Cifuentes M E, Espinet C, Lange A J, Pilkis S J, Hod Y
Department of Physiology and Biophysics, State University of New York, Stony Brook 11794-8661.
J Biol Chem. 1991 Jan 25;266(3):1557-63.
The hormonal control of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase gene expression was studied in the rat hepatoma cells, FTO-2B. In contrast to another hepatoma cell line (HTC), the enzyme in FTO-2B cells displays both kinase and bisphosphatase activities. As in rat liver, the mRNA in FTO-2B cells is 2.2-kilobases in length. However, the 5' region of the mRNA differs from the mRNA in the liver in that it contains sequences unique to 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase mRNA from skeletal muscle. These results suggest that the mRNA in FTO-2B cells may represent an additional alternative splicing product of the 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase gene. Exposure of FTO-2B cells to media containing either insulin (10(-7) M) or dexamethasone (10(-6) M) induced about a 10-fold increase in the level of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase mRNA within 6-10 h of hormone treatment. The concentrations of insulin or dexamethasone giving half-maximal stimulation were 10(-9) M and 2 x 10(-8) M, respectively, and dibutyryl cyclic AMP (5 x 10(-7) M) completely prevented the increase in enzyme mRNA induced by these hormones. Exposure of cells to glucose-free medium abolished the insulin-mediated enhancement in 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase mRNA, but not that induced by dexamethasone. No alteration in the degradation rate of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase mRNA was noted when cells were treated with insulin. Run-on transcription assays with isolated nuclei showed an increase in the relative transcription rate of the gene in cells treated with either insulin or dexamethasone. The time course of transcription activation preceded the increase in the level of the mRNA, indicating that the main mechanism for the induction of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase expression by insulin and dexamethasone is mediated by stimulation of gene transcription.
在大鼠肝癌细胞FTO-2B中研究了6-磷酸果糖-2-激酶/果糖-2,6-二磷酸酶基因表达的激素调控。与另一种肝癌细胞系（HTC）不同，FTO-2B细胞中的该酶同时具有激酶和双磷酸酶活性。与大鼠肝脏一样，FTO-2B细胞中的mRNA长度为2.2千碱基。然而，该mRNA的5'区域与肝脏中的mRNA不同，因为它包含骨骼肌中6-磷酸果糖-2-激酶/果糖-2,6-二磷酸酶mRNA特有的序列。这些结果表明，FTO-2B细胞中的mRNA可能代表6-磷酸果糖-2-激酶/果糖-2,6-二磷酸酶基因的另一种可变剪接产物。将FTO-2B细胞暴露于含有胰岛素（10^(-7) M）或地塞米松（10^(-6) M）的培养基中，在激素处理6-10小时内，6-磷酸果糖-2-激酶/果糖-2,6-二磷酸酶mRNA水平增加了约10倍。产生半最大刺激的胰岛素或地塞米松浓度分别为10^(-9) M和2×10^(-8) M，二丁酰环磷酸腺苷（5×10^(-7) M）完全阻止了这些激素诱导的酶mRNA增加。将细胞暴露于无葡萄糖培养基中消除了胰岛素介导的6-磷酸果糖-2-激酶/果糖-2,6-二磷酸酶mRNA增强作用，但未消除地塞米松诱导的增强作用。用胰岛素处理细胞时，未观察到6-磷酸果糖-2-激酶/果糖-2,6-二磷酸酶mRNA降解速率的改变。用分离的细胞核进行的连续转录分析表明，用胰岛素或地塞米松处理的细胞中该基因的相对转录速率增加。转录激活的时间进程先于mRNA水平的增加，表明胰岛素和地塞米松诱导6-磷酸果糖-2-激酶/果糖-2,6-二磷酸酶表达的主要机制是由基因转录的刺激介导的。
