Kole H K, Resnick R J, Van Doren M, Racker E
Section of Biochemistry, Molecular and Cell Biology, Cornell University, Ithaca, New York 14853.
Arch Biochem Biophys. 1991 May 1;286(2):586-90. doi: 10.1016/0003-9861(91)90084-v.
Fructose 2,6-bisphosphate (F-2,6-P2) stimulated glycolysis in cell-free extracts of both normal and ras-transfected rat-1 fibroblasts. The extract of the transformed cell glycolyzed more rapidly in both the absence and the presence of F-2,6-P2 than the extract of the parent fibroblast. Addition of mitochondrial ATPase (F1) or inorganic phosphate (Pi) further stimulated lactate production in both cell lines. F-2,6-P2 stimulated the 6-phosphofructo-1-kinase (PFK-1) activity in extracts of normal and transfected cells. The activity in extracts of transformed cells tested with a fructose 6-phosphate regenerating system was considerably higher than in the extract of normal cells. Stimulation of PFK-1 activity by cAMP of both cell lines was not as pronounced as that by F-2,6-P2. In the absence of F-2,6-P2 the PFK-1 activity was strongly inhibited in the transformed cell by ATP concentrations higher than 1 mM, whereas in the normal cell only a marginal inhibition was noted even at 2 or 3 mM ATP. F-2,6-P2 reversed the inhibition of PFK-1 by ATP. Nicotinamide adenine dinucleotide (NAD) at 100 microM (in the presence of 2 mM ATP and 1 microM F-2,6-P2) stimulated PFK-1 activity only in the transformed cell, whereas nicotinamide adenine dinucleotide phosphate (NADP) inhibited PFK-1 activity (in the presence or absence of 1 microM F-2,6-P2) in extracts of both cell lines. No previous observations of stimulation or inhibition by NAD or NADP on PFK-1 activity appear to have been reported. A threefold increase in the intracellular concentration of F-2,6-P2 was observed after transfection of rat-1 fibroblast by the ras oncogene. We conclude from these data that the PFK-1 activity of ras-transfected rat-1 fibroblasts shows a greater response to certain stimulating and inhibitory regulating factors than that of the parent cell.
果糖2,6 - 二磷酸（F - 2,6 - P2）可刺激正常和经ras基因转染的大鼠1型成纤维细胞的无细胞提取物中的糖酵解。在有无F - 2,6 - P2的情况下，转化细胞的提取物比亲代成纤维细胞的提取物糖酵解速度更快。添加线粒体ATP酶（F1）或无机磷酸（Pi）可进一步刺激两种细胞系中的乳酸生成。F - 2,6 - P2可刺激正常和转染细胞提取物中的6 - 磷酸果糖 - 1 - 激酶（PFK - 1）活性。用6 - 磷酸果糖再生系统检测的转化细胞提取物中的活性明显高于正常细胞提取物中的活性。两种细胞系的cAMP对PFK - 1活性的刺激不如F - 2,6 - P2明显。在没有F - 2,6 - P2的情况下，ATP浓度高于1 mM时，转化细胞中的PFK - 1活性受到强烈抑制，而在正常细胞中，即使ATP浓度为2或3 mM时也仅观察到轻微抑制。F - 2,6 - P2可逆转ATP对PFK - 1的抑制作用。100微摩尔的烟酰胺腺嘌呤二核苷酸（NAD）（在存在2 mM ATP和1微摩尔F - 2,6 - P2的情况下）仅在转化细胞中刺激PFK - 1活性，而烟酰胺腺嘌呤二核苷酸磷酸（NADP）在两种细胞系的提取物中均抑制PFK - 1活性（无论是否存在1微摩尔F - 2,6 - P2）。此前似乎没有关于NAD或NADP对PFK - 1活性的刺激或抑制作用的报道。用ras癌基因转染大鼠1型成纤维细胞后，观察到细胞内F - 2,6 - P2浓度增加了三倍。我们从这些数据得出结论，经ras基因转染的大鼠1型成纤维细胞的PFK - 1活性对某些刺激和抑制调节因子的反应比亲代细胞更大。
