Glycolysis and methylaminoisobutyrate uptake in rat-1 cells transfected with ras or myc oncogenes.

Abstract

A high rate of aerobic glycolysis was catalyzed by rat-i cells transfected with a ras oncogene (ras cells); rat-i cells and rat-i cells transtected with myc oncogene (myc cells) showed a low rate of glycolysis that was increased after exposure of the cells to type B transforming growth factor (TGF-P). The uptake of radioactive methylaminoisobutyric acid or L-methionine via system A of amino acid transport also was accelerated after exposure of these cells to TGF-p, with the myc cells being most sensitive and the ras cells least sensitive. Methionine was found to be a potent inhibitor of glycolysis in ras cells as well as In rat-i or myc cells that were exposed to TGF-P. We propose a relationship between the product of the ras oncogene (p21) and the protein(s) induced by exposure to TGF-,B.In a landmark paper (1) We have observed (9) that normal rat kidney NRK-49F cells exhibited on exposure to type 3 transforming growth factor (TGF-,B) an increase in the rate of glycolysis and in the uptake of methylaminoisobutyrate (MeAIB), a specific substrate of system A of amino acid transport (10). Independently, Inman and Colowick showed that glucose uptake was stimulated by TGF-/3 in 3T3 cells (11). We also have reported (9) that glycolysis in NRK-49F cells that had been exposed to TGF-P was markedly inhibited by 25 mM methionine, a substrate for system A of amino acid transport, whereas little or no inhibition was noted in the controls that were not exposed to TGF-f3. Examination of various established transformed cell lines revealed that in all of them glycolysis was sensitive to methionine, whereas several nontransformed cell lines showed no or moderate inhibition (unpublished data). Although there were differences between cell lines with respect to the time of exposure to methionine (2-16 hr) required to give rise to an inhibition of 50% or more, thus far no exception has been observed among 10 transformed cell lines grown in tissue culture that were tested. However, glycolysis in suspended EAT cells harvested from infected mice was not inhibited by methionine under the conditions tested. Yet, EAT cells grown in tissue culture were among the most sensitive cell lines, showing a significant inhibition after only 30 min exposure to 10 mM methionine. Removal of methionine from the medium resulted in a complete reversal of the inhibition within 2 hr (unpublished data). The capacity to inhibit glycolysis was shared by other substrates transported via system A, including MeAIB, a synthetic compound that is not metabolized.In view of the fact that methionine inhibits glycolysis in transformed cell lines, we explored the effect of this amino acid and of TGF-,B on rat-1 cells and rat-1 variants transfected with ras or myc oncogenes. We describe in this paper some profound differences in glycolysis, MeAIB uptake, and response to TGF-/3 that we observed between these cell lines. MATERIALS AND METHODSThe rat-i, myc (R1-CMYC), and ras (R1-EJ2) cells were obtained from R. Weinberg. They were grown in Falcon dishes Abbrevia...

Modulation of mammalian cell proliferation by a modified tRNA base of bacterial origin

“…This has been shown for EAT cells used in this study [35]. Racker et al described that transformation of rat cells with the activated ras gene _ but not the myc gene -increases aerobic glycolysis [31], suggesting that activation of some -but not alloncogenes may cause a shift of the metabolism towards glycolysis under aerobic conditions. Glycolysis is increased also after exposure of cells to hypoxic conditions as in the case of HeLa cells.…”

“…3) suggesting that hypoxically growing cells had shifted their energy metabolism towards glycolysis. Oncogen activation has been shown to increase glycolysis and LDH activity in rat cells under aerobic conditions [31,32]. Specific LDH activity was two-fold higher in rus-transformed NIH-3T3 cells than in non-transformed cells (Fig.…”

Regulation of glut1 mRNA by Hypoxia-inducible Factor-1

“…2). A number of oncogenes, including fps, src, and ras have been shown to increase glucose transport and to up-regulate glut1 mRNA and protein levels (3)(4)(5). glut1 gene expression and glucose transport are also stimulated in a variety of cells under hypoxic conditions, a response that is mediated by the transcription factor HIF-1.…”

Ras transformation requires metabolic control by 6-phosphofructo-2-kinase

“…Stable transfection of oncogenic H-ras V12 into Rat-1 immortalized fibroblasts causes increased glucose uptake and lactate production (Racker et al, 1985;Kole et al, 1991). Moreover, recent studies by Ramanathan et al (2005) have demonstrated that the introduction of an oncogenic allele of H-ras into human fibroblasts already immortalized with the telomerase catalytic subunit, SV40 large T antigen (LT) and small T antigen causes: (i) increased glucose uptake and lactate production; (ii) increased ribose-5-phosphate, a glucose-derived anabolic precursor of DNA and RNA; and (iii) increased sensitivity to the glycolytic inhibitors, 2-deoxyglucose and oxamate.…”