Interactions between Fc(epsilon)RI and lipid raft components are regulated by the actin cytoskeleton.
Interactions between Fc(epsilon)RI and lipid raft components are regulated by the actin cytoskeleton.
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发表时间：
2000-03
影响因子：
4
通讯作者：
D. Holowka;Erin D. Sheets;Barbara Baird
中科院分区：
文献类型：
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作者：
D. Holowka;Erin D. Sheets;Barbara Baird
先前的研究表明，RBL-2H3肥大细胞上的IgE-Fc(Epsilon)RI复合体的交联会导致它们与分离的抗洗涤剂膜(也称为脂筏)结合，这是一个胆固醇依赖的过程，在这些受体启动信号传递之前。为了研究这些在完整细胞上的相互作用，我们用共聚焦显微镜检测了RAFT组分与交联型IgE-Fc(Epsilon)RI的共分布。在4℃下交联数小时后，糖基磷脂酰肌醇连接蛋白Thy-1和Src家族酪氨酸激酶Lyn与IgE-Fc(Epsilon)RI在质膜上以大片状重新分布。在这些条件下，F-肌动蛋白也与Fc(Epsilon)RI和RAFT组分发生戏剧性的共分离，但在37℃的短暂预热后分散。当在更高的温度下启动IgE-Fc(Epsilon)RI的交联时，除非细胞松弛素D抑制刺激的F-肌动蛋白聚合，否则不容易检测到RAFT组分与补丁Fc(Epsilon)RI的共分布。裂解细胞的蔗糖梯度分析显示，在瞬时磷酸化反应的整个过程中，交联型IgE-Fc(Epsilon)RI仍然与脂筏相关，但经历了一种时间依赖的向高密度的转变，这被细胞松弛素D阻止。我们的结果表明，Lyn和交联型IgE-Fc(Epsilon)RI之间的相互作用受到刺激的F-肌动蛋白聚合的调节，这可以用锚定的RAFT组分与更多流动性的RAFT组分的分离来解释。
Previous studies showed that crosslinking of IgE-Fc(epsilon)RI complexes on RBL-2H3 mast cells causes their association with isolated detergent-resistant membranes, also known as lipid rafts, in a cholesterol-dependent process that precedes initiation of signaling by these receptors. To investigate these interactions on intact cells, we examined the co-redistribution of raft components with crosslinked IgE-Fc(epsilon)RI using confocal microscopy. After several hours of crosslinking at 4 degrees C, the glycosylphosphatidylinositol-linked protein Thy-1 and the Src-family tyrosine kinase Lyn co-redistribute with IgE-Fc(epsilon)RI in large patches at the plasma membrane. Under these conditions, F-actin also undergoes dramatic co-segregation with Fc(epsilon)RI and raft components but is dispersed following a brief warm-up to 37 degrees C. When crosslinking of IgE-Fc(epsilon)RI is initiated at higher temperatures, co-redistribution of raft components with patched Fc(epsilon)RI is not readily detected unless stimulated F-actin polymerization is inhibited by cytochalasin D. In parallel, cytochalasin D converts transient antigen-stimulated tyrosine phosphorylation to a more sustained response. Sucrose gradient analysis of lysed cells reveals that crosslinked IgE-Fc(epsilon)RI remains associated with lipid rafts throughout the time course of the transient phosphorylation response but undergoes a time-dependent shift to higher density that is prevented by cytochalasin D. Our results indicate that interactions between Lyn and crosslinked IgE-Fc(epsilon)RI are regulated by stimulated F-actin polymerization, and this is best explained by a segregation of anchored raft components from more mobile ones.