Control of receptor-induced signaling complex formation by the kinetics of ligand/receptor interaction

Tumor necrosis factor (TNF) exists both as a membrane-integrated type II precursor protein and a soluble cytokine that have different bioactivities on TNFR2 (CD120b) but not on TNFR1 (CD120a). To identify the molecular basis of this disparity, we have investigated receptor chimeras comprising the cytoplasmic part of Fas (CD95) and the extracellular domains of the two TNF receptors. The membrane form of TNF, but not its soluble form, was capable of inducing apoptosis as well as activation of c-Jun N-terminal kinase and NF-kappaB via the TNFR2-derived chimera. In contrast, the TNFR1-Fas chimera displayed strong responsiveness to both TNF forms. This pattern of responsiveness is identical to that of wild type TNF receptors, demonstrating that the underlying mechanisms are independent of the particular type of the intracellular signaling machinery and rather are controlled upstream of the intracellular domain. We further demonstrate that the signaling strength induced by a given ligand/receptor interaction is regulated at the level of adaptor protein recruitment, as shown for FADD, caspase-8, and TRAF2. Since both incidents, strong signaling and robust adapter protein recruitment, are paralleled by a high stability of individual ligand-receptor complexes, we propose that half-lives of individual ligand-receptor complexes control signaling at the level of adaptor protein recruitment.     

Reinvestigation of the <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> genome annotation by comparison to the genome of a related fungus: <Emphasis Type="Italic">Ashbya gossypii</Emphasis>

Background  

The recently sequenced genome of the filamentous fungus Ashbya gossypii revealed remarkable similarities to that of the budding yeast Saccharomyces cerevisiae both at the level of homology and synteny (conservation of gene order). Thus, it became possible to reinvestigate the S. cerevisiae genome in the syntenic regions leading to an improved annotation.     

Reduced-folate carrier (RFC) is expressed in placenta and yolk sac,as well as in cells of the developing forebrain,hindbrain, neural tube,craniofacial region,eye, limb buds and heart

Background  

Folate is essential for cellular proliferation and tissue regeneration. As mammalian cells cannot synthesize folates de novo, tightly regulated cellular uptake processes have evolved to sustain sufficient levels of intracellular tetrahydrofolate cofactors to support biosynthesis of purines, pyrimidines, and some amino acids (serine, methionine). Though reduced-folate carrier (RFC) is one of the major proteins mediating folate transport, knowledge of the developmental expression of RFC is lacking. We utilized in situ hybridization and immunolocalization to determine the developmental distribution of RFC message and protein, respectively.     

Comments on the sixth report of the Animal Welfare Advisory Committee

The Animal Welfare Advisory Committee (AWAC) was established in July 1996, to consider the care, welfare and use of animals involved in procedures for defence research purposes at Defence and Evaluation Research Agency (DERA) establishments in the UK. Two of the objectives of AWAC are to examine the broad trends in animal use at DERA establishments, and to implement and audit the application of the Three Rs principle. AWAC's sixth report addressed the period from 31 October 2000 to 28 February 2002. The statistics of animal use within the report are briefly examined, and some of the actions undertaken by defence research establishments to facilitate the application of the Three Rs are highlighted. It is recommended that, if possible (subject to security constraints), figures detailing the severity of the procedures undertaken should be included in future issues of the report, in order to provide a more-detailed account. It is concluded that Defence Science and Technology Laboratory establishments have made a contribution to the Three Rs, and that other establishments may be able to incorporate some of their actions into their own research programmes. There was an overall 36% increase in the number of procedures carried out by defence research establishments between 1995 and 2000, from 8,900 to 12,065. This probably reflects alterations in the research programme, which is, in turn, decided primarily by the Ministry of Defence's customers and the progress made with previous research programmes. It is therefore recommended that the UK Government allocates significantly more financial resources for the development and validation of alternatives, in order to maximise the potential for achieving the Three Rs in defence research, and to complement the existing initiatives within the defence research industry.     

Increased salt sensitivity secondary to leptin resistance in SHHF rats is mediated by endothelin

A link between leptin resistance, obesity, and salt sensitivity has been suggested. SHHF/Mcc-fa ^cp rats (SHHF) were used to study the effect of gene dosage of a null mutation of the leptin receptor (cp) on salt sensitivity and response to a combined endothelin A and B receptor antagonist (bosentan). Obese (cp/cp), heterozygous (+/cp), and homozygous lean (+/+) male SHHF were fed a low salt diet (0.3% NaCl) for 7 days, followed by a high salt diet (8.0% NaCl) for 7 days. There were no significant differences in systolic blood pressure between genotypes on low salt. In response to high salt, cp/cp had significantly greater systolic pressure than +/cp and +/+. On high salt diet, cp/cp showed a significant increase in 24 h urinary endothelin excretion and increased renal expression of preproendothelin mRNA. There was no effect of high salt diet on renal excretion of nitric oxide (NOx) or on gene expression of endothelial, neuronal, or cytokine-induced nitric oxide synthase isoforms (eNOS, nNOS, iNOS, respectively). Treatment with bosentan prevented the high salt-induced increment in systolic blood pressure in cp/cp. This was associated with a doubling of renal NOx excretion, but without changes in eNOS, nNOS, or iNOS expression. Endothelin receptor antagonism did not normalize systolic pressure in any of the genotypes. Our studies indicate that obesity secondary to leptin resistance (cp/cp) results in increased salt sensitivity that is mediated by endothelin in the SHHF rat.     

Regulation of taurine transporter expression by NO in cultured human retinal pigment epithelial cells

Taurine is activelytransported at the retinal pigment epithelial (RPE) apical membrane inan Na⁺- and Cl-dependent manner. Diabetes mayalter the function of the taurine transporter. Because nitric oxide(NO) is a molecule implicated in the pathogenesis of diabetes, we askedwhether NO would alter the activity of the taurine transporter incultured ARPE-19 cells. The activity of the transporter was stimulatedin the presence of the NO donor 3-morpholinosydnonimine. Thestimulatory effects of 3-morpholinosydnonimine were not observed duringthe initial 16-h treatment; however, stimulation of taurine uptake waselevated dramatically above control values with 20- and 24-htreatments. Kinetic analysis revealed that the stimulation wasassociated with an increase in the maximal velocity of the transporterwith no significant change in the substrate affinity. The NO-induced increase in taurine uptake was inhibited by actinomycin D and cycloheximide. RT-PCR analysis and nuclear run-on assays provided evidence for upregulation of the transporter gene. This study providesthe first evidence of an increase in taurine transporter geneexpression in human RPE cells cultured under conditions of elevatedlevels of NO.