The prion protein binds thiamine

Although highly conserved throughout evolution, the exact biological function of the prion protein is still unclear. In an effort to identify the potential biological functions of the prion protein we conducted a small-molecule screening assay using the Syrian hamster prion protein [shPrP(90-232)]. The screen was performed using a library of 149 water-soluble metabolites that are known to pass through the blood-brain barrier. Using a combination of 1D NMR, fluorescence quenching and surface plasmon resonance we identified thiamine (vitamin B1) as a specific prion ligand with a binding constant of ~60 μM. Subsequent studies showed that this interaction is evolutionarily conserved, with similar binding constants being seen for mouse, hamster and human prions. Various protein construct lengths, both with and without the unstructured N-terminal region in the presence and absence of copper, were examined. This indicates that the N-terminus has no influence on the protein's ability to interact with thiamine. In addition to thiamine, the more biologically abundant forms of vitamin B1 (thiamine monophosphate and thiamine diphosphate) were also found to bind the prion protein with similar affinity. Heteronuclear NMR experiments were used to determine thiamine's interaction site, which is located between helix 1 and the preceding loop. These data, in conjunction with computer-aided docking and molecular dynamics, were used to model the thiamine-binding pharmacophore and a comparison with other thiamine binding proteins was performed to reveal the common features of interaction.     

Orchids from coffee-plantations in mexico: an alternative for the sustainable use of tropical ecosystems

Life form, endemism, conservation status, and horticultural interest are detailed for orchid species associated to shade coffee-plantations in Mexico. About 11% of the orchid taxa (214 species) found in these agroecosystems are in the Mexican list of species requiring some form of protection. Almost 40% of the species are of horticultural interest. The importance of promoting shaded coffee plantations as an alternative to the conservation of primary plant communities in Mexican and other tropical regions is clear. Long term management plans are recommended.     

Proteomic analysis of reaper 5' untranslated region-interacting factors isolated by tobramycin affinity-selection reveals a role for La antigen in reaper mRNA translation

Translational control is a key step in gene expression regulation during apoptosis. To understand the mechanisms of mRNA translation of a pro-apoptotic gene, reaper (rpr), we adapted the tobramycin-aptamer technique described by Hartmuth et al. (Proc. Natl. Acad. Sci. USA 2002, 99, 16719-16724) for the analysis of proteins interacting with rpr 5' untranslated region (UTR). We assembled ribonucleoprotein complexes in vitro using translation extracts derived from Drosophila embryos and purified the RNA-protein complexes for mas spectrometry analysis. We identified the proteins bound to the 5' UTR of rpr. One of them, the La antigen, was validated by RNA-crosslinking experiments using recombinant protein and by the translation efficiency of reporter mRNAs in Drosophila cells after RNAinterference experiments. Our data provide evidence of the involvement of La antigen in the translation of rpr and set a protocol for purification of tagged-RNA-protein complexes from cytoplasmic extracts.     

Functional-cranial approach to the influence of economic strategy on skull morphology

Environmental factors are assumed to play an important role in the shaping of craniofacial morphology. Here we propose a statistical approach which can be of utility in estimating the magnitude and localization of a particular nongenetic factor upon the specific functional components of the skull. Our analysis is a combination of previous attempts of apportionment of variance and the application of craniofunctional theory. The effect of subsistence strategy on craniofacial functional components was studied on 18 populations of hunter-gatherers and farmers from South America. Results demonstrate that the environmental factors studied likely influenced the masticatory component's size and shape. Even when this effect is not large enough to clearly differentiate among subsistence strategies (since whole craniofacial variation among populations remains greater), the method used here provides interesting clues to localize plastic or adaptive responses to external stimuli.     

Quantitative analysis of the interaction between the fluorescent probe eosin and the Na+/K+-ATPase studied through Rb+ occlusion

We report a study on the effect of the fluorescent probe eosin on some of the reactions involved in the conformational transitions that lead to the occlusion of the K(+)-congener Rb(+) in the Na(+)/K(+)-ATPase. Eosin decreases the equilibrium levels of occluded Rb(+), this effect being fully attributable to a decrease in the apparent affinity of the enzyme for Rb(+) since the capacity for occlusion remains independent of eosin concentration. The results can be quantitatively described by a model that assumes that two molecules of eosin are able to bind to the Na(+)/K(+)-ATPase, both to the Rb(+)-free and to the Rb(+)-occluded enzyme regardless of the degree of cation occlusion. Concerning the effect on the affinity for Rb(+) occlusion, transient state experiments show that eosin reduces the initial velocity of occlusion, and that, like ATP, it increases the velocity of deocclusion of Rb(+). Interactions between eosin and ATP on Rb(+)-release experiments seem to indicate that eosin binds to the low-affinity site of ATP from which it exerts effects that are similar to those of the nucleotide.     

Intracellular delivery of phosphatidylinositol (3,4,5)-trisphosphate causes incorporation of glucose transporter 4 into the plasma membrane of muscle and fat cells without increasing glucose uptake

Insulin stimulates glucose uptake into muscle and fat cells by translocating glucose transporter 4 (GLUT4) to the cell surface, with input from phosphatidylinositol (PI) 3-kinase and its downstream effector Akt/protein kinase B. Whether PI 3,4,5-trisphosphate (PI(3,4,5)P(3)) suffices to produce GLUT4 translocation is unknown. We used two strategies to deliver PI(3,4,5)P(3) intracellularly and two insulin-sensitive cell lines to examine Akt activation and GLUT4 translocation. In 3T3-L1 adipocytes, the acetoxymethyl ester of PI(3,4,5)P(3) caused GLUT4 migration to the cell periphery and increased the amount of plasma membrane-associated phospho-Akt and GLUT4. Intracellular delivery of PI(3,4,5)P(3) using polyamine carriers also induced translocation of myc-tagged GLUT4 to the surface of intact L6 myoblasts, demonstrating membrane insertion of the transporter. GLUT4 translocation caused by carrier-delivered PI(3,4,5)P(3) was not reproduced by carrier-PI 4,5-bisphosphate or carrier alone. Like insulin, carrier-mediated delivery of PI(3,4,5)P(3) elicited redistribution of perinuclear GLUT4 and Akt phosphorylation at the cell periphery. In contrast to its effect on GLUT4 mobilization, delivered PI(3,4,5)P(3) did not increase 2-deoxyglucose uptake in either L6GLUT4myc myoblasts or 3T3-L1 adipocytes. The ability of exogenously delivered PI(3,4,5)P(3) to augment plasma membrane GLUT4 content without increasing glucose uptake suggests that input at the level of PI 3-kinase suffices for GLUT4 translocation but is insufficient to stimulate glucose transport.     

Phenotypic evolution of human craniofacial morphology after admixture: a geometric morphometrics approach

An evolutionary, diachronic approach to the phenotypic craniofacial pattern arisen in a human population after high levels of admixture and gene flow was achieved by means of geometric morphometrics. Admixture has long been studied after molecular data. Nevertheless, few efforts have been made to explain the morphological outcome in human craniofacial samples. The Spanish-Amerindian contact can be considered a good scenario for such an analysis. Here we present a comparative analysis of craniofacial shape changes observed between two putative ancestor groups, Spanish and precontact Aztecs, and two diachronic admixed groups, corresponding to early and late colonial periods from the Mexico's Central Valley. Quantitative shape comparisons of Amerindian, Spanish, and admixed groups were used to test the expectations of quantitative genetics for admixture events. In its simplest form, this prediction states that an admixed group will present phenotypic values falling between those of both parental groups. Results show that, in general terms, although the human skull is a complex, integrated structure, the craniofacial morphology observed fits the theoretical expectations of quantitative genetics. Thus, it is predictive of population structure and history. In fact, results obtained after the craniofacial analysis are in accordance with previous molecular and historical interpretations, providing evidence that admixture is a main microevolutionary agent influencing modern Mexican gene pool. However, expectations are not straightforward when moderate shape changes are considered. Deviations detected at localized structures, such as the upper and lower face, highlight the evolution of a craniofacial pattern exclusively inherent to the admixed groups, indicating that quantitative characters might respond to admixture in a complicated, nondirectional way.     

Rate of oxidant stress regulates balance between rat gastric mucosa proliferation and apoptosis

We have characterized an experimental model of ethanol-induced chronic gastritis in which a compensatory mucosal cell proliferation is apparently regulated by lipoperoxidative events. Therefore, the present study is an attempt to further assess the participation of oxidant stress during gastric mucosa proliferation, by administering alpha-tocopherol (vitamin E) to rats with gastritis. A morphometric analysis was done, and parameters indicative of oxidant stress, cellular proliferation (including cyclin D1 levels), apoptotic events, and activities of endogenous antioxidant systems were measured in gastric mucosa from our experimental groups. After ethanol withdrawal, restitution of surface epithelium coincided with increased lipid peroxidation and cell proliferation and further active apoptosis. High alpha-tocopherol dosing (100 IU/kg bw) showed a clear antioxidant effect, abolished cell proliferation, and promoted an early and progressive apoptosis, despite vitamin E also enhancing levels of endogenous antioxidants. Indicators of cell proliferation inversely correlated with apoptotic events, and this relationship was blunted by administering vitamin E, probably by affecting translocation of active cyclin D1 into the nucleus. In conclusion, alpha-tocopherol administration inhibited cell proliferation, leading to a predominance of apoptotic events in ethanol-induced gastric damage. Therefore, the timing and magnitude of lipoperoxidative events seemed to synchronize in vivo cell proliferative and apoptotic events, probably by changing the cell redox state.