Exploring the Mechanism of Biocatalyst Inhibition in Microbial Desulfurization

Microbial desulfurization, or biodesulfurization (BDS), of fuels is a promising technology because it can desulfurize compounds that are recalcitrant to the current standard technology in the oil industry. One of the obstacles to the commercialization of BDS is the reduction in biocatalyst activity concomitant with the accumulation of the end product, 2-hydroxybiphenyl (HBP), during the process. BDS experiments were performed by incubating Rhodococcus erythropolis IGTS8 resting-cell suspensions with hexadecane at 0.50 (vol/vol) containing 10 mM dibenzothiophene. The resin Dowex Optipore SD-2 was added to the BDS experiments at resin concentrations of 0, 10, or 50 g resin/liter total volume. The HBP concentration within the cytoplasm was estimated to decrease from 1,100 to 260 μM with increasing resin concentration. Despite this finding, productivity did not increase with the resin concentration. This led us to focus on the susceptibility of the desulfurization enzymes toward HBP. Dose-response experiments were performed to identify major inhibitory interactions in the most common BDS pathway, the 4S pathway. HBP was responsible for three of the four major inhibitory interactions identified. The concentrations of HBP that led to a 50% reduction in the enzymes'' activities (IC₅₀s) for DszA, DszB, and DszC were measured to be 60 ± 5 μM, 110 ± 10 μM, and 50 ± 5 μM, respectively. The fact that the IC₅₀s for HBP are all significantly lower than the cytoplasmic HBP concentration suggests that the inhibition of the desulfurization enzymes by HBP is responsible for the observed reduction in biocatalyst activity concomitant with HBP generation.     

Genomewide predictions from maize single-cross data

Maize (Zea mays L.) breeders evaluate many single-cross hybrids each year in multiple environments. Our objective was to determine the usefulness of genomewide predictions, based on marker effects from maize single-cross data, for identifying the best untested single crosses and the best inbreds within a biparental cross. We considered 479 experimental maize single crosses between 59 Iowa Stiff Stalk Synthetic (BSSS) inbreds and 44 non-BSSS inbreds. The single crosses were evaluated in multilocation experiments from 2001 to 2009 and the BSSS and non-BSSS inbreds had genotypic data for 669 single nucleotide polymorphism (SNP) markers. Single-cross performance was predicted by a previous best linear unbiased prediction (BLUP) approach that utilized marker-based relatedness and information on relatives, and from genomewide marker effects calculated by ridge-regression BLUP (RR-BLUP). With BLUP, the mean prediction accuracy (r _MG) of single-cross performance was 0.87 for grain yield, 0.90 for grain moisture, 0.69 for stalk lodging, and 0.84 for root lodging. The BLUP and RR-BLUP models did not lead to r _MG values that differed significantly. We then used the RR-BLUP model, developed from single-cross data, to predict the performance of testcrosses within 14 biparental populations. The r _MG values within each testcross population were generally low and were often negative. These results were obtained despite the above-average level of linkage disequilibrium, i.e., r ² between adjacent markers of 0.35 in the BSSS inbreds and 0.26 in the non-BSSS inbreds. Overall, our results suggested that genomewide marker effects estimated from maize single crosses are not advantageous (compared with BLUP) for predicting single-cross performance and have erratic usefulness for predicting testcross performance within a biparental cross.     

Genomic Pathology of SLE-Associated Copy-Number Variation at the FCGR2C/FCGR3B/FCGR2B Locus

Reduced FCGR3B copy number is associated with increased risk of systemic lupus erythematosus (SLE). The five FCGR2/FCGR3 genes are arranged across two highly paralogous genomic segments on chromosome 1q23. Previous studies have suggested mechanisms for structural rearrangements at the FCGR2/FCGR3 locus and have proposed mechanisms whereby altered FCGR3B copy number predisposes to autoimmunity, but the high degree of sequence similarity between paralogous segments has prevented precise definition of the molecular events and their functional consequences. To pursue the genomic pathology associated with FCGR3B copy-number variation, we integrated sequencing data from fosmid and bacterial artificial chromosome clones and sequence-captured DNA from FCGR3B-deleted genomes to establish a detailed map of allelic and paralogous sequence variation across the FCGR2/FCGR3 locus. This analysis identified two highly paralogous 24.5 kb blocks within the FCGR2C/FCGR3B/FCGR2B locus that are devoid of nonpolymorphic paralogous sequence variations and that define the limits of the genomic regions in which nonallelic homologous recombination leads to FCGR2C/FCGR3B copy-number variation. Further, the data showed evidence of swapping of haplotype blocks between these highly paralogous blocks that most likely arose from sequential ancestral recombination events across the region. Functionally, we found by flow cytometry, immunoblotting and cDNA sequencing that individuals with FCGR3B-deleted alleles show ectopic presence of FcγRIIb on natural killer (NK) cells. We conclude that FCGR3B deletion juxtaposes the 5′-regulatory sequences of FCGR2C with the coding sequence of FCGR2B, creating a chimeric gene that results in an ectopic accumulation of FcγRIIb on NK cells and provides an explanation for SLE risk associated with reduced FCGR3B gene copy number.     

Past and future demographic dynamics of alpine species: limited genetic consequences despite dramatic range contraction in a plant from the Spanish Sierra Nevada

Anthropogenic global climate change is expected to cause severe range contractions among alpine plants. Alpine areas in the Mediterranean region are of special concern because of the high abundance of endemic species with narrow ranges. This study combined species distribution models, population structure analyses and Bayesian skyline plots to trace the past and future distribution and diversity of Linaria glacialis, an endangered narrow endemic species that inhabits summits of Sierra Nevada (Spain). The results showed that: (i) the habitat of this alpine‐Mediterranean species in Sierra Nevada suffered little changes during glacial and interglacial stages of late Quaternary; (ii) climatic oscillations in the last millennium (Medieval Warm Period and Little Ice Age) moderately affected the demographic trends of L. glacialis; (iii) future warming conditions will cause severe range contractions; and (iv) genetic diversity will not diminish at the same pace as the distribution range. As a consequence of the low population structure of this species, genetic impoverishment in the alpine zones of Sierra Nevada should be limited during range contraction. We conclude that maintenance of large effective population sizes via high mutation rates and high levels of gene flow may promote the resilience of alpine plant species when confronted with global warming.     

Exclusive rewards in mutualisms: ant proteases and plant protease inhibitors create a lock–key system to protect Acacia food bodies from exploitation

Myrmecophytic Acacia species produce food bodies (FBs) to nourish ants of the Pseudomyrmex ferrugineus group, with which they live in an obligate mutualism. We investigated how the FBs are protected from exploiting nonmutualists. Two‐dimensional gel electrophoresis of the FB proteomes and consecutive protein sequencing indicated the presence of several Kunitz‐type protease inhibitors (PIs). PIs extracted from Acacia FBs were biologically active, as they effectively reduced the trypsin‐like and elastase‐like proteolytic activity in the guts of seed‐feeding beetles (Prostephanus truncatus and Zabrotes subfasciatus), which were used as nonadapted herbivores representing potential exploiters. By contrast, the legitimate mutualistic consumers maintained high proteolytic activity dominated by chymotrypsin 1, which was insensitive to the FB PIs. Larvae of an exploiter ant (Pseudomyrmex gracilis) taken from Acacia hosts exhibited lower overall proteolytic activity than the mutualists. The proteases of this exploiter exhibited mainly elastase‐like and to a lower degree chymotrypsin 1‐like activity. We conclude that the mutualist ants possess specifically those proteases that are least sensitive to the PIs in their specific food source, whereas the congeneric exploiter ant appears partly, but not completely, adapted to consume Acacia FBs. By contrast, any consumption of the FBs by nonadapted exploiters would effectively inhibit their digestive capacities. We suggest that the term ‘exclusive rewards’ can be used to describe situations similar to the one that has evolved in myrmecophytic Acacia species, which reward mutualists with FBs but safeguard the reward from exploitation by generalists by making the FBs difficult for the nonadapted consumer to use.     

TNF-related weak inducer of apoptosis (TWEAK) promotes kidney fibrosis and Ras-dependent proliferation of cultured renal fibroblast

Tumor necrosis factor-like weak inducer of apoptosis (TWEAK) regulates apoptosis, proliferation and inflammation in renal epithelial cells and plays a role in acute kidney injury. However, there is little information on the chronic effects of TWEAK. We hypothesized that TWEAK may influence renal fibrosis and regulate kidney fibroblast biology, in part, through Ras pathway.We studied a chronic model of experimental unilateral ureteral obstruction in wild type and TWEAK deficient mice, and a murine model of systemic TWEAK overexpression. TWEAK actions were also explored in cultured renal and embryonic fibroblasts.TWEAK and TWEAK receptor expression was increased in the obstructed kidneys. The absence of TWEAK decreased early kidney tubular damage, inflammatory infiltrates and myofibroblast number. TWEAK deficient mice had decreased renal fibrosis 21 days after obstruction, as assessed by extracellular matrix staining. In mice without prior underlying kidney disease, systemic overexpression of TWEAK induced kidney inflammation and fibrosis. In cultured fibroblasts, TWEAK induced proliferation through activation of the Ras/ERK pathway. TWEAK also activated nuclear factor κB (NFκB)-dependent inflammatory chemokine production in murine renal fibroblasts.In conclusion, lack of TWEAK reduces renal fibrosis in a model of persistent kidney insult and overexpression of TWEAK led to renal fibrosis. TWEAK actions on renal fibroblasts may contribute to the in vivo observations, as TWEAK promotes inflammatory activity and proliferation in fibroblast cultures.     

Functional and molecular insights into KSRP function in mRNA decay

KSRP is a single strand nucleic acid binding protein that controls gene expression at multiple levels. In this review we focus on the recent molecular, cellular, and structural insights into the mRNA decay promoting function of KSRP. We discuss also some aspects of KSRP-dependent microRNA maturation from precursors that are related to its mRNA destabilizing function. This article is part of a Special Issue entitled: RNA Decay mechanisms.     

Increased Learning and Brain Long-Term Potentiation in Aged Mice Lacking DNA Polymerase μ

A definitive consequence of the aging process is the progressive deterioration of higher cognitive functions. Defects in DNA repair mechanisms mostly result in accelerated aging and reduced brain function. DNA polymerase µ is a novel accessory partner for the non-homologous end-joining DNA repair pathway for double-strand breaks, and its deficiency causes reduced DNA repair. Using associative learning and long-term potentiation experiments, we demonstrate that Polµ^−/− mice, however, maintain the ability to learn at ages when wild-type mice do not. Expression and biochemical analyses suggest that brain aging is delayed in Polµ^−/− mice, being associated with a reduced error-prone DNA oxidative repair activity and a more efficient mitochondrial function. This is the first example in which the genetic ablation of a DNA-repair function results in a substantially better maintenance of learning abilities, together with fewer signs of brain aging, in old mice.     

An Optimized Immunohistochemistry Technique Improves NMO-IgG Detection: Study Comparison with Cell-Based Assays

Cell-based assays (CBA) have increased the sensitivity of the neuromyelitis optica (NMO)-IgG/aquaporin-4-antibody detection compared to classical tissue-based indirect assays. We describe the sensitivity of an optimized immunohistochemistry (IHC-o) to detect NMO-IgG/aquaporin-4-antibody in comparison with that of two CBA: an in-house (CBA-ih) and a commercial (CBA-c) assay (Euroimmun, Germany). Coded serum from 103 patients with definite NMO and 122 inflammatory controls were studied by IHC-o, CBA-ih, and CBA-c. IHC-o used the same protocol described to detect antibodies against cell surface antigens. CBA-ih used live cells transfected with the aquaporin-4-M23-isoform. The sensitivity of the IHC-o was 74.8% (95% confidence interval [CI] 65-83) and was similar to that of the CBA-ih 75.7% (95% CI 66-84) and the CBA-c 73.8% (95% CI 64-82). The specificity of the three assays was 100% (95% CI 97-100). Interassay concordance was high, 100 of 103 samples were coincident in all techniques. The optimized immunohistochemistry proves to be as sensitive and specific as the cell-based assays. This assay extends the available tools for NMO-IgG/aquaporin-4-antibody detection.