Genetic studies of two sister species in the Drosophila melanogaster subgroup, D. yakuba and D. santomea

We performed genetic analysis of hybrid sterility and of one morphological difference (sex-comb tooth number) on D. yakuba and D. santomea, the former species widespread in Africa and the latter endemic to the oceanic island of S?o Tomé, on which there is a hybrid zone. The sterility of hybrid males is due to at least three genes on the X chromosome and at least one on the Y, with the cytoplasm and large sections of the autosomes having no effect. F1 hybrid females carrying two X chromosomes from either species are perfectly fertile despite their genetic similarity to completely sterile F1 hybrid males. This implies that the appearance of Haldane's rule in this cross is at least partially due to the faster accumulation of genes causing male than female sterility. The larger effects of the X and Y chromosomes than of the autosomes, however, also suggest that the genes causing male sterility are recessive in hybrids. Some female sterility is also seen in interspecific crosses, but this does not occur between all strains. This is seen in pure-species females inseminated by heterospecific males (probably reflecting incompatibility between the sperm of one species and the female reproductive tract of the other) as well as in inseminated F1 and backcross females, probably reflecting genetically based incompatibilities in hybrids that affect the reproductive system. The latter 'innate' sterility appears to involve deleterious interactions between D. santomea chromosomes and D. yakuba cytoplasm. The difference in male sex-comb tooth number appears to involve fairly large effects of the X chromosome. We discuss the striking evolutionary parallels in the genetic basis of sterility, in the nature of sexual isolation, and in morphological differences between the D. santomea/D. yakuba divergence and two other speciation events in the D. melanogaster subgroup involving island colonization.     

Resistance Gene Analog Polymorphism (RGAP) Markers Co-Localize with Disease Resistance Genes and QTL in Common Bean

Resistance (R) genes containing nucleotide-binding site (NBS)-leucine rich repeats (LRR) are the most prevalent types of R gene in plants. The objective of this study was to develop PCR-based R-gene analog polymorphism (RGAP) markers for common bean (Phaseolus vulgaris L). Twenty degenerate primers were designed from the conserved kinase-1a (GVGKTT) and hydrophobic domains (GLPLAL) of known NBS-LRR type R-genes and from EST databases. Sixty-six of the 100 primer combinations tested yielded polymorphism. Thirty-two RGAP markers were mapped in the BAT 93/Jalo EEP558 core mapping population for common bean. The markers mapped to 10 of 11 linkage groups with a strong tendency for clustering. In addition, the RGAP markers co-located, on six linkage groups, with 15 resistance gene analogs (RGAs) that were previously mapped in other populations of common bean. The distance between the priming sites in NBS-LRR type R-genes is around 500 bp. Of the 32 RGAP markers, 19 had sizes larger and 13 less than 500 bp. RGAP markers mapped close to known R-genes on B11, and to QTLs for resistance on B1, B2, B6, B7, B8, B10, and B11. RGAP appears to provide a useful marker technique for tagging and mapping R-genes in segregating common bean populations, discovery of candidate genes underlying resistance QTL, and future cloning of R-genes in common bean.     

An interaction between benzodiazepines and neuroactive steroids at GABA A receptors in cultured hippocampal neurons

Neurosteroids are modulators of several receptors and ion channels and are implicated in the pathophysiology of several neuropsychiatric diseases including hepatic encephalopathy (HE). The neurosteroid, allopregnanolone, a positive allosteric modulator of GABA_A receptors, accumulates in the brains of HE patients where it can potentiate GABA_A receptor-mediated responses. Attenuation of the effects of neurosteroids on GABA-ergic neurotransmission is therefore of interest for the management of HE. In the present study, we determined the effect of the benzodiazepine partial inverse agonist, Ro15-4513, and the benzodiazepine antagonist, flumazenil on modulation of the GABA_A mediated chloride currents by allopregnanolone and on spontaneous synaptic activity in cultured hippocampal neurons using the patch-clamp technique. Allopregnanolone (0.03–0.3 μM), dose-dependently potentiated GABA-induced currents, an action significantly reduced by Ro15-4513 (10 μM). In contrast, flumazenil (10 μM) had no effect on the ability of allopregnanolone to potentiate GABA_A currents but it blocked the effects of Ro15-4513. The frequency of spontaneous synaptic activity was significantly reduced in the presence of allopregnanolone (0.1 μM) from 1.5 ± 0.7 to 0.1 ± 0.04 Hz. This action was partially reversed by Ro15-4513 (10 μM) but was not significantly influenced by flumazenil (10 μM). These findings suggest that the beneficial affects of Ro15-4513 in experimental HE result from attenuation of the effects of neurosteroids at GABA_A receptors. Our results may provide a rational basis for the use of benzodiazepine inverse agonists in the management and treatment of hepatic encephalopathy in patients with liver failure.     

CbgA, a protein involved in cortex formation and stress resistance in Myxococcus xanthus spores

CbgA plays a role in cortex formation and the acquisition of a subset of stress resistance properties in Myxococcus xanthus spores. The cbgA mutant produces spores with thin or no cortex layers, and these spores are more sensitive to heat and sodium dodecyl sulfate than their wild-type counterparts.     

DESCRIPTION OF VIRIDILOBUS MARINUS (GEN. ET SP. NOV.), A NEW RAPHIDOPHYTE FROM DELAWARE'S INLAND BAYS

Delaware's Inland Bays (DIB), USA, are subject to blooms of potentially harmful raphidophytes, including Heterosigma akashiwo. In 2004, a dense bloom was observed in a low salinity tributary of the DIB. Light microscopy initially suggested that the species was H. akashiwo; however, the cells were smaller than anticipated. 18S rDNA sequences of isolated cultures differed substantially from all raphidophyte sequences in GenBank. Phylogenetic analysis placed it approximately equidistant from Chattonella and Heterosigma with only ~96% sequence homology with either group. Here, we describe this marine raphidophyte as a novel genus and species, Viridilobus marinus (gen. et sp. nov.). We also compared this species with H. akashiwo, because both species are superficially similar with respect to morphology and their ecological niches overlap. V. marinus cells are ovoid to spherical (11.4 × 9.4 μm), and the average number of chloroplasts (4 per cell) is lower than in H. akashiwo (15 per cell). Pigment analysis of V. marinus revealed the presence of fucoxanthin, violaxanthin, and zeaxanthin, which are characteristic of marine raphidophytes within the family Chattonellaceae of the Raphidophyceae. TEM and confocal microscopy, however, revealed diagnostic microscopic and ultrastructural characteristics that distinguish it from other raphidophytes. Chloroplasts were in close association with the nucleus and thylakoids were arranged either parallel or perpendicular to the cell surface. Putative mucocysts were identified, but trichocysts were not observed. These features, along with DNA sequence data, distinguish this species from all other raphidophyte genera within the family Chattonellaceae of the Raphidophyceae.     

Lipoteichoic acid is an important microbe-associated molecular pattern of Lactobacillus rhamnosus GG

Background

Receptors with a single transmembrane (TM) domain are essential for the signal transduction across the cell membrane. NMR spectroscopy is a powerful tool to study structure of the single TM domain. The expression and purification of a TM domain in Escherichia coli (E.coli) is challenging due to its small molecular weight. Although ketosteroid isomerase (KSI) is a commonly used affinity tag for expression and purification of short peptides, KSI tag needs to be removed with the toxic reagent cyanogen bromide (CNBr).

Result

The purification of the TM domain of p75 neurotrophin receptor using a KSI tag with the introduction of a thrombin cleavage site is described herein. The recombinant fusion protein was refolded into micelles and was cleaved with thrombin. Studies showed that purified protein could be used for structural study using NMR spectroscopy.

Conclusions

These results provide another strategy for obtaining a single TM domain for structural studies without using toxic chemical digestion or acid to remove the fusion tag. The purified TM domain of p75 neurotrophin receptor will be useful for structural studies.     

Effects of synthetic cohesin-containing scaffold protein architecture on binding dockerin-enzyme fusions on the surface of Lactococcus lactis

Background

In the last years, the biotechnological production of platform chemicals for fuel components has become a major focus of interest. Although ligno-cellulosic material is considered as suitable feedstock, the almost inevitable pretreatment of this recalcitrant material may interfere with the subsequent fermentation steps. In this study, the fungus Ustilago maydis was used to produce itaconic acid as platform chemical for the synthesis of potential biofuels such as 3-methyltetrahydrofuran. No studies, however, have investigated how pretreatment of ligno-cellulosic biomass precisely influences the subsequent fermentation by U. maydis. Thus, this current study aims to first characterize U. maydis in shake flasks and then to evaluate the influence of three exemplary pretreatment methods on the cultivation and itaconic acid production of this fungus. Cellulose enzymatically hydrolysed in seawater and salt-assisted organic-acid catalysed cellulose were investigated as substrates. Lastly, hydrolysed hemicellulose from fractionated beech wood was applied as substrate.

Results

U. maydis was characterized on shake flask level regarding its itaconic acid production on glucose. Nitrogen limitation was shown to be a crucial condition for the production of itaconic acid. For itaconic acid concentrations above 25 g/L, a significant product inhibition was observed. Performing experiments that simulated influences of possible pretreatment methods, U. maydis was only slightly affected by high osmolarities up to 3.5 osmol/L as well as of 0.1 M oxalic acid. The production of itaconic acid was achieved on pretreated cellulose in seawater and on the hydrolysed hemicellulosic fraction of pretreated beech wood.

Conclusion

The fungus U. maydis is a promising producer of itaconic acid, since it grows as single cells (yeast-like) in submerged cultivations and it is extremely robust in high osmotic media and real seawater. Moreover, U. maydis can grow on the hemicellulosic fraction of pretreated beech wood. Thereby, this fungus combines important advantages of yeasts and filamentous fungi. Nevertheless, the biomass pretreatment does indeed affect the subsequent itaconic acid production. Although U. maydis is insusceptible to most possible impurities from pretreatment, high amounts of salts or residues of organic acids can slow microbial growth and decrease the production. Consequently, the pretreatment step needs to fit the prerequisites defined by the actual microorganisms applied for fermentation.     

Clinical features and predictors for disease natural progression in adults with Pompe disease: a nationwide prospective observational study

Background

Due partly to physicians’ unawareness, many adults with Pompe disease are diagnosed with great delay. Besides, it is not well known which factors influence the rate of disease progression, and thus disease outcome. We delineated the specific clinical features of Pompe disease in adults, and mapped out the distribution and severity of muscle weakness, and the sequence of involvement of the individual muscle groups. Furthermore, we defined the natural disease course and identified prognostic factors for disease progression.

Methods

We conducted a single-center, prospective, observational study. Muscle strength (manual muscle testing, and hand-held dynamometry), muscle function (quick motor function test), and pulmonary function (forced vital capacity in sitting and supine positions) were assessed every 3–6 months and analyzed using repeated-measures ANOVA.

Results

Between October 2004 and August 2009, 94 patients aged between 25 and 75 years were included in the study. Although skeletal muscle weakness was typically distributed in a limb-girdle pattern, many patients had unfamiliar features such as ptosis (23%), bulbar weakness (28%), and scapular winging (33%). During follow-up (average 1.6 years, range 0.5-4.2 years), skeletal muscle strength deteriorated significantly (mean declines of ?1.3% point/year for manual muscle testing and of ?2.6% points/year for hand-held dynamometry; both p<0.001). Longer disease duration (>15 years) and pulmonary involvement (forced vital capacity in sitting position <80%) at study entry predicted faster decline. On average, forced vital capacity in supine position deteriorated by 1.3% points per year (p=0.02). Decline in pulmonary function was consistent across subgroups. Ten percent of patients declined unexpectedly fast.

Conclusions

Recognizing patterns of common and less familiar characteristics in adults with Pompe disease facilitates timely diagnosis. Longer disease duration and reduced pulmonary function stand out as predictors of rapid disease progression, and aid in deciding whether to initiate enzyme replacement therapy, or when.

     

Improving efficiency of breeding for higher crop yield

Summary Exclusive selection for yield raises, the harvest index of self-pollinated crops with little or no gain in total bipmass. In addition to selection for yield, it is suggested that efficient breeding for higher yield requires simultaneous selection for yield's three major, genetically controlled physiological components. The following are needed: (1) a superior rate of biomass accumulation. (2) a superior rate of actual yield accumulation in order to acquire a high harvest index, and (3) a time to harvest maturity that is neither shorter nor longer than the duration of the growing season. That duration is provided by the environment, which is the fourth major determinant of yield. Simultaneous selection is required because genetically established interconnections among the three major physiological components cause: (a) a correlation between the harvest index and days to maturity that is usually negative; (b) a correlation between the harvest index and total biomass that is often negative, and (c) a correlation between biomass and days to maturity that is usually positive. All three physiological components and the correlations among them can be quantified by yield system analysis (YSA) of yield trials. An additive main effects and multiplicative interaction (AMMI) statistical analysis can separate and quantify the genotype × environment interaction (G × E) effect on yield and on each physiological component that is caused by each genotype and by the different environment of each yield trial. The use of yield trials to select parents which have the highest rates of accumulation of both biomass and yield, in addition to selecting for the G × E that is specifically adapted to the site can accelerate advance toward the highest potential yield at each geographical site. Higher yield for many sites will raise average regional yield. Higher yield for multiple regions and continents will raise average yield on a world-wide basis. Genetic and physiological bases for lack of indirect selection for biomass from exclusive selection for yield are explained.