Hemopoietic cell transformation is associated with failure to downregulate glucose uptake during the G2/M phase of the cell cycle

Growth factors and cytokines initiate multiple signal transduction pathways that lead to cell survival, cell cycle progression or differentiation. A common feature of these pathways is increased cellular metabolism and glucose uptake. Furthermore, the energy requirements of many cancers and transformed cell lines are met by constitutive upregulation of glucose uptake. Relationships among transforming events, glucose uptake and cell cycle progression are not well understood. Here we investigated the regulation of glucose transport during the cell cycle of growth factor-dependent 32D cells, primary T-cells, src-transformed 32D cells and Jurkat cells. Cells were enriched in the G1, S and G2/M phases of the cell cycle, and glucose transporter expression and 2-deoxyglucose uptake were measured. Glucose transporter expression increased with cell volume as cells progressed through the cell cycle. Growth factor-dependent 32D cells and T-lymphocytes were characterised by increased 2-deoxyglucose uptake from G1 to S and reduced uptake at G2/M, with the highest specific activity of transporters in the S phase. In contrast, src-transformed 32D cells and Jurkat cells showed increased 2-deoxyglucose uptake from S to G2/M, with the highest glucose transporter specific activity in G2/M. Our results show that glucose transport is regulated in a cell cycle-dependent manner and suggest that this regulation may be altered in transformed cells.     

Cell surface oxygen consumption by mitochondrial gene knockout cells

Mitochondrial gene knockout (rho(0)) cells that depend on glycolysis for their energy requirements show an increased ability to reduce cell-impermeable tetrazolium dyes by electron transport across the plasma membrane. In this report, we show for the first time, that oxygen functions as a terminal electron acceptor for trans-plasma membrane electron transport (tPMET) in HL60rho(0) cells, and that this cell surface oxygen consumption is associated with oxygen-dependent cell growth in the absence of mitochondrial electron transport function. Non-mitochondrial oxygen consumption by HL60rho(0) cells was extensively inhibited by extracellular NADH and NADPH, but not by NAD(+), localizing this process at the cell surface. Mitochondrial electron transport inhibitors and the uncoupler, FCCP, did not affect oxygen consumption by HL60rho(0) cells. Inhibitors of glucose uptake and glycolysis, the ubiquinone redox cycle inhibitors, capsaicin and resiniferatoxin, the flavin centre inhibitor, diphenyleneiodonium, and the NQO1 inhibitor, dicoumarol, all inhibited oxygen consumption by HL60rho(0) cells. Similarities in inhibition profiles between non-mitochondrial oxygen consumption and reduction of the cell-impermeable tetrazolium dye, WST-1, suggest that both systems may share a common tPMET pathway. This is supported by the finding that terminal electron acceptors from both pathways compete for electrons from intracellular NADH.     

Dopamine D1 activation shortens the duration of phases in stereotyped grooming sequences

Rats frequently emit grooming actions in a highly stereotyped, syntactic chain in which three distinct phases of facially directed forearm movements are sequentially emitted in a rule-governed pattern and followed by body-directed licking. The present study evaluated the effects of the full dopamine D1 agonist, SKF 81297, and the partial dopamine D1 agonist, SKF 38393, on the duration of individual phases of stereotyped grooming chains. We found that systemic administration of SKF 81297 significantly shortened grooming chain duration. An examination of the fine temporal structure of syntactic grooming chain actions showed that duration changes were correlated with decreased numbers of actions in phases I and IV of the chain. Phases II and III were not changed in duration, although there were some structural distortions introduced. The partial D1 agonist, SKF 38393, had no effect on duration or number of component actions in the grooming chain. Based on these results, we hypothesize that the timing of syntactic grooming phase transitions may involve a D1-mediated internal clock process that is altered by full D1 agonist activation. By this model, SKF 81297 increases the speed of the clock used for the temporal control of grooming actions, and thus shortens phase durations.     

Robust Modeling of Differential Gene Expression Data Using Normal/Independent Distributions: A Bayesian Approach

In this paper, the problem of identifying differentially expressed genes under different conditions using gene expression microarray data, in the presence of outliers, is discussed. For this purpose, the robust modeling of gene expression data using some powerful distributions known as normal/independent distributions is considered. These distributions include the Student’s t and normal distributions which have been used previously, but also include extensions such as the slash, the contaminated normal and the Laplace distributions. The purpose of this paper is to identify differentially expressed genes by considering these distributional assumptions instead of the normal distribution. A Bayesian approach using the Markov Chain Monte Carlo method is adopted for parameter estimation. Two publicly available gene expression data sets are analyzed using the proposed approach. The use of the robust models for detecting differentially expressed genes is investigated. This investigation shows that the choice of model for differentiating gene expression data is very important. This is due to the small number of replicates for each gene and the existence of outlying data. Comparison of the performance of these models is made using different statistical criteria and the ROC curve. The method is illustrated using some simulation studies. We demonstrate the flexibility of these robust models in identifying differentially expressed genes.     

Caffeine inhibits inositol-trisphosphate-induced membrane potential oscillations in Xenopus oocytes.

Immature Xenopus oocytes injected with inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) give a complex electrophysiological response comprising an a early depolarizing spike followed by a burst of oscillations. These two components have been interpreted on the basis of an interaction between two internal calcium stores: an Ins(1,4,5) P3-sensitive pool responsible for the early spike which then primes an Ins(1,4,5) P3-insensitive pool to begin to oscillate through a process of calcium-induced calcium release (Berridge, M. J., J. Physiol., Lond. 403, 589-599 (1988)). The role of the latter was investigated in Xenopus oocytes by using the drug caffeine which can trigger calcium-induced calcium release in muscle cells. Caffeine had no effect on the early Ins(1,4,5)P3-induced spike but it suppressed the subsequent oscillations. The spontaneous oscillations observed in some oocytes were also abolished by caffeine. Oscillation amplitude and duration was slightly reduced following incubation of oocytes with adenosine or isobutylmethylxanthine. Because these two agents gave large membrane hyperpolarizations indicative of an increase in cyclic AMP, it can be concluded that this second messenger is not responsible for the inhibitory action of caffeine. The ability of caffeine to abolish oscillations while not affecting the early Ins(1,4,5) P3 response is discussed with regard to the two-pool model for generating calcium oscillations.     

The caffeine-sensitive Ca2+ store in bovine adrenal chromaffin cells; an examination of its role in triggering secretion and Ca2+ homeostasis

The effect of caffeine on catecholamine secretion and intracellular free Ca2+ concentration [( Ca2+]i) in bovine adrenal chromaffin cells was examined using single fura-2-loaded cells and cell populations. In cell populations caffeine elicited a large (approximately 200 nM) transient rise in [Ca2+]i that was independent of external Ca2+. This rise in [Ca2+]i triggered little secretion. Single cell measurements of [Ca2+]i showed that most cells responded with a large (greater than 200 nM) rise in [Ca2+]i, whereas a minority failed to respond. The latter, whose caffeine-sensitive store was empty, buffered a Ca2+ load induced by a depolarizing stimulus more effectively than those whose store was full. The caffeine-sensitive store in bovine chromaffin cells may be involved in Ca2+ homeostasis rather than in triggering exocytosis.