Heat shock protein 27 regulates neutrophil chemotaxis and exocytosis through two independent mechanisms
Heat shock protein 27 regulates neutrophil chemotaxis and exocytosis through two independent mechanisms
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DOI：
10.4049/jimmunol.178.4.2421
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发表时间：
2007-02-15
影响因子：
4.4
通讯作者：
McLeish, Kenneth R.
中科院分区：
文献类型：
--
作者：
Jog, Neelakshi R.;Jala, Venkatakrishna R.;McLeish, Kenneth R.
负责调节中性粒细胞趋化性和胞吐作用的p38 MAPK途径的靶点尚不清楚。该途径的一个靶点是肌动蛋白结合蛋白，热休克蛋白27（Hsp 27）。因此，我们测试的假设，热休克蛋白27介导p38 MAPK依赖的趋化性和胞吐作用，在人类中性粒细胞通过调节肌动蛋白重组。通过引入抗Hsp 27抗体而不是同种型抗体来隔离Hsp 27，抑制fMLP刺激的趋化性，在没有fMLP刺激的情况下增加皮质F-肌动蛋白，并抑制fMLP刺激的胞吐作用。用latrunculin A预处理防止肌动蛋白重组和由Hsp 27隔离诱导的fMLP刺激的胞吐的变化。为了确定Hsp 27磷酸化的作用，通过电穿孔将野生型、磷酸化抗性或磷酸化模拟重组Hsp 27引入中性粒细胞。抗磷酸化突变体显著减少了向fMLP的迁移，而没有一个Hsp 27蛋白影响fMLP刺激或TNF-α刺激的胞吐或肌动蛋白聚合。内源性热休克蛋白27与F-肌动蛋白共定位在未受刺激和fMLP刺激的中性粒细胞，而磷酸化的热休克蛋白27除了与F-肌动蛋白共定位显示胞质定位。我们的研究结果表明，热休克蛋白27调节中性粒细胞的趋化性和胞吐作用的肌动蛋白依赖性，磷酸化独立的方式。Hsp 27的磷酸化调节趋化性，而不是胞吐作用，独立于肌动蛋白重组的调节。
The targets of the p38 MAPK pathway responsible for regulation of neutrophil chemotaxis and exocytosis are unknown. One target of this pathway is the actin-binding protein, heat shock protein 27 (Hsp27). Therefore, we tested the hypothesis that Hsp27 mediates p38 MAPK-dependent chemotaxis and exocytosis in human neutrophils through regulation of actin reorganization. Sequestration of Hsp27 by introduction of anti-Hsp27 Ab, but not an isotype Ab, inhibited fMLP-stimulated chernotaxis, increased cortical F-actin in the absence of fMLP stimulation, and inhibited fMLP-stimulated exocytosis. Pretreatment with latrunculin A prevented actin reorganization and the changes in fMLP-stimulated exocytosis induced by Hsp27 sequestration. To determine the role of Hsp27 phosphorylation, wild-type, phosphorylation-resistant, or phosphorylation-mimicking recombinant Hsp27 was introduced into neutrophils by electroporation. The phosphorylation-resistant mutant significantly reduced migration toward fMLP, whereas none of the Hsp27 proteins affected fMLP-stimulated or TNF-alpha-stimulated exocytosis or actin polymerization. Endogenous Hsp27 colocalized with F-actin in unstimulated and fMLP-stimulated neutrophils, whereas phosphorylated Hsp27 showed cytosolic localization in addition to colocalization with F-actin. Our results suggest that Hsp27 regulates neutrophil chemotaxis and exocytosis in an actin-dependent, phosphorylation-independent manner. Phosphorylation of Hsp27 regulates chernotaxis, but not exocytosis, independent of regulation of actin reorganization.