di Campli A, Valderrama F, Babià T, De Matteis M A, Luini A, Egea G
Department of Cell Biology and Oncology, Istituto di Ricerche Farmacologiche Mario Negri, Consorzio Mario Negri Sud, Santa Maria Imbaro (Chieti), Italy.
Cell Motil Cytoskeleton. 1999;43(4):334-48. doi: 10.1002/(SICI)1097-0169(1999)43:4<334::AID-CM6>3.0.CO;2-3.
In this report we have studied the morphological changes of the Golgi complex (GC) that specifically accompany F-actin reorganizations. In starved rat RBL-2H3 tumor mast cells, the GC, that was visualized at immunofluorescence level with antibodies raised against the Golgi-resident proteins giantin, mannosidase II, or TGN-38, showed a compacted morphology with a supranuclear positioning. Concomitant to membrane ruffle formation induced by epidermal growth factor (EGF) or phorbol 12-myristate 13-acetate (PMA), and stress fiber formation induced by lysophosphatidic acid (LPA), specific GC morphological changes were observed. When cells were stimulated with EGF or PMA, the compacted GC morphology was transformed into a reticular network that was extended towards the cell periphery. When cells were incubated with LPA, the GC acquired a characteristic ring-shaped morphology. Brefeldin A (BFA) did not affect the PMA- or LPA-induced membrane ruffling and stress fiber formation, respectively, indicating that actin rearrangements occurred independent of the presence of the GC. Upon BFA removal, the presence of PMA or LPA during the recovery process induced the GC to acquire the morphological appearance described above for each agent. Moreover, the PMA- but not the LPA-induced GC rearrangements were sensitive to the actin perturbing agents cytochalasin D and jasplakinolide. When cells were preincubated with the phosphatidylinositide 3-kinase (PI3K) inhibitors wortmannin or LY294002, the PMA-induced GC morphological changes were inhibited but not membrane ruffles. Finally, the PMA-induced increase in the post-Golgi transport of glycosaminoglycans to the cell surface was not altered by cytochalasin D or jasplakinolide. Altogether, these data suggest that: (1) the shape of the GC is influenced by the 3D arrangement of actin microfilaments; (2) PI3K regulates the association of the GC with actin microfilaments; and (3) actin microfilaments are not essential for the post-Golgi transport to the plasma membrane.
在本报告中，我们研究了与F-肌动蛋白重组特异性相关的高尔基体复合物（GC）的形态变化。在饥饿的大鼠RBL-2H3肿瘤肥大细胞中，通过用针对高尔基体驻留蛋白巨蛋白、甘露糖苷酶II或TGN-38产生的抗体在免疫荧光水平可视化的GC，呈现出一种致密形态并位于核上。伴随着表皮生长因子（EGF）或佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA）诱导的膜皱褶形成，以及溶血磷脂酸（LPA）诱导的应力纤维形成，观察到了特定的GC形态变化。当用EGF或PMA刺激细胞时，致密的GC形态转变为向细胞周边延伸的网状网络。当细胞与LPA孵育时，GC获得了特征性的环形形态。布雷菲德菌素A（BFA）分别不影响PMA或LPA诱导的膜皱褶和应力纤维形成，表明肌动蛋白重排独立于GC的存在而发生。去除BFA后，恢复过程中PMA或LPA的存在诱导GC获得上述每种试剂所描述的形态外观。此外，PMA诱导的而非LPA诱导的GC重排对肌动蛋白干扰剂细胞松弛素D和茉莉酸内酯敏感。当细胞用磷脂酰肌醇3-激酶（PI3K）抑制剂渥曼青霉素或LY294002预孵育时，PMA诱导的GC形态变化受到抑制，但膜皱褶不受影响。最后，细胞松弛素D或茉莉酸内酯未改变PMA诱导的糖胺聚糖从高尔基体后运输到细胞表面的增加。总之，这些数据表明：（1）GC的形状受肌动蛋白微丝的三维排列影响；（2）PI3K调节GC与肌动蛋白微丝的关联；（3）肌动蛋白微丝对于从高尔基体后运输到质膜不是必需的。
