Rodland K D, Muldoon L L, Lenormand P, Magun B E
Department of Cell Biology and Anatomy, Oregon Health Sciences University, Portland 97201-3098.
J Biol Chem. 1990 Jul 5;265(19):11000-7.
We investigated the role of intracellular [Ca2+] in mediating the independent signal transduction pathways leading to induction of VL30 RNA expression by multiple agonists in the Rat-1-derived RVL-3 cell line. This cell line contains a single integrated VL30 element, and displays a rapid transcriptional activation of VL30 following stimulation by epidermal growth factor, endothelin, or the phorbol ester tumor promoter 12-O-tetradecanoylphorbol acetate (Rodland, K. D., Brown, A. M. C., and Magun, B. E. (1987) M. Cell. Biol. 7, 2296-2298). Neither epidermal growth factor nor endothelin is dependent upon protein kinase C for activation of VL30 expression, as both of these agonists induce normal levels of VL30 RNA expression, even in cells which have been severely depleted of protein kinase C following chronic 12-O-tetradecanoylphorbol acetate exposure. Induction of VL30 RNA expression by either endothelin or 12-O-tetradecanoylphorbol acetate was blocked by concomitant exposure of RVL-3 cells to the intracellular Ca2(+)-chelating agent 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid at a concentration sufficient to buffer intracellular [Ca2+] below 200 nM, and VL30 RNA was induced by the application of the Ca2+ ionophore A23187 in the absence of agonist. Normal levels of VL30 expression in response to epidermal growth factor were observed at 165 nM [Ca2+], but were significantly inhibited at 115 nM [Ca2+]. Both the protein kinase C-dependent and protein kinase C-independent pathways leading to VL30 transcription were dependent upon the presence of an intracellular [Ca2+] exceeding 115 nM. The dependence upon intracellular Ca2+ transients for transcriptional induction by endothelin appears to be a characteristic of VL30 expression, as 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid treatment did not prevent the endothelin-induced transcription of the protooncogenes c-jun and c-fos.
我们研究了细胞内[Ca2+]在介导独立信号转导途径中的作用，该途径可导致大鼠1衍生的RVL-3细胞系中多种激动剂诱导VL30 RNA表达。该细胞系含有单个整合的VL30元件，在表皮生长因子、内皮素或佛波酯肿瘤启动子12-O-十四酰佛波醇-13-乙酸酯（TPA）刺激后，VL30会迅速发生转录激活（罗德兰，K.D.，布朗，A.M.C.，马贡，B.E.（1987年）《分子细胞生物学》7，2296 - 2298）。表皮生长因子和内皮素激活VL30表达均不依赖蛋白激酶C，因为即使在长期暴露于TPA后蛋白激酶C严重耗竭的细胞中，这两种激动剂仍能诱导正常水平的VL30 RNA表达。内皮素或TPA诱导VL30 RNA表达可被RVL-3细胞同时暴露于细胞内Ca2+螯合剂1,2-双（邻氨基苯氧基）乙烷-N,N,N',N'-四乙酸所阻断，该螯合剂浓度足以将细胞内[Ca2+]缓冲至200 nM以下，并且在无激动剂情况下，应用Ca2+离子载体A23187可诱导VL30 RNA表达。在[Ca2+]为165 nM时观察到对表皮生长因子的正常VL30表达水平，但在[Ca2+]为115 nM时显著受到抑制。导致VL30转录的蛋白激酶C依赖性和蛋白激酶C非依赖性途径均依赖于细胞内[Ca2+]超过115 nM。内皮素转录诱导对细胞内Ca2+瞬变的依赖性似乎是VL30表达的一个特征，因为1,2-双（邻氨基苯氧基）乙烷-N,N,N',N'-四乙酸处理并未阻止内皮素诱导的原癌基因c-jun和c-fos的转录。
