SELECTIVE-INHIBITION OF RAS-DEPENDENT CELL-GROWTH BY FARNESYLTHIOSALICYLIC ACID
SELECTIVE-INHIBITION OF RAS-DEPENDENT CELL-GROWTH BY FARNESYLTHIOSALICYLIC ACID
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DOI：
10.1074/jbc.270.38.22263
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发表时间：
1995-09-22
影响因子：
4.8
通讯作者：
KLOOG, Y
中科院分区：
文献类型：
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作者：
MAROM, M;HAKLAI, R;KLOOG, Y
S-反式，反式-法尼基硫代水杨酸（FTS）是一种新的法尼基化刚性羧酸衍生物，在无细胞体系中，它作为异戊二烯化蛋白质甲基转移酶（PPMTase）的有效竞争性抑制剂（Ki = 2.6 μ M），PPMTase使大量异戊二烯化蛋白质中的羧基末端S-异戊二烯基半胱氨酸甲基化，包括HAS。FTS在各种组织和细胞系的匀浆或膜中对PPMT酶的抑制是从阻断外源添加的底物如N-乙酰基-S-反式，反式-法呢基-L-半胱氨酸和内源底物包括小GTP结合蛋白的甲基化推断的，FTS也可以抑制完整细胞中这些蛋白的甲基化（例如在Rat-1成纤维细胞、Ras转化的Rat-1和B16黑素瘤细胞中）。（50-100 μ M）是部分阻断（10-40%）完整细胞中蛋白质甲基化所需的。因此，FTS是完整细胞中甲基化的弱抑制剂，由于甲基化是Ras和相关蛋白质加工的最后一步，因此FTS不太可能影响之前的步骤，例如蛋白质异戊烯化。这可以解释为什么高达25 μ M的FTS不会影响多种培养细胞类型（包括中国仓鼠卵巢、NIH 3 T3、Rat 1、B16黑色素瘤和PC 12）的生长和大体形态，这与观察到的缺乏FTS诱导的细胞毒性一致。然而，FTS降低细胞膜中Ras的水平，并能抑制Ras依赖性细胞体外生长，与甲基化无关。它抑制人Ha-ras转化细胞（EJ细胞）的生长，并以剂量依赖性方式（0.1-10 μ M）逆转其转化形态。该药物不干扰由v-Raf或T-抗原转化的细胞的生长，但抑制ErbB 2转化的细胞的生长并阻断表皮和碱性成纤维细胞生长因子的促有丝分裂作用，因此暗示其对Ras生长信号传导的选择性，可能通过调节Ras-Raf通讯。该结果提高了FTS可能特异性干扰Ras与细胞膜中法尼基半胱氨酸识别结构域的相互作用的可能性。该药物，以及可能的其它法尼基化的刚性羧酸类似物，可用于PPMT酶的体外表征和用于细胞膜中推定的Ras法尼基半胱氨酸识别结构域的鉴定。
S-trans,trans-FarnesylthiosalicyIic acid (FTS) is a novel farnesylated rigid carboxylic acid derivative, In cell-free systems, it acts as a potent competitive inhibitor (K-i = 2.6 mu M) of the enzyme prenylated protein methyltransferase (PPMTase), which methylates the carboxyl-terminal S-prenylcysteine in a large number of prenylated proteins including has, In such systems, FTS inhibits Ras methylation but not Ras farnesylation. Inhibition of the PPMTase by FTS in homogenates or membranes of a variety of tissues and cell lines is inferred from a block in the methylation of exogenously added substrates such as N-acetyl-S-trans,trans-farnesyl-L-cysteine and of endogenous substrates including small GTP-binding proteins, FTS can also inhibit methylation of these proteins in intact cells (e.g. in Rat-1 fibroblasts, Ras transformed Rat-1, and B16 melanoma cells), Unlike in cell-free systems, however, relatively high concentrations of FTS (50-100 mu M) are required for partial blocking (10-40%) of protein methylation in the intact cells, Thus, FTS is a weak inhibitor of methylation in intact cells, Because methylation is the last step in the processing of Ras and related proteins, FTS is not likely to affect steps that precede it, e.g. protein prenylation. This may explain why the growth and gross morphology of a variety of cultured cell types (including Chinese hamster ovary, NIH3T3, Rat1, B16 melanoma, and PC12) is not affected by up to 25 mu M FTS and is consistent with the observed lack of FTS-induced cytotoxicity. Nevertheless, FTS reduces the levels of Ras in cell membranes and can inhibit Ras-dependent cell growth in vitro, independently of methylation, It inhibits the growth of human Ha-ras-transformed cells (EJ cells) and reverses their transformed morphology in a dose-dependent manner (0.1-10 mu M). The drug does not interfere with the growth of cells transformed by v-Raf or T-antigen but inhibits the growth of ErbB2-transformed cells and blocks the mitogenic effects of epidermal and basic fibroblast growth factors, thus implying its selectivity toward Ras growth signaling, possibly via modulation of Ras-Raf communication, Taken together, the results raise the possibility that FTS may specifically interfere with the interaction of Ras with a farnesylcysteine recognition domain in the cell membrane, This drug, and perhaps other farnesylated rigid carboxylic acid analogs, may be used for in vitro characterization of the PPMTase and for the identification of a putative Ras farnesylcysteine recognition domain in cell membranes.