Altin J G, Kujubu D A, Raffioni S, Eveleth D D, Herschman H R, Bradshaw R A
Department of Biological Chemistry, College of Medicine, University of California, Irvine 92717.
J Biol Chem. 1991 Mar 25;266(9):5401-6.
As a measure of the transmembrane signals that they transduce, two neurotrophic agents, nerve growth factor (NGF) and basic fibroblast growth factor (bFGF), and the muscarinic agonist carbachol were compared for their ability to induce TIS (tetradecanoyl phorbol acetate-inducible sequences) transcripts, representing a family of immediate early response genes, in the rat pheochromocytoma cell line PC12 and the morphologically unresponsive variant PC12nnr5. Three genes, TIS1 (also designated NGFIB), TIS8 (also designated NGFIA), and TIS21, induced in these cells by NGF (Kujubu, D.A., Lim, R.W., Varnum, B.C., and Herschman, H.R. (1987) Oncogene 1, 257-262, 1987), are also induced by bFGF and carbachol. In native PC12 cells the level of expression of TIS8 and TIS21 is similar for all three stimuli, as well as for tetradecanoyl phorbol acetate (TPA). In contrast, the induction of TIS1 by NGF and TPA is slight and is only just detectable after stimulation by bFGF, but is strong for carbachol. Thus, although all of these agents can stimulate protein kinase (PK-C), at least one TIS gene can apparently be differentially regulated by these ligands, suggesting that alternative signaling pathways must also exist. In keeping with this view, bFGF, and to a lesser degree NGF, can elicit a TIS gene response in PC12 cells in which PK-C has been down-regulated with TPA. The response to carbachol (and TPA) is effectively blocked under these conditions. Since both NGF and bFGF stimulate neurite outgrowth in such cells, PK-C is apparently not essential, i.e. does not represent the sole mechanism, for signal transduction leading to modulation of gene expression for these factors. Consistent with this model, putative protein kinase inhibitors, K252a and sphingosine, did not inhibit the TIS gene responses to bFGF. However, these agents also failed to block TIS gene responses to carbachol and TPA indicating that they were ineffective as PK-C inhibitors under these conditions. The NGF-induced response was, however, blocked by K252a indicating a unique step in the mechanism of this factor not shared by the other ligands. Sphingosine did not block TIS induction with NGF. The mutant cell line PC12 nnr5 does not respond morphologically to either NGF or bFGF. However, TIS gene responses to bFGF are unaffected, whereas those to NGF are completely abolished. The response to TPA is altered quantitatively but not qualitatively; the induction by carbachol is largely eliminated, apparently as a result of a 90% reduction in muscarinic receptors.(ABSTRACT TRUNCATED AT 400 WORDS)
作为对它们所转导的跨膜信号的一种衡量，比较了两种神经营养因子，即神经生长因子（NGF）和碱性成纤维细胞生长因子（bFGF），以及毒蕈碱激动剂卡巴胆碱在大鼠嗜铬细胞瘤细胞系PC12和形态学上无反应的变体PC12nnr5中诱导TIS（十四烷酰佛波醇乙酸酯诱导序列）转录本的能力，这些转录本代表了一个即时早期反应基因家族。NGF（库朱布，D.A.，林，R.W.，瓦尔纳姆，B.C.，和赫希曼，H.R.（1987年）《癌基因》1，257 - 262，1987年）在这些细胞中诱导的三个基因，即TIS1（也称为NGFIB）、TIS8（也称为NGFIA）和TIS21，也可被bFGF和卡巴胆碱诱导。在天然PC12细胞中，对于所有三种刺激物以及十四烷酰佛波醇乙酸酯（TPA），TIS8和TIS21的表达水平相似。相比之下，NGF和TPA对TIS1的诱导很轻微，仅在bFGF刺激后勉强可检测到，但对卡巴胆碱的诱导很强。因此，尽管所有这些试剂都能刺激蛋白激酶（PK - C），但至少有一个TIS基因显然可被这些配体差异调节，这表明也必定存在其他信号通路。与此观点一致的是，bFGF，以及程度较轻的NGF，能在PK - C已被TPA下调的PC12细胞中引发TIS基因反应。在这些条件下，对卡巴胆碱（和TPA）的反应被有效阻断。由于NGF和bFGF都能刺激此类细胞中的神经突生长，PK - C显然不是这些因子导致基因表达调节的信号转导所必需的，即不代表唯一机制。与该模型一致的是，推定的蛋白激酶抑制剂K252a和鞘氨醇并未抑制TIS基因对bFGF的反应。然而，这些试剂也未能阻断TIS基因对卡巴胆碱和TPA的反应，表明它们在这些条件下作为PK - C抑制剂无效。然而，K252a阻断了NGF诱导的反应，表明该因子的机制中有一个其他配体所没有的独特步骤。鞘氨醇未阻断NGF诱导的TIS反应。突变细胞系PC12 nnr5对NGF或bFGF在形态上均无反应。然而，TIS基因对bFGF的反应未受影响，而对NGF的反应则完全消失。对TPA的反应在数量上有所改变但性质未变；卡巴胆碱的诱导作用在很大程度上被消除，显然是由于毒蕈碱受体减少了90%。（摘要截短至400字）
