Transcriptional regulation of murine NADP+-dependent methylenetetrahydrofolate dehydrogenase-cyclohydrolase-synthetase

AbstractThe cytosolic NADP+-dependent methylenetetrahydrofolate dehydrogenase-methenyltetrahydrofolate cyclohydrolase-formyltetrahydrofolate synthetase is ubiquitously expressed in all mouse tissues and cell lines examined. Northern analyses of the RNA indicated that there is an extensive variation in the levels of mRNA in different tissues. However, the gene is refractory to induction by serum, phorbol esters or growth factors in cultured fibroblasts. The mRNA of the NADP′-dependent trifunctional enzyme is stabilized post-transcriptionally by insulin-like growth factor-1

Evidence for the phosphorylation of enzyme IIglucose of the phosphoenolpyruvate—sugar phosphotransferase system of Escherichia coli and Salmonella typhimurium

AbstractA membrane-bound phosphoprotein, of subunit Mr 48 000 by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, has been found in Escherichia coli and Salmonella typhimurium. The phosphorylation of this protein is dependent upon phosphoenolpyruvate and the protein of the sugar phosphotransferase (PT) system: enzyme I, HPr, and IIIglucose. The membrane-bound phosphoprotein is identified as enzyme IIglucose. Other membrane-bound phosphoproteins, that are also dependent upon the PT system, remain to be identified

Nuclear localization of prostaglandin E(2) receptors

Prostaglandin E(2) receptors (EP) were detected by radioligand binding in nuclear fractions isolated from porcine brain and myometrium. Intracellular localization by immunocytofluorescence revealed perinuclear localization of EPs in porcine cerebral microvascular endothelial cells. Nuclear association of EP(1) was also found in fibroblast Swiss 3T3 cells stably overexpressing EP(1) and in human embryonic kidney 293 (Epstein–Barr virus-encoded nuclear antigen) cells expressing EP(1) fused to green fluorescent protein. High-resolution immunostaining of EP(1) revealed their presence in the nuclear envelope of isolated (cultured) endothelial cells and in situ in brain (cortex) endothelial cells and neurons. Stimulation of these nuclear receptors modulate nuclear calcium and gene transcription

Plateletactivating factor in vasoobliteration of oxygen-induced retinopathy. Invest Ophthalmol Vis Sci 43

PURPOSE. To test whether platelet-activating factor (PAF) directly causes retinovascular endothelial cell (EC) death. METHODS. Retinovascular density was calculated in rat pups exposed to 80% O 2 from postnatal days (P)6 to P14 (to produce oxygen-induced retinopathy [OIR]), using the adenosine diphosphatase (ADPase) technique, in animals treated with distinct PAF receptor blockers (PCA-4248, BN52021, or THG315). PAF levels were then measured in the retinas. Viability of ECs from piglets and humans in response to C-PAF (a stable PAF analogue) was determined by the reduction of the tetrazolium salt 3-(4,5-dimethyl thiazol-2yl)-2,5-diphenyl tetrazolium bromide (MTT) by viable cells, incorporation of propidium iodide (PI), TUNEL assay, and release of lactate dehydrogenase. Release of thromboxane (TX) was measured in the cell media. RESULTS. PAF levels in retina were markedly increased by exposure of isolated rat retinas to H 2 O 2 (1 M) and of rat pups placed in 80% O 2 . Exposure to 80% O 2 induced retinal vasoobliteration, which was equally significantly inhibited (ϳ60%) by all PAF receptor blockers tested. C-PAF increased incorporation of PI by isolated rat retinal microvasculature. Also, C-PAF caused time-and concentration-dependent death of cultured retinal ECs, which was prevented by the PAF receptor antagonist CV-3988. This effect of C-PAF was selective on retinal and neurovascular ECs, but not on other ECs. DNA fragmentation (TUNEL) was hardly detected, and inhibition of apoptosis-related processes by nicotinamide, cyclosporin A, and Z-DEVD-FMK and Z-VAD-FMK (caspase inhibitors) barely protected against death in EC, whereas C-PAF increased release of lactate dehydrogenase, implying that necrosis is the nature of EC death. Finally, C-PAF-induced cell death was preceded by an increase in TXB 2 levels and was prevented by TXA 2 synthase inhibition (with CGS12970). CONCLUSIONS. The data suggest PAF plays a major role in vasoobliteration in OIR by triggering death of neuroretinal microvascular ECs. The cell death seems to be mediated at least in part by TXA 2 . These effects of PAF may participate in ischemic retinopathies such as diabetes and retinopathy of prematurity. (Invest Ophthalmol Vis Sci. 2002;43:3327-3337