Cyclic AMP and cyclic GMP concentrations in serum- and density-restricted fibroblast cultures.

Abstract

Mouse fibroblasts transformed by simian virus 40 (SV3T3 cells) are characterized by cyclic AMP and cyclic GMP levels, respectively, about half and twice those found in growing untransformed 3T3 cells. Densitydependent inhibition of growth is correlated with reduced cyclic GMP concentrations in 3T3 and four different density-restricted revertant lines derived from SV3T3. The levels of cyclic AMP are not increased at confluence. Upon serum restriction, serum-dependent cell lines show a greater increase in intracellular cAMP than seruminsensitive lines. Cyclic GMP levels are greatly reduced, even in serum-insensitive density revertants, but not in SV3T3. Serum readdition to all serum-dependent lines is followed by a rapid decrease in cyclic AMP and increase in cyclic GMP concentrations. The magnitude of these responses is decreased in SV3T3 and density revertants.The growth of nontransformed fibroblasts, such as the mouse 3T3 line, is regulated by serum factors and cell-to-cell contact (1-3). Transition from a growing state to a resting state occurs upon depletion or deprivation of serum and upon cell confluence. Simian virus 40 (SV40) virus-transformed 3T3 clones often lose sensitivity to these environmental factors (3). Using negative selection pressures on SV40-transformed 3T3 fibroblasts, one can isolate revertant cell lines; they have regained growth control by serum or cell density or by both signals (4-6).Recent work from various laboratories has implicated 3': 5'-cyclic AMP (cANIP) and 3': 5'-cyclic GMIP (cGMP) in the regulation of proliferation in fibroblast cultures.On one hand, the induction of fibroblast proliferation, by a variety of mitogenic signals, is associated with early decreases in intracellular cAMP (7-15) and increases in the cGMP content (14,15).On the other hand, the addition of dibutyryl cyclic AMP to the culture medium partially prevents the stimulation of cell growth (12, 13), whereas exogenous addition of cyclic GMP counteracts the inhibitory effects of dibutyryl cyclic AMP upon a number of biochemical processes activated early in the mitogenic response (16 We report here that density-dependent inhibition of growth is correlated with decreased cGMP concentrations in 3T3 and four different lines of density-dependent revertant cell lines. In contrast, the cGMMP levels remain almost unaffected over the same range of cell densities, in the non "contact-inhibited" clone SV101 of SV40-transformed 3T3 fibroblasts. MATERIALS AND METHODSCell Lines. The cell lines used in this study are Swiss 3T3, clone SV101 of SV40-transformed 3T3, and four revertant lines derived from SV101 by Pollack and coworkers (4-6). The density-dependent revertant clones FlSV101 and BuSV2 were obtained by negative selection with fluorodeoxyuridine and bromodeoxyuridine, respectively (4, 5). The serum-dependent clones (LsSV2 and AySV5) were selected for their inability to grow at low serum concentrations (1%) and in gammadepleted (= agamma) 10% calf serum, respectively (6). The density revertants have regained den...

Cystathionine β‐synthase is coordinately regulated with proliferation through a redox‐sensitive mechanism in cultured human cells and Saccharomyces cerevisiae

“…probably induced by characteristic changes in the ratio of cAMP to cGMP (Moens et al, 1974(Moens et al, , 1975. Previous work has also suggested that CBS is regulated at least in part by cAMP (Goss, 1986).…”

Multiple phosphorylation of ribosomal protein S6 during transition of quiescent 3T3 cells into early G1, and cellular compartmentalization of the phosphate donor.

“…However, in serum-stimulated 3T3 cells, cyclic AMP levels are greatly decreased (35)(36)(37)(38). On the other hand, cyclic GMP levels have been reported to be increasing (36,37) and this has led others (27) to suggest that this compound may be the S6-phosphokinase activator. But again there is a great deal of controversy concerning whether cyclic GMP levels are increasing or decreasing during this time (38,39).…”

Induction of c-fos and c-myc mRNA by epidermal growth factor or calcium ionophore is cAMP dependent.

“…Pruss and Herschman (20) and Pledger and coworkers (22) have shown that in some cell lines the mitogenic activity of EGF is potentiated by cholera toxin, which activates adenylate cyclase thereby raising intracellular cAMP. In addition, earlier studies showed that serum deprivation increased intracellular cAMP in some cell lines (28)(29)(30). We, therefore, determined the cAMP level in density-arrested and serum-deprived A31 cells and found nearly twice as much as in serum-deprived cells (Fig.…”