Alterations in the cyclic AMP signal transduction pathway regulating ribonucleotide reductase gene expression in malignant H‐ras transformed cell lines

Ribonucleotide reductase is a highly regulated activity responsible for reducing ribonucleotides to deoxyribonucleotides, which are required for DNA synthesis and DNA repair. We have tested the hypothesis that malignant cell populations contain alterations in signal pathways important in controlling the expression of the two genes that code for ribonucleotide reductase, R1 and R2. A series of radiation and H-ras transformed mouse 10T1/2 cell lines with increasing malignant potential were exposed to stimulators of cAMP synthesis (forskolin and cholera toxin), an inhibitor of cAMP degradation (3-isobutyl-1-methylxanthine) and a biologically stable analogue of cAMP (8-bromo-cAMP). Dramatic elevations in the expression of the R1 and R2 genes at the message and protein levels were observed in malignant metastatic populations, which were not detected in the normal parental cell line or in cells capable of benign tumor formation. These changes in ribonucleotide reductase gene expression occurred without any detectable modifications in the rates of DNA synthesis, showing that they were regulated by a novel mechanism independent of the S phase of the cell cycle. Furthermore, studies with forskolin (a stimulator of the protein kinase A signal pathway) and the tumor promoter 12-0-tetradecanoylphorbol-1 3-acetate (a stimulator of the protein kinase C signal pathway), alone or in combination, indicated that their effects on R1 and R2 gene expression in a highly malignant cell line were greater than when they were tested individually, suggesting that the two pathways modulating R1 and R2 gene expression can cooperate to regulate ribonucleotide reduction, and interestingly this can occur in a synergistic fashion. Also, a direct relationship between H-ras expression and ribonucleotide reductase gene expression was observed; analysis of forskolin mediated elevations in R1 and R2 message levels closely correlated with the levels of H-ras expression in the various cell lines. In total, these studies demonstrate that ribonucleotide reductase expression is controlled by a complex process, and malignant ras transformed cells contain alterations in the regulation of signal transduction pathways that lead to novel modifications in ribonucleotide reductase gene expression. This signal mechanism, which is aberrantly regulated in malignant cells, may be related to regulatory pathways involved in determining ribonucleotide reductase expression in a S phase independent manner during periods of DNA repair. (C) 1994 Wiley-Liss, Inc.PT: J; CR: ALBERT DA, 1991, P NATL ACAD SCI USA, V89, P1597 BLAKE MS, 1984, ANAL BIOCHEM, V136, P175 BOYTON AL, 1983, ADV CYCLIC NUCLEOTID, P193 CHEN FY, 1993, EMBO J, V12, P3977 CHOY BK, 1988, CANCER RES, V48, P2029 CHOY BK, 1989, BIOCHEM BIOPH RES CO, V162, P1417 EDWARDS DR, 1985, MOL CELL BIOL, V5, P3280 EGAN SE, 1987, MOL CELL BIOL, V7, P830 GILMAN AG, 1987, ANNU REV BIOCHEM, P615 GONZALEZ IL, 1985, P NATL ACAD SCI USA, V82, P7666 GOUGH NM, 1988, ANAL BIOCHEM, V173, P93 HURTA RAR, 1991, J BIOL CHEM, V266 HURTA RAR, 1992, BIOCH CELL BIOL, V70, P1081 HURTA RAR, 1992, J BIOL CHEM, V267 LAURENZA A, 1989, TRENDS PHARMACOL SCI, V10, P442 LE AQ, 1992, J BIOL CHEM, V267, P1072 LEWIS WH, 1978, J CELL PHYSL, V94, P287 MCCLARTY GA, 1986, SOMAT CELL MOLEC GEN, V12, P121 MCCLARTY GA, 1987, BIOCHEMISTRY-US, V26, P8004 MCCLARTY GA, 1988, BIOCHEMISTRY-US, V27, P7524 MCCLARTY GA, 1990, J BIOL CHEM, V265, P7539 PELECH SL, 1990, BIOCHEM CELL BIOL, V68, P1297 PHILLIPS DR, 1986, BIOCHEMISTRY-US, V25, P7355 QURESHI SA, 1991, MOL CELL BIOL, V11 SCHWARZ LC, 1988, CANCER RES, V48, P6999 SCHWARZ LC, 1990, GROWTH FACTORS, V3, P115 STRYER L, 1986, ANNU REV CELL BIOL, V2, P391 THELANDER L, 1980, J BIOL CHEM, V255, P7426 TOWBIN H, 1979, P NATL ACAD SCI USA, V76, P4350 ULLMAN B, 1980, J BIOL CHEM, V255, P8308 WEBER G, 1983, CANCER RES, V43, P3466 WEINBERG G, 1981, P NATL ACAD SCI USA, V78, P2447 WRIGHT JA, 1989, DRUG RESISTANCE MAMM, P15 WRIGHT JA, 1989, INT ENCY PHARM THERA, V128, P89 WRIGHT JA, 1990, ANTICANCER RES, V10, P1247 WRIGHT JA, 1990, BIOCH CELL BIOL, V68, P1364 WRIGHT JA, 1993, CRIT REV ONCOGENESIS, V4, P473; NR: 37; TC: 12; J9: J CELL PHYSIOL; PG: 11; GA: MM962Source type: Electronic(1