的结构性分泌，它们调节蛋白质分泌的能力可能影响一系列生物过程。
Dynamins are fission proteins that mediate endocytic and exocytic membrane events and are pharmacological therapeutic targets. These studies investigate whether dynamin II regulates constitutive protein secretion and show for the first time that pharmacological inhibition of dynamin decreases secretion of apolipoprotein E (apoE) and several other proteins constitutively secreted from primary human macrophages. Inhibitors that target recruitment of dynamin to membranes (MiTMABs) or directly target the GTPase domain (Dyngo or Dynole series), dose- and time-dependently reduced the secretion of apoE. SiRNA oligo's targeting all isoforms of dynamin II confirmed the involvement of dynamin II in apoE secretion. Inhibition of secretion was not mediated via effects on mRNA or protein synthesis. 2D-gel electrophoresis showed that inhibition occurred after apoE was processed and glycosylated in the Golgi and live cell imaging showed that inhibited secretion was associated with reduced post-Golgi movement of apoE-GFP-containing vesicles. The effect was not restricted to macrophages, and was not mediated by the effects of the inhibitors on microtubules. Inhibition of dynamin also altered the constitutive secretion of other proteins, decreasing the secretion of fibronectin, matrix metalloproteinase 9, Chitinase-3-like protein 1 and lysozyme but unexpectedly increasing the secretion of the inflammatory mediator cyclophilin A. We conclude that pharmacological inhibitors of dynamin II modulate the constitutive secretion of macrophage apoE as a class effect, and that their capacity to modulate protein secretion may affect a range of biological processes.