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. 1990 Jun 1;268(2):465–470. doi: 10.1042/bj2680465

Phosphofructokinase 2 and glycolysis in HT29 human colon adenocarcinoma cell line. Regulation by insulin and phorbol esters.

C Denis-Pouxviel 1, T Gauthier 1, D Daviaud 1, J C Murat 1
PMCID: PMC1131455  PMID: 2163613

Abstract

Kinetic properties of phosphofructokinase 2 (PFK2) and regulation of glycolysis by phorbol 12-myristate 13-acetate (PMA) and insulin were investigated in highly glycolytic HT29 colon cancer cells. PFK2 was found to be inhibited by citrate and, to a lesser extent, by phosphoenolpyruvate and ADP, but to be insensitive to inhibition by sn-glycerol phosphate. From these kinetic data, PFK2 from HT29 cells appears different from the liver form, but resembles somewhat the heart isoenzyme. Fructose 2,6-bisphosphate (Fru-2,6-P2) levels, glucose consumption and lactate production are increased in a dose-dependent manner in HT29 cells treated with PMA or insulin. The increase in Fru-2,6-P2 can be related to an increase in the Vmax. of PFK2, persisting after the enzyme has been precipitated with poly(ethylene glycol), without change in the Km for fructose 6-phosphate. The most striking effects of PMA and insulin on Fru-2,6-P2 production are observed after long-term treatment (24 h) and are abolished by actinomycin, cycloheximide and puromycin, suggesting that protein synthesis is involved. Furthermore, the effects of insulin and PMA on glucose consumption, lactate production, Fru-2,6-P2 levels and PFK2 activity are additive, and the effect of insulin on Fru-2,6-P2 production is not altered by pre-treatment of the cells with the phorbol ester. This suggests that these effects are exerted by separate mechanisms.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Angel P., Imagawa M., Chiu R., Stein B., Imbra R. J., Rahmsdorf H. J., Jonat C., Herrlich P., Karin M. Phorbol ester-inducible genes contain a common cis element recognized by a TPA-modulated trans-acting factor. Cell. 1987 Jun 19;49(6):729–739. doi: 10.1016/0092-8674(87)90611-8. [DOI] [PubMed] [Google Scholar]
  2. Babia T., Denis-Pouxviel C., Murat J. C., Gomez-Foix A. M., Trocheris V., Guinovart J. J., Bosch F. Insulin controls key steps of carbohydrate metabolism in cultured HT29 colon cancer cells. Biochem J. 1989 Jul 1;261(1):175–179. doi: 10.1042/bj2610175. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Berridge M. J. Inositol lipids and cell proliferation. Biochim Biophys Acta. 1987 Apr 20;907(1):33–45. doi: 10.1016/0304-419x(87)90017-5. [DOI] [PubMed] [Google Scholar]
  4. Blumberg P. M. Protein kinase C as the receptor for the phorbol ester tumor promoters: sixth Rhoads memorial award lecture. Cancer Res. 1988 Jan 1;48(1):1–8. [PubMed] [Google Scholar]
  5. Bosca L., Mojena M., Diaz-Guerra J. M., Marquez C. Phorbol 12,13-dibutyrate and mitogens increase fructose 2,6-bisphosphate in lymphocytes. Comparison of lymphocyte and rat-liver 6-phosphofructo-2-kinase. Eur J Biochem. 1988 Aug 1;175(2):317–323. doi: 10.1111/j.1432-1033.1988.tb14199.x. [DOI] [PubMed] [Google Scholar]
  6. Bosca L., Rousseau G. G., Hue L. Phorbol 12-myristate 13-acetate and insulin increase the concentration of fructose 2,6-bisphosphate and stimulate glycolysis in chicken embryo fibroblasts. Proc Natl Acad Sci U S A. 1985 Oct;82(19):6440–6444. doi: 10.1073/pnas.82.19.6440. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  8. Carreras M., Bassols A. M., Carreras J., Climent F. Effects of vanadate and insulin on glucose 1,6-P2 and fructose 2,6-P2 levels in rat skeletal muscle. Biochem Int. 1988 Aug;17(2):359–366. [PubMed] [Google Scholar]
  9. Colosia A. D., Marker A. J., Lange A. J., el-Maghrabi M. R., Granner D. K., Tauler A., Pilkis J., Pilkis S. J. Induction of rat liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase mRNA by refeeding and insulin. J Biol Chem. 1988 Dec 15;263(35):18669–18677. [PubMed] [Google Scholar]
  10. Crepin K. M., Darville M. I., Hue L., Rousseau G. G. Starvation or diabetes decreases the content but not the mRNA of 6-phosphofructo-2-kinase in rat liver. FEBS Lett. 1988 Jan 25;227(2):136–140. doi: 10.1016/0014-5793(88)80884-6. [DOI] [PubMed] [Google Scholar]
  11. Denis C., Murat J. C., Trocheris V., Paris H. Variations of fructose-2,6-diphosphate levels in cultured HT29 human colon cancer cells: influence of hexoses and lactate concentrations. Int J Biochem. 1986;18(12):1123–1128. doi: 10.1016/0020-711x(86)90086-8. [DOI] [PubMed] [Google Scholar]
  12. Denis C., Paris H., Murat J. C. Hormonal control of fructose 2,6-bisphosphate concentration in the HT29 human colon adenocarcinoma cell line. Alpha 2-adrenergic agonists counteract effect of vasoactive intestinal peptide. Biochem J. 1986 Nov 1;239(3):531–536. doi: 10.1042/bj2390531. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Farnararo M., Vasta V., Bruni P., D'Alessandro A. The effect of insulin on Fru-2,6-P2 levels in human fibroblasts. FEBS Lett. 1984 Jun 4;171(1):117–120. doi: 10.1016/0014-5793(84)80470-6. [DOI] [PubMed] [Google Scholar]
  14. Fogh J., Fogh J. M., Orfeo T. One hundred and twenty-seven cultured human tumor cell lines producing tumors in nude mice. J Natl Cancer Inst. 1977 Jul;59(1):221–226. doi: 10.1093/jnci/59.1.221. [DOI] [PubMed] [Google Scholar]
  15. Franklin C. C., Chin P. C., Turner J. T., Kim H. D. Insulin regulation of glucose metabolism in HT29 colonic adenocarcinoma cells: activation of glycolysis without augmentation of glucose transport. Biochim Biophys Acta. 1988 Oct 28;972(1):60–68. doi: 10.1016/0167-4889(88)90103-6. [DOI] [PubMed] [Google Scholar]
  16. Franklin C. C., Turner J. T., Kim H. D. Regulation of Na+/K+/Cl- cotransport and [3H]bumetanide binding site density by phorbol esters in HT29 cells. J Biol Chem. 1989 Apr 25;264(12):6667–6673. [PubMed] [Google Scholar]
  17. Goodman S. A., Esau B., Koontz J. W. Insulin and phorbol myristic acetate induce ornithine decarboxylase in Reuber H35 rat hepatoma cells by different mechanisms. Arch Biochem Biophys. 1988 Nov 1;266(2):343–350. doi: 10.1016/0003-9861(88)90266-4. [DOI] [PubMed] [Google Scholar]
  18. Hue L., Blackmore P. F., Shikama H., Robinson-Steiner A., Exton J. H. Regulation of fructose-2,6-bisphosphate content in rat hepatocytes, perfused hearts, and perfused hindlimbs. J Biol Chem. 1982 Apr 25;257(8):4308–4313. [PubMed] [Google Scholar]
  19. Hue L., Rider M. H. Role of fructose 2,6-bisphosphate in the control of glycolysis in mammalian tissues. Biochem J. 1987 Jul 15;245(2):313–324. doi: 10.1042/bj2450313. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kitamura K., Kangawa K., Matsuo H., Uyeda K. Phosphorylation of myocardial fructose-6-phosphate,2-kinase: fructose-2,6-bisphosphatase by cAMP-dependent protein kinase and protein kinase C. Activation by phosphorylation and amino acid sequences of the phosphorylation sites. J Biol Chem. 1988 Nov 15;263(32):16796–16801. [PubMed] [Google Scholar]
  21. Kitamura K., Uyeda K. Purification and characterization of myocardial fructose-6-phosphate,2-kinase and fructose-2,6-bisphosphatase. J Biol Chem. 1988 Jun 25;263(18):9027–9033. [PubMed] [Google Scholar]
  22. Klip A., Ramlal T. Protein kinase C is not required for insulin stimulation of hexose uptake in muscle cells in culture. Biochem J. 1987 Feb 15;242(1):131–136. doi: 10.1042/bj2420131. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Loiseau A. M., Rider M. H., Foret D., Rousseau G. G., Hue L. Rat hepatoma (HTC) cell 6-phosphofructo-2-kinase differs from that in liver and can be separated from fructose-2,6-bisphosphatase. Eur J Biochem. 1988 Jul 15;175(1):27–32. doi: 10.1111/j.1432-1033.1988.tb14161.x. [DOI] [PubMed] [Google Scholar]
  24. Munnich A., Marie J., Reach G., Vaulont S., Simon M. P., Kahn A. In vivo hormonal control of L-type pyruvate kinase gene expression. Effects of glucagon, cyclic AMP, insulin, cortisone, and thyroid hormones on the dietary induction of mRNAs in the liver. J Biol Chem. 1984 Aug 25;259(16):10228–10231. [PubMed] [Google Scholar]
  25. Pilkis S. J., Chrisman T. D., El-Maghrabi M. R., Colosia A., Fox E., Pilkis J., Claus T. H. The action of insulin on hepatic fructose 2,6-bisphosphate metabolism. J Biol Chem. 1983 Feb 10;258(3):1495–1503. [PubMed] [Google Scholar]
  26. Pilkis S. J., el-Maghrabi M. R., Claus T. H. Hormonal regulation of hepatic gluconeogenesis and glycolysis. Annu Rev Biochem. 1988;57:755–783. doi: 10.1146/annurev.bi.57.070188.003543. [DOI] [PubMed] [Google Scholar]
  27. Richards C. S., Uyeda K. The effect of insulin and glucose on fructose-2,6-P2 in hepatocytes. Biochem Biophys Res Commun. 1982 Nov 30;109(2):394–401. doi: 10.1016/0006-291x(82)91734-x. [DOI] [PubMed] [Google Scholar]
  28. Rider M. H., Foret D., Hue L. Comparison of purified bovine heart and rat liver 6-phosphofructo-2-kinase. Evidence for distinct isoenzymes. Biochem J. 1985 Oct 1;231(1):193–196. doi: 10.1042/bj2310193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Rider M. H., Hue L. Activation of rat heart phosphofructokinase-2 by insulin in vivo. FEBS Lett. 1984 Oct 29;176(2):484–488. doi: 10.1016/0014-5793(84)81223-5. [DOI] [PubMed] [Google Scholar]
  30. Rider M. H., Hue L. Phosphorylation of purified bovine heart and rat liver 6-phosphofructo-2-kinase by protein kinase C and comparison of the fructose-2,6-bisphosphatase activity of the two enzymes. Biochem J. 1986 Nov 15;240(1):57–61. doi: 10.1042/bj2400057. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Rochette-Egly C., Kedinger M., Haffen K. Modulation of HT-29 human colonic cancer cell differentiation with calmidazolium and 12-O-tetradecanoylphorbol-13-acetate. Cancer Res. 1988 Nov 1;48(21):6173–6182. [PubMed] [Google Scholar]
  32. Rousset M., Paris H., Chevalier G., Terrain B., Murat J. C., Zweibaum A. Growth-related enzymatic control of glycogen metabolism in cultured human tumor cells. Cancer Res. 1984 Jan;44(1):154–160. [PubMed] [Google Scholar]
  33. Sato T., Ye T., Larner A., Larner J. Contrasting interactions between phorbol ester and insulin on the regulation of glycogen synthase activity and p33 mRNA accumulation in rat hepatoma cells. Arch Biochem Biophys. 1988 Jan;260(1):377–387. doi: 10.1016/0003-9861(88)90461-4. [DOI] [PubMed] [Google Scholar]
  34. Taniyama M., Kitamura K., Thomas H., Lawson J. W., Uyeda K. Isozymes of fructose 6-phosphate,2-kinase:fructose-2,6-bisphosphatase in rat and bovine heart, liver, and skeletal muscle. Biochem Biophys Res Commun. 1988 Dec 30;157(3):949–954. doi: 10.1016/s0006-291x(88)80966-5. [DOI] [PubMed] [Google Scholar]
  35. Taylor D. J., Evanson J. M., Woolley D. E. Comparative effects of interleukin 1 and a phorbol ester on rheumatoid synovial cell fructose 2,6-bisphosphate content and prostaglandin E production. Biochem Biophys Res Commun. 1988 Jan 15;150(1):349–354. doi: 10.1016/0006-291x(88)90527-x. [DOI] [PubMed] [Google Scholar]
  36. Tejwani G. A., Chauhan S., Duruibe V., Vaswani K. K. Enhancement in the activities of mouse epidermal glucose-6-phosphate dehydrogenase, hexokinase, phosphofructokinase, and pyruvate kinase by 12-O-tetradecanoyl-phorbol-13-acetate. Arch Biochem Biophys. 1985 Jun;239(2):462–466. doi: 10.1016/0003-9861(85)90712-x. [DOI] [PubMed] [Google Scholar]
  37. Van Schaftingen E., Hers H. G. Purification and properties of phosphofructokinase 2/fructose 2,6-bisphosphatase from chicken liver and from pigeon muscle. Eur J Biochem. 1986 Sep 1;159(2):359–365. doi: 10.1111/j.1432-1033.1986.tb09876.x. [DOI] [PubMed] [Google Scholar]
  38. Van Schaftingen E., Hue L., Hers H. G. Fructose 2,6-bisphosphate, the probably structure of the glucose- and glucagon-sensitive stimulator of phosphofructokinase. Biochem J. 1980 Dec 15;192(3):897–901. doi: 10.1042/bj1920897. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Viallard V., Denis C., Trocheris V., Murat J. C. Effect of glutamine deprivation and glutamate or ammonium chloride addition on growth rate, metabolism and differentiation of human colon cancer cell-line HT29. Int J Biochem. 1986;18(3):263–269. doi: 10.1016/0020-711x(86)90116-3. [DOI] [PubMed] [Google Scholar]
  40. Weinhouse S. Glycolysis, respiration, and anomalous gene expression in experimental hepatomas: G.H.A. Clowes memorial lecture. Cancer Res. 1972 Oct;32(10):2007–2016. [PubMed] [Google Scholar]
  41. de Miguel C., Gualberto A., Sobrino F. ACTH stimulates fructose 2,6-bisphosphate synthesis and glycolysis in Y-1 adrenal tumor cells. Biochem Int. 1988 Jul;17(1):69–76. [PubMed] [Google Scholar]

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