Search for cellular phosphorylation products of D-allose.

The all-cis hexose D-allose, fed to hamster fibroblast cultures over 14-20 hr, brings about a striking down-regulation of hexose transport. This down-regulation by allose is inhibited by 2,4-dinitrophenol. By using D-[14C]allose and thin-layer chromatography, two types of products accumulated that have been identified as allose and allose phosphate.

50 years of biological research--from oxidative phosphorylation to energy requiring transport regulation.

In 1930 adenosine triphosphate appeared in the literature from W. A. Engelhardt's work on avian erythrocytes.

Concerted hexose transport curb by tunicamycin is rendered irreversible by glucose or allose in medium containing L-glutamine.

The hexose transport in a hamster fibroblast mutant (DS7), unable to use glucose for generation of energy, is nevertheless subject to a marked down-regulation ("curb") after prolonged incubation of monolayer cultures with glucose; fructose is unable to exert any curb. D-Allose, an all-cis hexose, mediates a vigorous curb of the transport system. Moreover, prolonged coincubation of glucose or allose with tunicamycin (TM) brings about an additional effect that is not an inhibition of the transport system, which we shall call the "concerted" transport curb.

Tunicamycin tightens the glucose- and D-allose-mediated control of hexose transport in a metabolic fibroblast mutant.

The hexose transport system of a fibroblast mutant, DS7, unable to convert glucose 6-phosphate to fructose 6-phosphate ("the phosphoglucose isomerase mutant"), is subject to a specific down-regulation ("curb") evoked by only glucose or D-allose. Neither fructose nor mannose has a curbing effect on this mutant. Further addition of tunicamycin intensified the transport curb on the mutant mediated by glucose or allose.

Oligonucleotides with rapid turnover of the phosphate groups occur endogenously in eukaryotic cells.

Endogenous oligonucleotides were found in trichloroacetic acid extracts of hamster lung fibroblasts and Tetrahymena cells. Peaks of radioactivity that eluted with retention times similar to oligonucleotide markers (5- to 50-mer) were found by HPLC in cells labeled briefly with 32Pi. Only minute amounts of UV-absorbing material were detected, consistent with a rapid turnover of phosphate groups.

Hexose transport control in a fibroblast metabolic mutant can be promoted more effectively by D-allose than by glucose.

By studying the energy-requiring control of the hexose transport system (the transport "curb") in a lung fibroblast mutant called the phosphoglucose isomerase mutant (because it is devoid of the enzyme phosphoglucose isomerase) the following features were noted. The aldohexose D-allose, if added over 20 hr to a culture of the mutant, promotes the development of an intense curb of the hexose transport system, greatly surpassing that brought about by incubation with glucose. The allose-mediated curb can be circumvented by various metabolic inhibitors as well as by the presence of other aldohexoses such as mannose.

Insulin binding and insulin action in cultured fibroblasts: significant differences between a phosphoglucose isomerase-deficient mutant and the parental strain.

The interrelationships of hexose feeding and insulin action were studied in the Chinese hamster fibroblast cell lines 023 and DS-7. The latter, derived from 023 and deficient in phosphoglucose isomerase, has been used to map the metabolic requirements for aldohexose-mediated down-regulation or "curbing" of hexose transport. We have characterized insulin binding and the response to insulin in both cell lines to determine if the insulin-mediated stimulation of transport is similarly dependent on hexose metabolism.

Mutations in the phosphoglucose isomerase gene can lead to marked alterations in cellular ATP levels in cultured fibroblasts exposed to simple nutrient shifts.

The generation of ATP in a hamster fibroblast mutant devoid of the enzyme phosphoglucose isomerase (PGI) has been studied and compared with that in the parental line, which is PGI+. Both cell lines could be maintained for 24 hr in hexose media devoid of L-glutamine. Under these conditions in mannose medium, both the parental line and the PGI mutant maintained high intracellular ATP levels.

Hexose uptake regulation mediated through aerobic pathways: schism in a fibroblast mutant.

A protracted type of down-regulation of the hexose transport system in cultured fibroblasts that depends on one main factor in their nutritional state, the presence or absence of metabolizable D- aldohexoses in the culture fluid, is discussed. Fructose feeding is unable to elicit a down-regulation, whereas mannose and D-glucosamine, regulation of the transport system. This down-regulation or transport curb depends on oxidative energy metabolism, because inhibitors of this type of metabolism bring about a striking release of the transport curb.

Further clues concerning the vectors essential to regulation of hexose transport, as studied in fibroblast cultures from a metabolic mutant.

A close study of the metabolic regulation of hexose transport in a hamster fibroblast mutant, highly defective in the enzyme phosphoglucose isomerase (PGI mutant), reveals the requirement for at least three vectors for transport regulation. The downward regulation of the hexose transport system, called the "transport curb," requires (i) a ligand for the transport system, (ii) oxidative energy metabolism, and (iii) some specific enzymes of the glucose-6-phosphate metabolism. Deprivation of glucose was shown to deprive the PGI mutant of UDP hexose, whereas the glucose-fed mutant contained high levels.

Down-regulation of the hexose transport system: metabolic basis studied with a fibroblast mutant lacking phosphoglucose isomerase.

Down-regulation ("curb") of hexose transport in Chinese hamster lung fibroblasts has been studied in a metabolic mutant highly defective in phosphoglucose isomerase (PGI; glucosephosphate isomerase; D-glucose-6-phosphate ketol-isomerase, EC 5.3.1.

Effects of combined glutamine and serum deprivation on glucose control of hexose transport in mammalian fibroblast cultures.

Regulation of hexose transport in NIL hamster fibroblasts has been studied in confluent cultures preconditioned for 24 hr in media deprived of glutamine or of serum or of both. Cultures maintained in media containing dialyzed fetal calf serum and 4 mM glutamine accumulated up to 72 nmol of glutamine per mg of cell protein; in contrast, cells deprived of glutamine contained less than 1 nmol/mg of cell protein. Glutamine elicited a general enhancement of hexose transport compared with transport in glutamine-deprived cultures.

Uncouplers of oxidative phosphorylation promote derepression of the hexose transport system in cultures of hamster cells.

Long-term (18-24 hr) preincubation of NIL hamster cell cultures with D-glucose or D-glucosamine (both of which repress the hexose transport system) gave rise to a striking loss of the hexose transport system ("super-repression") when cycloheximide was also present in the culture medium. However, if 0.2 mM 2,4-dinitrophenol (DNP) was also present, the cycloheximide-mediated super-repression was prevented.

Selective high metabolic lability of uridine, guanosine and cytosine triphosphates in response to glucose deprivation and refeeding of untransformed and polyoma virus-transformed hamster fibroblasts.

Sugar deprivation of hamster fibroblasts (NIL) affected the steady state levels (pool sizes) of cellular acid soluble nucleotides in the folloing fashion: the pools of UTP, GTP and CTP decreased to a much greater extent than the cellular ATP pools, with the UTP pools undergoing the most dramatic reduction. Sugar deprivation of polyoma-transformed NIL cells (PyNIL) yielded even sharper decreases in the nucleoside triphosphate pools with relative changes similar to those of the untransformed cells. Inhibition of protein synthesis by cycloheximide, initiated at the onset of (and continued during) sugar deprivation, prevented the reduction in pool sizes and yielded values slightly higher than those observed for pool sizes in cells cultured in sugar-supplemented medium.

Catabolic control of the enhanced alanine-preferring system for amino acid transport in glucose-starved hamster cells requires protein synthesis.

In cultured hamster cells starved for glucose for 24 hr there is an enhancement of the rate of alpha-aminoisobutyric acid transport ("shiftup"). When the starved cells are re-fed with glucose, the rate of transport shifts back down to the low, "regulated" rate typical of cells continuously fed with medium containing glucose ("shiftdown"). The high, deregulated rate of transport is maintained, however, when cycloheximide is present for 24 hr during the re-feeding with glucose.

Comparative studies of glucose-fed and glucose-starved hamster cell cultures: responses in galactose metabolism.

The metabolic flow of trace amounts of D-[14C]-galactose was followed in cultures of transformed and untransformed hamster cells over a period ranging from five minutes to two hours. The results of chromatographic and enzymatic analyses of the soluble pools are described. Non-glycolytic cells(previously deprived of sugar periods of up to 24 hours) convert D-galactose to galactose-1-phosphate and uridine diphosphoglucuronic acid in 10 to 20 minutes.