The effects of a number of metabolic inhibitors on the influx of Ca2+ activated by stimulation of receptors coupled to inositol 1,4,5-trisphosphate generation or by depletion of intracellular Ca2+ stores with thapsigargin were investigated in four different cell types: Ehrlich ascites tumor cells, Jurkat and HeLa cell lines, and rat hepatocytes. Independently of their chemical structure and site of inhibition, all of these metabolic poisons markedly inhibited Ca2+ influx without significantly affecting Ca2+ release. This inhibition was not due to membrane potential depolarization or to alteration in cytosolic pH but appeared correlated to a drop in the cellular concentration of ATP. The decreases in cellular [ATP] were paralleled by decreases in [GTP] and by increases in [ADP] and [GDP]. The reduction in ATP level necessary to drastically reduce Ca2+ influx was quite small, e.g. a 50% inhibition for a 5% reduction in [ATP], thus within the range of fluctuation presumably occurring under physiological conditions. We suggest that changes in the adenine or guanine nucleotide concentrations may represent an important modulatory mechanism of Ca2+ influx activated by store depletion.

The constitutive/inducible association of the T cell receptor (TCR) with isolated detergent-resistant, lipid raft-derived membranes has been studied in Jurkat T lymphocytes. Membranes resistant to 1% Triton X-100 contained virtually no CD3ε, part of the TCR complex, irrespective of cell stimulation. On the other hand, membranes resistant either to a lower Triton X-100 concentration (i.e. 0.2%) or to the less hydrophobic detergent Brij 58 (1%) contained (i) a low CD3ε amount (approximate 2.7% of total) in resting cells and (ii) a several times higher amount of the TCR component, after T cell stimulation with either antigen-presenting cells or with phytohemagglutinin. It appeared that CD3/TCR was constitutively associated with and recruited to a raft-derived membrane subset because (i) all three membrane preparations contained a similar amount of the raft marker tyrosine kinase Lck but no detectable amounts of the conventional membrane markers, CD45 phosphatase and transferrin receptor; (ii) a larger amount of particulate membranes were resistant to solubilization with 0.2% Triton X-100 and Brij 58 than to solubilization with 1% Triton X-100; and (iii) higher cholesterol levels were present in membranes resistant to either the lower Triton X-100 concentration or to Brij 58, as compared with those resistant to 1% Triton X-100. The recruitment of CD3 to the raft-derived membrane subset appeared (i) to occur independently of cell signaling events, such as protein-tyrosine phosphorylation and Ca²⁺mobilization/influx, and (ii) to be associated with clustering of plasma membrane rafts induced by multiple cross-linking of either TCR or the raft component, ganglioside GM₁. We suggest that during T cell stimulation a lateral reorganization of rafts into polarized larger domains can determine the recruitment of TCR into these domains, which favors a polarization of the signaling cascade.

ShcA is an important mediator of Ras/MAPK activation in PTK-regulated pathways triggered by surface receptors. This function is subserved by the constitutively expressed p52-kDa isoform. Besides activating Ras, p52Shc couples the TCR to Rho GTPases, and thereby participates in actin cytoskeleton remodeling in T cells. Here we have addressed the potential involvement of p52Shc in T-cell chemotaxis and the role of the phosphorylatable tyrosine residues, YY239/240 and Y317, in this process. We show that CXCR4 engagement by the homeostatic chemokine, SDF-1α, results in p52Shc phosphorylation and its assembly into a complex that includes Lck, ZAP-70, and Vav. This process was found to be both Lck and Gi dependent. Expression of p52Shc mutants lacking YY239/240 or Y317, or p52Shc deficiency, resulted in a profound impairment in CXCR4 signaling and SDF-1α–dependent chemotaxis, underscoring a crucial role of p52Shc as an early component of the CXCR4 signaling cascade. p52Shc was also found to be required for ligand-dependent CXCR4 internalization independently of tyrosine phosphorylation. Remarkably, CXCR4 engagement promoted phosphorylation of the ζ chain of the TCR/CD3 complex, which was found to be essential for CXCR4 signaling, as well as for SDF-1α–dependent receptor endocytosis and chemotaxis, indicating that CXCR4 signals by transactivating the TCR.

The modern Western society lifestyle is characterized by a hyperenergetic, high sugar containing food intake. Sugar intake increased dramatically during the last few decades, due to the excessive consumption of high-sugar drinks and high-fructose corn syrup. Current evidence suggests that high fructose intake when combined with overeating and adiposity promotes adverse metabolic health effects including dyslipidemia, insulin resistance, type II diabetes, and inflammation. Similarly, elevated glucocorticoid levels, especially the enhanced generation of active glucocorticoids in the adipose tissue due to increased 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1) activity, have been associated with metabolic diseases. Moreover, recent evidence suggests that fructose stimulates the 11β-HSD1-mediated glucocorticoid activation by enhancing the availability of its cofactor NADPH. In adipocytes, fructose was found to stimulate 11β-HSD1 expression and activity, thereby promoting the adipogenic effects of glucocorticoids. This article aims to highlight the interconnections between overwhelmed fructose metabolism, intracellular glucocorticoid activation in adipose tissue, and their metabolic effects on the progression of the metabolic syndrome.