Phosphatidylinositol 3-kinase (PI 3-kinase) is a lipid kinase which has been implicated in mitogenesis, protein trafficking, inhibition of apoptosis, and integrin and actin functions. Here we show using a green fluorescent protein–tagged p85 subunit that phosphatidylinositol 3-kinase is distributed throughout the cytoplasm and is localized to focal adhesion complexes in resting NIH-3T3, A431, and MCF-7 cells. Ligand stimulation of an epidermal growth factor receptor/c-erbB-3 chimera expressed in these cells results in a redistribution of p85 to the cell membrane which is independent of the catalytic activity of the enzyme and the integrity of the actin cytoskeleton. The movement is, however, dependent on the phosphorylation status of the erbB-3 chimera. Using rhodamine-labeled epidermal growth factor we show that the phosphatidylinositol 3-kinase and the receptors colocalize in discrete patches on the cell surface. Low concentrations of ligand cause patching only at the periphery of the cells, whereas at high concentrations patches were seen over the whole cell surface. Using green fluorescent protein–tagged fragments of p85 we show that binding to the receptor requires the NH2-terminal part of the protein as well as its SH2 domains.

Overexpression of the liver subunit of 6-phosphofructo-1-kinase in Chinese hamster ovary K1 cells was shown to increase the steady-state level of the enzyme's product, fructose 1,6-bisphosphate, and to produce a small but significant decrease in the concentration of fructose 2,6-bisphosphate, which is an allosteric activator of the enzyme. However, overexpression of the enzyme had no effect on glycolytic flux under a variety of different substrate conditions. This latter observation is consistent with similar studies in fungi and in potato tubers which indicate that 6-phosphofructo-1-kinase has very little control over flux in glycolysis.

31P NMR measurements on extracts prepared from a variety of cultured mammalian cell lines and primary rat hepatocytes have shown changes in the levels of several phospholipid metabolites after incubation of cells with unsaturated fatty acids. These data suggest a possible link between the accumulation of neutral lipid and the changes in phospholipid metabolite concentrations that have been observed in some tumor cells and other rapidly growing tissues such as the regenerating liver and mitogen-stimulated lymphocytes.

The techniques of NMR spectroscopy and molecular genetics have provided new and powerful approaches to studying the control and organisation of cellular metabolism in vivo. We review here our recent applications of these methodologies to the study of energy metabolism in yeast and mammalian cells.

The techniques of NMR spectroscopy and molecular genetics have provided new and powerful approaches to studying the control and organisation of cellular metabolism in vivo. We review here our recent applications of these methodologies to the study of energy metabolism in yeast and mammalian cells.