A distinctive RNA fold: the solution structure of an analogue of the yeast tRNAPhe T Psi C domain.

The structure of an analogue of the yeast tRNAPhe T Psi C stem-loop has been determined by NMR spectroscopy and restrained molecular dynamics. The molecule contained the highly conserved modification ribothymidine at its naturally occurring position. The ribothymidine-modified T Psi C stem-loop is the product of the m5U54-tRNA methyltransferase, but is not a substrate for the m1A58-tRNA methyltransferase. Site-specific substitutions and 15N labels were used to confirm the assignment of NOESY cross-peaks critical in defining the global fold of the molecule. The structure is unusual in that the loop folds far over into the major groove of the curved stem. This conformation is stabilized by both stacking interactions and hydrogen bond formation. Furthermore, this conformation appears to be unique among RNA hairpins of similar size. There is, however, a considerable resemblance to the analogous domain in the crystal structure of the full-length yeast tRNAPhe. We believe, therefore, that the structure we have determined may represent an intermediate in the folding pathway during the maturation of tRNA.     

Modelling changes in VOC emission in response to climate change in the continental United States

The alteration of climate is driven not only by anthropogenic activities, but also by biosphere processes that change in conjunction with climate. Emission of volatile organic compounds (VOCs) from vegetation may be particularly sensitive to changes in climate and may play an important role in climate forcing through their influence on the atmospheric oxidative balance, greenhouse gas concentration, and the formation of aerosols. Using the VEMAP vegetation database and associated vegetation responses to climate change, this study examined the independent and combined effects of simulated changes in temperature, CO₂ concentration, and vegetation distribution on annual emissions of isoprene, monoterpenes, and other reactive VOCs (ORVOCs) from potential vegetation of the continental United States. Temperature effects were modelled according to the direct influence of temperature on enzymatic isoprene production and the vapour pressure of monoterpenes and ORVOCs. The effect of elevated CO₂ concentration was modelled according to increases in foliar biomass per unit of emitting surface area. The effects of vegetation distribution reflects simulated changes in species spatial distribution and areal coverage by 21 different vegetation classes. Simulated climate warming associated with a doubled atmospheric CO₂ concentration enhanced total modelled VOC emission by 81.8% (isoprene + 82.1%, monoterpenes + 81.6%, ORVOC + 81.1%), whereas a simulated doubled CO₂ alone enhanced total modelled VOC emission by only + 11.8% (isoprene + 13.7%, monoterpenes + 4.1%, ORVOC + 11.7%). A simulated redistribution of vegetation in response to altered temperatures and precipitation patterns caused total modelled VOC emission to decline by 10.4% (isoprene – 11.7%, monoterpenes – 18.6%, ORVOC 0.0%) driven by a decline in area covered by vegetation classes emitting VOCs at high rates. Thus, the positive effect of leaf-level adjustments to elevated CO₂ (i.e. increases in foliar biomass) is balanced by the negative effect of ecosystem-level adjustments to climate (i.e. decreases in areal coverage of species emitting VOC at high rates).     

The effects of infant births on the sociosexual behavior and hormonal patterns of a cooperatively breeding primate (Cebuella pygmaea)

This study examined changes in the behavioral and hormonal patterns of cooperatively breeding pairs in a primate species with the passing of time and with specific reproductive events. We (1) compared patterns of sexual, agonistic, and affiliative behavior of newly paired pygmy marmosets with the same behavioral patterns immediately after the birth of their first set of infants; (2) determined if postpartum behavioral differences existed between pairs whose infants lived and those whose infants died; and (3) examined whether behavioral patterns changed over the course of ovarian cycles in parous pygmy marmosets as had been documented in nulliparous pairs. The behavior of pairs was recorded during daily half-hour focal samples for 60 days after pairing, and 30 or 60 days after the birth of infants for pairs whose litters died or lived, respectively. Daily urine samples from females during the study were analyzed for luteinizing hormone and pregnanediol glucuronide concentrations to determine dates of ovulation. The results indicated that males consistently altered their sexual behavior and olfactory monitoring of mates during periovulatory periods in the females' cycles both postpairing and postpartum, while similar rates of social and sexual behavior were maintained between the conditions. Sexual behavior occurred throughout the females' ovarian cycles. Peaks in sexual behavior during the periovulatory period in nulliparous pairs disappeared after the birth of infants. Pairs whose infants died showed higher rates of sexual behavior than pairs with surviving infants. Social and sexual behavior may function to maintain the relationship both during and outside of ovulation, especially with the loss of infants. © 1996 Wiley-Liss, Inc.     

Close and distant: Contrasting the metabolism of two closely related subspecies of Scots pine under the effects of folivory and summer drought

Metabolomes, as chemical phenotypes of organisms, are likely not only shaped by the environment but also by common ancestry. If this is the case, we expect that closely related species of pines will tend to reach similar metabolomic solutions to the same environmental stressors. We examined the metabolomes of two sympatric subspecies of Pinus sylvestris in Sierra Nevada (southern Iberian Peninsula), in summer and winter and exposed to folivory by the pine processionary moth. The overall metabolomes differed between the subspecies but both tended to respond more similarly to folivory. The metabolomes of the subspecies were more dissimilar in summer than in winter, and iberica trees had higher concentrations of metabolites directly related to drought stress. Our results are consistent with the notion that certain plant metabolic responses associated with folivory have been phylogenetically conserved. The larger divergence between subspecies metabolomes in summer is likely due to the warmer and drier conditions that the northern iberica subspecies experience in Sierra Nevada. Our results provide crucial insights into how iberica populations would respond to the predicted conditions of climate change under an increased defoliation in the Mediterranean Basin.     

The adipose triglyceride lipase,adiponectin and visfatin are downregulated by tumor necrosis factor-α (TNF-α) in vivo

Inflammatory cytokines have been linked to obesity-related insulin resistance. To investigate the effect of TNF-α, an inflammatory cytokine, on insulin action, C57BL/6J mice were treated with TNF-α for 7 days after which we examined the in vivo effects of TNF-α on glucose tolerance and insulin sensitivity with IV glucose tolerance tests and hyperinsulinemic-euglycemic clamps. In addition, we analyzed the in vivo effect of TNF-α on several metabolism-related genes and adipocytokines implicated in the development of insulin resistance. TNF-α treatment resulted in markedly increased fasting blood glucose, insulin and free fatty acids (FFA) levels and reduced glucose tolerance. During the clamps, the rates insulin-stimulated whole body (G_Rd) and skeletal muscle glucose uptake (MGU) and insulin’s ability to suppress hepatic glucose production (HGP) were decreased in TNF-α treated animals, indicating insulin resistance. In addition, both PPARγ and ATGL mRNA expression in adipose tissues as well as ATGL protein levels in plasma were downregulated. Moreover, adipose mRNA expression and plasma protein levels of adiponectin and visfatin were significantly down-regulated. We conclude that the alterations of PPARγ, ATGL, adiponectin and visfatin may contribute to the development of insulin resistance mediated by TNF-α.     

The relationships of cholesterol metabolism and plasma plant sterols with the severity of coronary artery disease

Changes in the balance of cholesterol absorption and synthesis and moderately elevated plasma plant sterols have been suggested to be atherogenic. Measuring cholestanol, lathosterol, campesterol, and sitosterol, we investigated the relationships of cholesterol metabolism and plasma plant sterols with the severity of coronary artery disease (CAD) in 2,440 participants of the Ludwigshafen Risk and Cardiovascular health (LURIC) study. The coronary status was determined by angiography, and the severity of CAD was assessed by the Friesinger Score (FS). An increase in the ratio of cholestanol to cholesterol was associated with high FS (P = 0.006). In contrast, a high ratio of lathosterol to cholesterol went in parallel with low FS (P < 0.001). Whereas the campesterol to cholesterol ratio significantly correlated with the FS (P = 0.026), the relationship of the sitosterol to cholesterol ratio with the FS did not reach statistical significance in the whole group. Increased campesterol, sitosterol, and cholestanol to lathosterol ratios were associated high FS (P < 0.001). To conclude, there is a modest association of high cholesterol absorption and low cholesterol synthesis with an increased severity of CAD. An atherogenic role of plasma plant sterols themselves, however, seems unlikely in subjects without sitosterolaemia.     

Identification of phosphatidylserine decarboxylases 1 and 2 from Pichia pastoris

Genetic manipulation of lipid biosynthetic enzymes allows modification of cellular membranes. We made use of this strategy and constructed mutants in phospholipid metabolism of Pichia pastoris , which is widely used in biotechnology for expression of heterologous proteins. Here we describe identification of two P. pastoris phosphatidylserine decarboxylases (PSDs) encoded by genes homologous to PSD1 and PSD2 from Saccharomyces cerevisiae . Using P. pastoris psd1 Δ and psd2 Δ mutants we investigated the contribution of the respective gene products to phosphatidylethanolamine synthesis, membrane composition and cell growth. Deletion of PSD1 caused loss of PSD activity in mitochondria, a severe growth defect on minimal media and depletion of cellular and mitochondrial phosphatidylethanolamine levels. This defect could not be compensated by Psd2p, but by supplementation with ethanolamine, which is the substrate for the cytidine diphosphate (CDP)–ethanolamine pathway, the third route of phosphatidylethanolamine synthesis in yeast. Fatty acid analysis showed selectivity of both Psd1p and Psd2p in vivo for the synthesis of unsaturated phosphatidylethanolamine species. Phosphatidylethanolamine species containing palmitic acid (16:0), however, were preferentially assembled into mitochondria. In summary, this study provides first insight into membrane manipulation of P. pastoris , which may serve as a useful method to modify cell biological properties of this microorganism for biotechnological purposes.     

Enhancement of Tumour-Specific Immune Responses In Vivo by ‘MHC Loading-Enhancer’ (MLE)

Background

Class II MHC molecules (MHC II) are cell surface receptors displaying short protein fragments for the surveillance by CD4+ T cells. Antigens therefore have to be loaded onto this receptor in order to induce productive immune responses. On the cell surface, most MHC II molecules are either occupied by ligands or their binding cleft has been blocked by the acquisition of a non-receptive state. Direct loading with antigens, as required during peptide vaccinations, is therefore hindered.

Principal Findings

Here we show, that the in vivo response of CD4+ T cells can be improved, when the antigens are administered together with ‘MHC-loading enhancer’ (MLE). MLE are small catalytic compounds able to open up the MHC binding site by triggering ligand-release and stabilizing the receptive state. Their enhancing effect on the immune response was demonstrated here with an antigen from the influenza virus and tumour associated antigens (TAA) derived from the NY-ESO-1 protein. The application of these antigens in combination with adamantane ethanol (AdEtOH), an MLE compound active on human HLA-DR molecules, significantly increased the frequency of antigen-specific CD4+ T cells in mice transgenic for the human MHC II molecule. Notably, the effect was evident only with the MLE-susceptible HLA-DR molecule and not with murine MHC II molecules non-susceptible for the catalytic effect of the MLE.

Conclusion

MLE can specifically increase the potency of a vaccine by facilitating the efficient transfer of the antigen onto the MHC molecule. They may therefore open a new way to improve vaccination efficacy and tumour-immunotherapy.