Polyamine deficiency alters EGF receptor distribution and signaling effectiveness in IEC-6 cells.
Polyamine deficiency alters EGF receptor distribution and signaling effectiveness in IEC-6 cells.
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DOI：
10.1152/ajpcell.1998.274.1.c192
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发表时间：
1998
期刊：
影响因子：
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通讯作者：
S. McCormack;P. Blanner;B. Zimmerman;R. Ray;H. Poppleton;T. Patel;L. Johnson
中科院分区：
文献类型：
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作者：
S. McCormack;P. Blanner;B. Zimmerman;R. Ray;H. Poppleton;T. Patel;L. Johnson
细胞的生长和迁移是肠道上皮细胞分化、维持和修复的重要过程。表皮生长因子(EGF)是这两个过程所需的细胞骨架重组的重要因素。因为我们之前在多胺缺乏期间发现了细胞骨架的显著变化，所以有兴趣知道这些变化是否可以阻止EGF刺激生长和迁移。α-二氟甲基鸟氨酸(DFMO)是鸟氨酸脱羧酶的特异性抑制剂，是多胺生物合成的主要限速酶，可中断多胺的生物合成。DFMO抑制细胞增殖，抑制细胞迁移，EGF不能正常刺激这两种功能。转铁蛋白受体(作为内吞途径的标志)免疫细胞化学显示，EGF受体(EGFR)在与EGF结合10分钟后出现异常分布。多胺缺乏会耗尽体内微丝的细胞，使肌动蛋白皮质增厚，并阻止EGF结合的EGFR与肌动蛋白的迅速结合。EGF刺激的170 kDa蛋白酪氨酸磷酸化和纯化的膜EGFR的激酶活性降低了50%。然而，免疫沉淀的EGFR蛋白浓度并不因多胺缺乏而降低。所有这些变化都可以通过补充腐胺来预防。细胞骨架的破坏、EGFR磷酸化和激酶活性的降低、EGFR在细胞内的异常分布以及与肌动蛋白细丝的延迟结合提示了上皮细胞生长和迁移依赖于多胺的部分原因。
Cell growth and migration are essential processes for the differentiation, maintenance, and repair of the intestinal epithelium. Epidermal growth factor (EGF) is an important factor in the reorganization of the cytoskeleton required for both processes. Because we had previously found significant changes in the cytoskeleton during polyamine deficiency, it was of interest to know whether those changes could prevent EGF from stimulating growth and migration. Polyamine biosynthesis in IEC-6 cells was interrupted by treatment with α-difluoromethylornithine (DFMO), a specific inhibitor of ornithine decarboxylase, the primary rate-limiting enzyme of polyamine biosynthesis. DFMO halted cell proliferation and inhibited cell migration, and neither function could be normally stimulated by EGF. Immunocytochemistry of the transferrin receptor (used as a marker for the endocytic pathway) revealed an abnormal distribution of the EGF receptor (EGFR) 10 min after binding EGF. Polyamine deficiency depleted the cells of interior microfilaments, thickened the actin cortex, and prevented the prompt association of EGF-bound EGFR with actin. EGF-stimulated 170-kDa protein tyrosine phosphorylation and the kinase activity of purified membrane EGFR were reduced by 50%. Immunoprecipatated EGFR protein concentration, however, was not reduced by polyamine deficiency. All of these changes could be prevented by supplementation with putrescine. Cytoskeletal disruption, reduced EGFR phosphorylation and kinase activity, aberrant intracellular EGFR distribution, and delayed association with actin filaments suggest a partial explanation for the dependence of epithelial cell growth and migration on polyamines.