Tilly B C, van Paridon P A, Verlaan I, de Laat S W, Moolenaar W H
Hubrecht Laboratory, Netherlands Institute for Developmental Biology, Utrecht.
Biochem J. 1988 Jun 15;252(3):857-63. doi: 10.1042/bj2520857.
In human A431 epidermoid carcinoma cells, epidermal growth factor (EGF) rapidly stimulates the breakdown of inositol phospholipids and raises cytoplasmic free [Ca2+]. In this paper, we investigate the action of EGF on inositol phosphate metabolism, and we compare it with the previously described effects of bradykinin on the same cell system [Tilly, van Paridon, Verlaan, Wirtz, de Laat & Moolenaar (1987) Biochem. J. 244, 129-135]. In cells prelabelled with [3H]inositol, EGF slowly but persistently (for at least 30 min) stimulates the formation of [3H]inositol phosphates, whereas bradykinin causes an immediate but transient release of inositol phosphates, which lasts for only a few minutes. The EGF effect is additive to bradykinin stimulation and does not require extracellular Ca2+. In contrast, inositol phosphate formation induced by Ca2+-ionophore A23187 has an absolute requirement for external Ca2+. Treatment of the cells with 12-O-tetradecanoylphorbol 13-acetate completely abolishes the response to EGF and to sub-optimal doses of bradykinin, suggesting a negative-feedback function of protein kinase C. Pretreatment of the cells with pertussis toxin has no effect on inositol phosphate formation induced by either EGF or bradykinin. Unlike bradykinin, EGF stimulates very little accumulation of inositol 1,4,5-trisphosphate, with only a small and rather variable release of Ca2+ from intracellular stores. EGF rapidly but transiently increases inositol 1,3,4-trisphosphate and 1,3,4,5-tetrakisphosphate, but the effects are much smaller than those of bradykinin. In addition, EGF increases both inositol mono- and bis-phosphate. At 10 min after EGF addition, inositol monophosphate, unlike the other inositol phosphates, is still increasing. It is concluded that the EGF-dependent pattern of stimulation is different from that observed in bradykinin-stimulated A431 cells, suggesting that there are separate mechanisms of inositol-lipid hydrolysis involved.
在人A431表皮癌细胞中，表皮生长因子（EGF）能迅速刺激肌醇磷脂的分解并提高细胞质游离[Ca2+]浓度。在本文中，我们研究了EGF对肌醇磷酸代谢的作用，并将其与先前描述的缓激肽对同一细胞系统的作用进行了比较[Tilly, van Paridon, Verlaan, Wirtz, de Laat & Moolenaar (1987) Biochem. J. 244, 129 - 135]。在用[3H]肌醇预标记的细胞中，EGF缓慢但持续地（至少30分钟）刺激[3H]肌醇磷酸的形成，而缓激肽则导致肌醇磷酸的立即但短暂释放，仅持续几分钟。EGF的作用与缓激肽刺激具有加和性，且不需要细胞外Ca2+。相反，Ca2+离子载体A23187诱导的肌醇磷酸形成绝对需要细胞外Ca2+。用12 - O - 十四烷酰佛波醇13 - 乙酸酯处理细胞可完全消除对EGF和次最佳剂量缓激肽的反应，提示蛋白激酶C具有负反馈功能。用百日咳毒素预处理细胞对EGF或缓激肽诱导的肌醇磷酸形成均无影响。与缓激肽不同，EGF刺激肌醇1,4,5 - 三磷酸的积累很少，从细胞内储存中释放的Ca2+量很小且变化较大。EGF迅速但短暂地增加肌醇1,3,4 - 三磷酸和1,3,4,5 - 四磷酸，但作用比缓激肽小得多。此外，EGF增加肌醇单磷酸和双磷酸。在添加EGF后10分钟，肌醇单磷酸与其他肌醇磷酸不同，仍在增加。结论是，EGF依赖的刺激模式与缓激肽刺激的A431细胞中观察到的模式不同，提示存在不同的肌醇 - 脂质水解机制。
