INTERNALIZATION OF ACTIVATED PLATELET-DERIVED GROWTH-FACTOR RECEPTOR-PHOSPHATIDYLINOSITOL-3' KINASE COMPLEXES - POTENTIAL INTERACTIONS WITH THE MICROTUBULE CYTOSKELETON
INTERNALIZATION OF ACTIVATED PLATELET-DERIVED GROWTH-FACTOR RECEPTOR-PHOSPHATIDYLINOSITOL-3' KINASE COMPLEXES - POTENTIAL INTERACTIONS WITH THE MICROTUBULE CYTOSKELETON
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DOI：
10.1128/mcb.13.10.6052
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发表时间：
1993-10-01
影响因子：
5.3
通讯作者：
CORVERA, S
中科院分区：
文献类型：
--
作者：
KAPELLER, R;CHAKRABARTI, R;CORVERA, S
磷脂酰肌醇（PI）-3'激酶催化PI 3，4-二磷酸和PI 3，4，5-三磷酸响应血小板衍生生长因子（PDGF）对细胞的刺激而形成。在这里，我们报告，酪氨酸磷酸化的PDGF受体，PI-3'激酶（p85）的p85亚基，并激活PI-3'激酶被发现在孤立的网格蛋白包被的囊泡内的细胞暴露于PDGF的2分钟内，表明受体和激活PI-3'激酶进入内吞途径。免疫荧光分析显示，p85在血清饥饿的细胞呈点状/网状染色模式，集中在核周区域，并显示高焦点浓度的中心体。此外，在光学切片显微镜和图像重建后观察到p85与微管的部分共排列。微管解聚药物诺考达唑（nocodazole）进一步证明了p85与微管网络的关联，诺考达唑导致p85从核周区域重新分布到细胞周边。有趣的是，PDGF对p85分布的最显著影响是核周区域中该蛋白质的染色强度增加，并且这种影响通过用诺考达唑预先处理细胞而消除。这些结果表明，PDGF受体-p85复合物的内化和运输与微管细胞骨架。此外，在没有PDGF刺激的情况下，细胞内结构中p85的高浓度表明这种蛋白质的额外作用，与其与受体酪氨酸激酶的关联无关。
Phosphatidylinositol (PI)-3' kinase catalyzes the formation of PI 3,4-diphosphate and PI 3,4,5-triphosphate in response to stimulation of cells by platelet-derived growth factor (PDGF). Here we report that tyrosine-phosphorylated PDGF receptors, the p85 subunit of PI-3' kinase (p85), and activated PI-3' kinase are found in isolated clathrin-coated vesicles within 2 min of exposure of cells to PDGF, indicating that both receptor and activated PI-3' kinase enter the endocytic pathway. Immunofluorescence analysis of p85 in serum-starved cells revealed a punctate/reticular staining pattern, concentrated in the perinuclear region and displaying high focal concentration at the centrosome. In addition, partial coalignment of p85 with microtubules was observed after optical sectioning microscopy and image reconstruction. The association of p85 with the microtubule network was further evidenced by the microtubule-depolymerizing drug nocodazole, which caused a redistribution of p85 from the perinuclear region to the cell periphery. Interestingly, the most significant effect of PDGF on the distribution of p85 was an increase in the staining intensity of this protein in the perinuclear region, and this effect was eliminated by prior treatment of cells with nocodazole. These results suggest that PDGF receptor-p85 complexes internalize and transit in association with the microtubule cytoskeleton. In addition, the high concentration of p85 in intracellular structures in the absence of PDGF stimulation suggests additional roles for this protein independent of its association with receptor tyrosine kinases.