Stimulation of 2-deoxyglucose uptake in growth-inhibited BALB/c 3T3 and revertant SV40-transformed 3T3 cells

Abstract

Sugar transport in the contact-inhibited BALB/c 3T3 and revenant SV40-transformed (contact-inhibited variants of transformed cells described previously) 3T3 cells was markedly diminished when these cells became growth-inhibited [22]. Several agents have been tested for their ability to stimulate transport of 2-deoxyglucose in growth-inhibited cells and their mechanism of action has been examined in some detail. Insulin rapidly stimulated transport in both normal 3T3 and revertant cells, when they were contact-inhibited, by increasing the Michaelis-Menton kinetic parameter V max—possibly a rapid increase in the number of transport sites on the surface of these cells. Insulin had no effect on sugar transport in SV40-transformed (SVT2) cells, growing 3T3 cells, or growing revertant cells. Increasing concentrations of fresh fetal calf serum caused a stimulation of transport in contact-inhibited 3T3 and revertant cells to levels identical with growing cells (there was no effect on growing cells or on SVT2 cells); this correlated with its ability to ‘turn-on’ DNA synthesis in the vast majority of these cells. This stimulation was maximal at serum concentrations of 35–80%. The mechanisms of stimulation for the two cell types were quite different—stimulation of transport in 3T3 cells was initially rapid with an increase in V max followed 1 h later by a second increase resulting from a diminished K m (an increased binding affinity). Stimulation in revertant cells proceeded slowly and linearly and was a result of an increased V max with no apparent change in the K m constant. Although revertant cells exhibited sugar transport properties which in general resembled those same properties in normal 3T3 cells and not of their parental SVT2 cells, the mechanisms of transport changes were quite different. The nature of the interaction of fresh fetal calf serum with BALB/c 3T3 cells in terms of stimulation of transport was examined in some detail. The serum was shown to be partially multipotential, while the cells were shown not to be. Serum was modified in that it lost its ability to cause the second, long-term stimulation of transport mentioned above ( K m decrease); this correlated with the loss of its ability to induce DNA synthesis in these cells. These experiments suggest a unique approach to the study of the interaction between cells and serum and the importance of this interaction in growth control of normal cells and its loss by virally-transformed cells.