Down-regulation of protein kinase C and of an endogenous 80-kDa substrate in transformed fibroblasts.
Down-regulation of protein kinase C and of an endogenous 80-kDa substrate in transformed fibroblasts.
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DOI：
10.1016/s0021-9258(18)49290-3
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发表时间：
1987-12
期刊：
影响因子：
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通讯作者：
A. Wolfman;T. Wingrove;P. Blackshear;I. Macara
中科院分区：
文献类型：
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作者：
A. Wolfman;T. Wingrove;P. Blackshear;I. Macara
NIH-3 T3成纤维细胞的Ha-ras，Ki-v-ras，v-src和v-fms癌基因蛋白转化的亚融合培养物都具有升高的稳态水平的二酰甘油，蛋白激酶C的内源性激活剂，与非转化的亲本系相比。这些癌基因转化的成纤维细胞也表现出显著降低的细胞蛋白激酶C活性水平，通过四种不同的标准测量：佛波酯刺激的内源性80千道尔顿（80 kDa）底物的磷酸化;佛波酯刺激的86 Rb摄取的变化;酶测定;和[3 H]佛波酯结合。在所有情况下，转化的细胞表现出减弱的响应佛波酯添加和较低的佛波酯结合能力相比，亲本系。蛋白激酶C的内源性80-kDa底物的Western分析显示，在转化细胞中这种蛋白质的水平显著低于未转化的对照组，并且这种降低可以通过与佛波醇酯长期孵育在亲本细胞中模拟，表明80-kDa蛋白质的水平受蛋白激酶C的活化状态调节。这些效应似乎不是对转化细胞自分泌的非特异性反应。它们可能代表转化细胞负调节癌基因产物产生的组成性增殖信号的不成功尝试。
Subconfluent cultures of NIH-3T3 fibroblasts transformed by the Ha-ras, Ki-v-ras, v-src, and v-fms oncogene proteins all possess elevated steady-state levels of diacylglycerol, the endogenous activator of protein kinase C, as compared to the nontransformed parental lines. These oncogene-transformed fibroblasts also exhibit a significantly decreased level of cellular protein kinase C activity as measured by four different criteria: phorbol ester-stimulated phosphorylation of an endogenous 80-kilodalton (80 kDa) substrate; phorbol ester-stimulated changes in 86Rb uptake; enzymatic assay; and [3H]phorbol ester binding. In all cases, the transformed cells demonstrated an attenuated response to phorbol ester addition and a lower phorbol ester binding capacity as compared to the parental lines. Western analysis of the endogenous 80-kDa substrate of protein kinase C revealed a significantly lower level of this protein in the transformed cells than in the untransformed controls, and this decrease could be mimicked in parental cells by long-term incubation with phorbol esters, suggesting that the level of the 80-kDa protein is regulated by the state of activation of protein kinase C. These effects do not appear to be nonspecific responses to autocrine secretions by the transformed cells. They may represent an unsuccessful attempt by the transformed cells to negatively modulate the constitutive proliferative signals generated by the oncogene products.