Major differences in antigen-processing correlate with a single Arg71<-->Lys substitution in HLA-DR molecules predisposing to rheumatoid arthritis and with their selective interactions with 70-kDa heat shock protein chaperones

Several HLA-DR alleles are genetically associated with rheumatoid arthritis. DRB1*0401 predominates in Northern Europe and has a characteristic (70)QKRAA motif. This sequence contacts bound peptides and the TCR. Further interactions have been suggested with additional proteins during Ag loading. We explored the much stronger processing/presentation of full-length recombinant human acetylcholine receptor alpha subunit to a specific T cell clone by APC from DRB1*0401+ than *0408+ donors. Using DR*04 transfectants, we show that this difference results largely from the single Lys71<-->Arg interchange (0401<-->0408), which scarcely affects epitope binding, rather than from any other associated polymorphism. Furthermore, we proved our recombinant polypeptides to contain the Escherichia coli 70-kDa heat shock protein molecule DnaK and its requirement for efficient processing and presentation of the epitope by DRB1*0401+ cells. According to a recent report, 70-kDa heat shock protein chaperones preferentially bind to the QKRAA, rather than the QRRAA, motif. Variations between the shared epitope motifs QKRAA and QRRAA are emphasized by underlining. We propose that such interactions enhance the intracellular epitope loading of *0401 molecules. They may thus broaden immune responses to pathogens and at least partially explain the distinct contributions of DRB1*0401 and other alleles to disease predisposition.     

The fission yeast ubiquitin-conjugating enzymes UbcP3, Ubc15, and Rhp6 affect transcriptional silencing of the mating-type region

Genes transcribed by RNA polymerase II are silenced when introduced near the mat2 or mat3 mating-type loci of the fission yeast Schizosaccharomyces pombe. Silencing is mediated by a number of gene products and cis-acting elements. We report here the finding of novel trans-acting factors identified in a screen for high-copy-number disruptors of silencing. Expression of cDNAs encoding the putative E2 ubiquitin-conjugating enzymes UbcP3, Ubc15 (ubiquitin-conjugating enzyme), or Rhp6 (Rad homolog pombe) from the strong nmt1 promoter derepressed the silent mating-type loci mat2 and mat3 and reporter genes inserted nearby. Deletion of rhp6 slightly derepressed an ade6 reporter gene placed in the mating-type region, whereas disruption of ubcP3 or ubc15 had no obvious effect on silencing. Rhp18 is the S. pombe homolog of Saccharomyces cerevisiae Rad18p, a DNA-binding protein that physically interacts with Rad6p. Rhp18 was not required for the derepression observed when UbcP3, Ubc15, or Rhp6 was overproduced. Overexpressing Rhp6 active-site mutants showed that the ubiquitin-conjugating activity of Rhp6 is essential for disruption of silencing. However, high dosage of UbcP3, Ubc15, or Rhp6 was not suppressed by a mutation in the 26S proteasome, suggesting that loss of silencing is not due to an increased degradation of silencing factors but rather to the posttranslational modification of proteins by ubiquitination. We discuss the implications of these results for the possible modes of action of UbcP3, Ubc15, and Rhp6.     

Lack of mannose-binding lectin-A enhances survival in a mouse model of acute septic peritonitis

The mannose-binding lectin (MBL) (also known as the mannose-binding protein) is a serum protein that plays a role as an "ante-antibody" in innate immunity. In man, MBL is encoded by a single gene, whereas in mice there are two homologous proteins, MBL-A and MBL-C. In order to evaluate the relative roles of these two forms of MBL, we created MBL-A null mice that were MBL-C sufficient. We found MBL-A null mice had enhanced survival in a septic peritonitis model compared to wild-type mice and complement 3 null mice at 24 h, 48 h and 10 d (P < 0.05). Reconstitution of these mice with human MBL reversed the phenotype. Surviving mice had significantly decreased TNF-alpha and IL-6 levels in the blood and peritoneal cavity (P < 0.01). In vitro studies indicate that bacteria opsonized with MBL-A-deficient serum induced significantly less cytokine by peritoneal macrophages compared to those with wild-type serum. Our results indicate that MBL-A is a modulator of inflammation in vivo and in vitro in the mouse and that the role of MBL may extend beyond its role as an opsonin.     

GeneCards 2002: towards a complete,object-oriented,human gene compendium

MOTIVATION: In the post-genomic era, functional analysis of genes requires a sophisticated interdisciplinary arsenal. Comprehensive resources are challenged to provide consistently improving, state-of-the-art tools. RESULTS: GeneCards (Rebhan et al., 1998) has made innovative strides: (a). regular updates and enhancements incorporating new genes enriched with sequences, genomic locations, cDNA assemblies, orthologies, medical information, 3D protein structures, gene expression, and focused SNP summaries; (b). restructured software using object-oriented Perl, migration to schema-driven XML, and (c). pilot studies, introducing methods to produce cards for novel and predicted genes.     

Mechanism of electrogenic cation transport by the cloned organic cation transporter 2 from rat

The organic cation transporter 2 (OCT2) is expressed in plasma membranes of kidney and brain. Its transport mechanism and substrates are debated. We studied substrate-induced changes of electrical current with the patch clamp technique after expression of rat OCT2 in oocytes. Activation of current, corresponding to efflux, was observed for small organic cations, e.g. choline. In contrast, the bigger cations quinine and tetrabutylammonium elicited no change in current. However, transport of choline could be inhibited by applying quinine or tetrabutylammonium to the cytoplasmic side. Inhibition of organic cation efflux by quinine was competitive with substrates. Quinine at the inside also inhibited substrate influx from the outside. Current-voltage analysis showed that both maximal turnover and apparent affinity to substrates are voltage-dependent. Substrate-induced currents with organic cations on both membrane sides reversed as predicted from the Nernst potential. Our results clearly identify the electrochemical potential as driving force for transport at neutral pH and exclude an electroneutral H(+)/organic cation(+) exchange. We suggest the existence of an electroneutral organic cation(+) exchange and propose a model for a carrier-type transport mechanism.     

Significant increase in natural disturbance impacts on European forests since 1950

Over the last decades, the natural disturbance is increasingly putting pressure on European forests. Shifts in disturbance regimes may compromise forest functioning and the continuous provisioning of ecosystem services to society, including their climate change mitigation potential. Although forests are central to many European policies, we lack the long-term empirical data needed for thoroughly understanding disturbance dynamics, modeling them, and developing adaptive management strategies. Here, we present a unique database of >170,000 records of ground-based natural disturbance observations in European forests from 1950 to 2019. Reported data confirm a significant increase in forest disturbance in 34 European countries, causing on an average of 43.8 million m³ of disturbed timber volume per year over the 70-year study period. This value is likely a conservative estimate due to under-reporting, especially of small-scale disturbances. We used machine learning techniques for assessing the magnitude of unreported disturbances, which are estimated to be between 8.6 and 18.3 million m³/year. In the last 20 years, disturbances on average accounted for 16% of the mean annual harvest in Europe. Wind was the most important disturbance agent over the study period (46% of total damage), followed by fire (24%) and bark beetles (17%). Bark beetle disturbance doubled its share of the total damage in the last 20 years. Forest disturbances can profoundly impact ecosystem services (e.g., climate change mitigation), affect regional forest resource provisioning and consequently disrupt long-term management planning objectives and timber markets. We conclude that adaptation to changing disturbance regimes must be placed at the core of the European forest management and policy debate. Furthermore, a coherent and homogeneous monitoring system of natural disturbances is urgently needed in Europe, to better observe and respond to the ongoing changes in forest disturbance regimes.     

Large‐scale disturbance legacies and the climate sensitivity of primary Picea abies forests

Determining the drivers of shifting forest disturbance rates remains a pressing global change issue. Large‐scale forest dynamics are commonly assumed to be climate driven, but appropriately scaled disturbance histories are rarely available to assess how disturbance legacies alter subsequent disturbance rates and the climate sensitivity of disturbance. We compiled multiple tree ring‐based disturbance histories from primary Picea abies forest fragments distributed throughout five European landscapes spanning the Bohemian Forest and the Carpathian Mountains. The regional chronology includes 11,595 tree cores, with ring dates spanning the years 1750–2000, collected from 560 inventory plots in 37 stands distributed across a 1,000 km geographic gradient, amounting to the largest disturbance chronology yet constructed in Europe. Decadal disturbance rates varied significantly through time and declined after 1920, resulting in widespread increases in canopy tree age. Approximately 75% of current canopy area recruited prior to 1900. Long‐term disturbance patterns were compared to an historical drought reconstruction, and further linked to spatial variation in stand structure and contemporary disturbance patterns derived from LANDSAT imagery. Historically, decadal Palmer drought severity index minima corresponded to higher rates of canopy removal. The severity of contemporary disturbances increased with each stand's estimated time since last major disturbance, increased with mean diameter, and declined with increasing within‐stand structural variability. Reconstructed spatial patterns suggest that high small‐scale structural variability has historically acted to reduce large‐scale susceptibility and climate sensitivity of disturbance. Reduced disturbance rates since 1920, a potential legacy of high 19th century disturbance rates, have contributed to a recent region‐wide increase in disturbance susceptibility. Increasingly common high‐severity disturbances throughout primary Picea forests of Central Europe should be reinterpreted in light of both legacy effects (resulting in increased susceptibility) and climate change (resulting in increased exposure to extreme events).     

Cortisol and progestin release, heart rate and heart rate variability in the pregnant and postpartum mare, fetus and newborn foal

The mechanisms leading to parturition in the horse in many aspects differ from those in other species. Pregnancy is maintained not by progesterone but by 5α-pregnanes and the progestin precursor pregnenolone originates from the fetus. As parturition approaches, the fetal adrenal switches from pregnenolone to cortisol synthesis but it is not known whether cortisol crosses the placenta. We hypothesized that in parallel to fetal cortisol release, cortisol in the maternal circulation increases before foaling and this increase can be determined in both saliva and plasma. In addition, maternal, fetal and neonatal heart rate and heart rate variability were measured. In 25 pregnant mares, saliva for cortisol analysis was collected 4 times daily from 15 days before to 5 days after foaling. In 13 mares, in addition, fetomaternal electrocardiogram (ECG) recordings were made and blood samples for progestin and cortisol analysis were collected once daily. Heart rate (HR) was recorded until 5 days after foaling. The heart rate variability (HRV) variables standard deviation of the beat-to-beat (RR) interval (SDRR) and root mean square of successive RR differences (RMSSD) were calculated. From Days 15 to 4 before parturition, progestin concentration increased (peak 267 ± 42 ng/mL) and decreased thereafter (P < 0.05, day of foaling 113 ± 18 ng/mL). A prepartum increase in maternal cortisol concentrations was evident in blood (P < 0.05) and saliva (P < 0.05) and paralleled the decrease in progestin concentrations. In mares, HR remained constant during the last days of pregnancy but decreased within one day after parturition (P < 0.05) while maternal HRV did not change. In the fetus and neonate, HR increased from before to after birth (P < 0.05) indicating increasing demands on the cardiovascular system with adaptation to extrauterine life.     

Responses of N fluxes and pools to elevated atmospheric CO2 in model forest ecosystems with acidic and calcareous soils

The objectives of this study were to estimate how soil type, elevated N deposition (0.7 vs. 7 g N m^–2y^–1) and tree species influence the potential effects of elevated CO₂ (370 vs. 570 mol CO₂ mol^–1) on N pools and fluxes in forest soils. Model spruce-beech forest ecosystems were established on a nutrient-rich calcareous sand and on a nutrient-poor acidic loam in large open-top chambers. In the fourth year of treatment, we measured N concentrations in the soil solution at different depths, estimated N accumulation by ion exchange resin (IER) bags, and quantified N export in drainage water, denitrification, and net N uptake by trees. Under elevated CO₂, concentrations of N in the soil solution were significantly reduced. In the nutrient-rich calcareous sand, CO₂ enrichment decreased N concentrations in the soil solution at all depths (–45 to –100%). In the nutrient-poor acidic loam, the negative CO₂ effect was restricted to the uppermost 5 cm of the soil. Increasing the N deposition stimulated the negative impact of CO₂ enrichment on soil solution N in the acidic loam at 5 cm depth from –20% at low N inputs to –70% at high N inputs. In the nutrient-rich calcareous sand, N additions did not influence the CO₂ effect on soil solution N. Accumulation of N by IER bags, which were installed under individual trees, was decreased at high CO₂ levels under spruce in both soil types. Under beech, this decrease occurred only in the calcareous sand. N accumulation by IER bags was negatively correlated with current-years foliage biomass, suggesting that the reduction of soil N availability indices was related to a CO₂-induced growth enhancement. However, the net N uptake by trees was not significantly increased by elevated CO₂. Thus, we suppose that the reduced N concentrations in the soil solution at elevated CO₂ concentrations were rather caused by an increased N immobilisation in the soil. Denitrification was not influenced by atmospheric CO₂ concentrations. CO₂ enrichment decreased nitrate leaching in drainage by 65%, which suggests that rising atmospheric CO₂ potentially increases the N retention capacity of forest ecosystems.