Ammi Visnaga Lam.—A medicinal plant

Extensive investigation has shown that khellin, an active principle obtained from this plant, is of value in treating angina pectoris and bronchial asthma.     

The European carbon balance. Part 2: croplands

We estimated the long‐term carbon balance [net biome production (NBP)] of European (EU‐25) croplands and its component fluxes, over the last two decades. Net primary production (NPP) estimates, from different data sources ranged between 490 and 846 gC m^?2 yr^?1, and mostly reflect uncertainties in allocation, and in cropland area when using yield statistics. Inventories of soil C change over arable lands may be the most reliable source of information on NBP, but inventories lack full and harmonized coverage of EU‐25. From a compilation of inventories we infer a mean loss of soil C amounting to 17 g m^?2 yr^?1. In addition, three process‐based models, driven by historical climate and evolving agricultural technology, estimate a small sink of 15 g C m^?2 yr^?1 or a small source of 7.6 g C m^?2 yr^?1. Neither the soil C inventory data, nor the process model results support the previous European‐scale NBP estimate by Janssens and colleagues of a large soil C loss of 90 ± 50 gC m^?2 yr^?1. Discrepancy between measured and modeled NBP is caused by erosion which is not inventoried, and the burning of harvest residues which is not modeled. When correcting the inventory NBP for the erosion flux, and the modeled NBP for agricultural fire losses, the discrepancy is reduced, and cropland NBP ranges between ?8.3 ± 13 and ?13 ± 33 g C m^?2 yr^?1 from the mean of the models and inventories, respectively. The mean nitrous oxide (N₂O) flux estimates ranges between 32 and 37 g C Eq m^?2 yr^?1, which nearly doubles the CO₂ losses. European croplands act as small CH₄ sink of 3.3 g C Eq m^?2 yr^?1. Considering ecosystem CO₂, N₂O and CH₄ fluxes provides for the net greenhouse gas balance a net source of 42–47 g C Eq m^?2 yr^?1. Intensifying agriculture in Eastern Europe to the same level Western Europe amounts is expected to result in a near doubling of the N₂O emissions in Eastern Europe. N₂O emissions will then become the main source of concern for the impact of European agriculture on climate.     

Dose-dependant hypothyroidism in mice induced by commercial trimethoprim-sulfamethoxazole rodent feed

Trimethoprim-sulfamethoxazole (TMP-SMX) medication in the feed or water is commonly administered to immunocompro mised mice to prevent the occurrence of Pneumocystis murina (formerly P. carinii) pneumonia. Therapeutic doses of SMX can cause decreased total and free thyroxine (T4) levels in dogs and thyroid hypertrophy and hyperplasia in mice, rats, and dogs. Our primary objective was to determine whether SMX at doses present in commercially available rodent TMP-SMX feed would pro duce hypothyroidism in mice. Plasma T4 levels were determined prior to and after placement of Brand A TMP-SMX feed (daily SMX dose, 240 mg/kg), Brand B TMP-SMX feed (daily SMX dose, 2400 mg/kg), and their respective controls (doses calculated for a 25-g mouse according to vendor's information). T4 levels in the mice fed Brand B TMP-SMX feed were significantly decreased by 2 wk after feed placement. Levels of thyroid stimulating hormone in male and female mice given Brand B TMP-SMX feed were significantly elevated compared with those of control groups at 6 wk after feed placement, when only these mice showed evidence of thyroid hypertrophy and hyperplasia. No significant change in T4 levels occurred over the course of 11 wk in mice given the Brand A TMP-SMX chow or either control feed. In light of the significant clinical hypothyroidism that occurred in our mice while receiving Brand B TMP-SMX diet, we recommend SMX levels more similar to that of Brand A to avoid such unwanted effects which could confound research data.     

The small GTPase Ran: interpreting the signs

The small GTPase Ran has roles in nuclear transport, mitotic spindle assembly and nuclear envelope assembly. During the past three years, it has become clear that many of these processes rely on conserved molecular mechanisms involving Ran-GTP-binding proteins of the importin-beta superfamily. Moreover, recent experimental evidence has documented the distribution of Ran-GTP within cells and supported the notion that Ran plays a central role in the spatial and temporal organization of the eukaryotic cell.     

Structural basis for the interaction between NTF2 and nucleoporin FxFG repeats

Interactions with nucleoporins containing FxFG-repeat cores are crucial for the nuclear import of RanGDP mediated by nuclear transport factor 2 (NTF2). We describe here the 1.9 A resolution crystal structure of yeast NTF2-N77Y bound to a FxFG-nucleoporin core, which provides a basis for understanding this interaction and its role in nuclear trafficking. The two identical FxFG binding sites on the dimeric molecule are formed by residues from each chain of NTF2. Engineered mutants at the interaction interface reduce the binding of NTF2 to nuclear pores and cause reduced growth rates and Ran mislocalization when substituted for the wild-type protein in yeast. Comparison with the crystal structure of FG-nucleoporin cores bound to importin-beta and TAP/p15 identified a number of common features of their binding sites. The structure of the binding interfaces on these transport factors provides a rationale for the specificity of their interactions with nucleoporins that, combined with their weak binding constants, facilitates rapid translocation through NPCs during nuclear trafficking.     

N-Linked glycosylation of a baculovirus-expressed recombinant glycoprotein in insect larvae and tissue culture cells

The potential of insect cell cultures and larvae infected with recombinant baculoviruses to produce authentic recombinant glycoproteins cloned from mammalian sources was investigated. A comparison was made of the N-linked glycans attached to secreted alkaline phosphatase (SEAP) produced in four species of insect larvae and their derived cell lines plus one additional insect cell line and larvae of one additional species. These data survey N-linked oligosaccharides produced in four families and six genera of the order Lepidoptera. Recombinant SEAP expressed by recombinant isolates of Autographa californica and Bombyx mori nucleopolyhedroviruses was purified from cell culture medium, larval hemolymph or larval homogenates by phosphate affinity chromatography. The N-linked oligosaccharides were released with PNGase-F, labeled with 8- aminonaphthalene-1-3-6-trisulfonic acid, fractionated by polyacrylamide gel electrophoresis, and analyzed by fluorescence imaging. The oligosaccharide structures were confirmed with exoglycosidase digestions. Recombinant SEAP produced in cell lines of Lymantria dispar (IPLB-LdEIta), Heliothis virescens (IPLB-HvT1), and Bombyx mori (BmN) and larvae of Spodoptera frugiperda, Trichoplusia ni , H.virescens , B.mori , and Danaus plexippus contained oligosaccharides that were structurally identical to the 10 oligosaccharides attached to SEAP produced in T.ni cell lines. The oligosaccharide structures were all mannose-terminated. Structures containing two or three mannose residues, with and without core fucosylation, constituted more than 75% of the oligosaccharides from the cell culture and larval samples.      

Sensing the effect of body load in legs: responses of tibial campaniform sensilla to forces applied to the thorax in freely standing cockroaches

Sense organs in the legs that detect body weight are an important component in the regulation of posture and locomotion. We tested the abilities of tibial campaniform sensilla, receptors that can monitor forces in the cockroach leg, to encode variations in body load in freely standing animals. Small magnets were attached to the thorax and currents were applied to a coil below the substrate. Sensory and motor activities were monitored neurographically. The tibial sensilla could show vigorous discharges to changing forces when animals stood upon their legs and actively supported the body weight. Firing of individual afferents depended upon the orientation of the receptors cuticular cap: proximal sensilla (oriented perpendicular to the leg axis) discharged to force increases while distal receptors (parallel to the leg) fired to decreasing forces. Proximal sensillum discharges were prolonged and could encode the level of load when increases were sustained. Firing of the trochanteral extensor motoneuron was also strongly modulated by changing load. In some postures, sensillum discharges paralleled changes in motor frequency consistent with a known interjoint reflex. These findings demonstrate that tibial campaniform sensilla can monitor the effects of body weight upon the legs and may aid in generating support of body load.     

Functional analysis of the hydrophobic patch on nuclear transport factor 2 involved in interactions with the nuclear pore in vivo

Nuclear transport factor 2 (NTF2) is a small homodimeric protein that interacts simultaneously with both RanGDP and FxFG nucleoporins. The interaction between NTF2 and Ran is essential for the import of Ran into the nucleus. Here we use mutational analysis to dissect the in vivo role of the interaction between NTF2 and nucleoporins. We identify a series of surface residues that form a hydrophobic patch on NTF2, which when mutated disrupt the NTF2-nucleoporin interaction. Analysis of these mutants in vivo demonstrates that the strength of this interaction can be significantly reduced without affecting cell viability. However, cells cease to be viable if the interaction between NTF2 and nucleoporins is abolished completely, indicating that this interaction is essential for the function of NTF2 in vivo. In addition, we have isolated a dominant negative mutant of NTF2, N77Y, which has increased affinity for nucleoporins. Overexpression of the N77Y protein blocks nuclear protein import and concentrates Ran at the nuclear rim. These data support a mechanism in which NTF2 interacts transiently with FxFG nucleoporins to translocate through the pore and import RanGDP into the nucleus.     

Evaluation of APOE polymorphisms and the risk for age-related macular degeneration in a Southeastern Brazilian population

This study aimed to evaluate the role of APOE polymorphisms (rs429358 and rs7412) in the risk of age-related macular degeneration in a sample of the Southeastern Brazilian population. Seven hundred and five unrelated individuals were analyzed, 334 with age-related macular degeneration (case group), and 371 without the disease (control group). In the case group, patients were further stratified according to disease phenotypes, divided into dry and wet age-related macular degeneration, and non-advanced and advanced age-related macular degeneration. APOE polymorphisms (rs429358 and rs7412) were evaluated through polymerase chain reaction and direct sequencing. In the comparison of cases vs. controls, none of the associations reached statistical significance, considering the Bonferroni-adjusted P-value, although there was a suggestive protection for the E3/E4 genotype (OR = 0.626; P-value = 0.037) and E4 carriers (OR = 0.6515; P-value = 0.047). Statistically significant protection for both the E3/E4 genotype and E4 carriers was observed in the comparisons: advanced age-related macular degeneration vs. controls (OR = 0.3665, P-value = 0.491 × 10⁻³ and OR = 0.4031, P-value = 0.814 × 10⁻³, respectively), advanced age-related macular degeneration vs. non-advanced age-related macular degeneration (OR = 0.2529, P-value = 0.659 × 10⁻⁴ and OR = 0.2692, P-value = 0.631 × 10⁻⁴, respectively). In the comparison of wet age-related macular degeneration vs. control, protection was statistically significant only for E3/E4 (OR = 0.4052, P-value = 0.001). None of the comparisons demonstrated any significant association for E2 genotypes or E2 carriers in age-related macular degeneration risk in this study. Findings suggest a protective role of the E4 haplotype in the APOE gene in the risk for advanced and wet forms of age-related macular degeneration, in a sample of the Brazilian population. To our knowledge, this is the first Brazilian study to show the association between APOE polymorphisms and age-related macular degeneration.