Decalactone Production by <Emphasis Type="Italic">Yarrowia lipolytica</Emphasis> under increased O<Subscript>2</Subscript> Transfer Rates

Yarrowia lipolytica converts methyl ricinoleate to γ-decalactone, a high-value fruity aroma compound. The highest amount of 3-hydroxy-γ-decalactone produced by the yeast (263 mg l^-1) occurred by increasing the k_La up to 120 h⁻¹ at atmospheric pressure; above it, its concentration decreased, suggesting a predominance of the activity of 3-hydroxyacyl-CoA dehydrogenase. Cultures were grown under high-pressure, i.e., under increased O₂ solubility, but, although growth was accelerated, γ-decalactone production decreased. However, by applying 0.5 MPa during growth and biotransformation gave increased concentrations of dec−2-en-4-olide and dec-3-en-4-olide (70 mg l⁻¹).     

Comparative analysis of tandem repeats from hundreds of species reveals unique insights into centromere evolution

Background

Centromeres are essential for chromosome segregation, yet their DNA sequences evolve rapidly. In most animals and plants that have been studied, centromeres contain megabase-scale arrays of tandem repeats. Despite their importance, very little is known about the degree to which centromere tandem repeats share common properties between different species across different phyla. We used bioinformatic methods to identify high-copy tandem repeats from 282 species using publicly available genomic sequence and our own data.

Results

Our methods are compatible with all current sequencing technologies. Long Pacific Biosciences sequence reads allowed us to find tandem repeat monomers up to 1,419 bp. We assumed that the most abundant tandem repeat is the centromere DNA, which was true for most species whose centromeres have been previously characterized, suggesting this is a general property of genomes. High-copy centromere tandem repeats were found in almost all animal and plant genomes, but repeat monomers were highly variable in sequence composition and length. Furthermore, phylogenetic analysis of sequence homology showed little evidence of sequence conservation beyond approximately 50 million years of divergence. We find that despite an overall lack of sequence conservation, centromere tandem repeats from diverse species showed similar modes of evolution.

Conclusions

While centromere position in most eukaryotes is epigenetically determined, our results indicate that tandem repeats are highly prevalent at centromeres of both animal and plant genomes. This suggests a functional role for such repeats, perhaps in promoting concerted evolution of centromere DNA across chromosomes.     

Involvement of the Dihydropyridine Receptor and Internal CaStores in Myoblast Fusion

The process of myoblast fusion during skeletal myogenesis is calcium regulated. Both dihydropyridine receptor and ryanodine receptor are already present on muscle precursors, at the prefusional stage, before they are required for excitation–contraction coupling. Previous pharmacological studies have shown the need for a special pool of Ca²⁺associated with the membrane for the fusion process to occur. We hypothesized that this pool of Ca²⁺is mobilized via a machinery similar to that involved in excitation–contraction coupling. The process of fusion in rat L6 muscle precursors was either totally or partially abolished in the presence of the L-type calcium channel inhibitors SR33557 and nifedipine (half inhibition towards 2 μM), respectively. The inhibition was reversible and dose-dependent. Drugs able to deplete internal calcium stores (caffeine, ryanodine, and thapsigargin) were also tested on the fusion. Both caffeine and thapsigargin drastically inhibited fusion whereas ryanodine had no effect. This suggests that fusion may be controlled by internal pools of Ca²⁺but that its regulation may be insensitive to ryanodine. We presumed that an early form of the ryanodine receptor may exist, with different pharmacological properties than the adult forms. Indeed, Western blot analysis of pre- and postfusional L6 cells demonstrated the presence, at the prefusional stage, of a transient form of the ryanodine receptor protein with an apparent molecular weight slightly different from those of the classical skeletal and cardiac forms. Taken together, these results support the hypothesis that the fusion process is driven by a mechanism involving both the dihydropyridine receptor (α1 subunit of the L-type Ca²⁺channel) and the internal stores of Ca²⁺. The machinery underlying this mechanism might consist of slightly different forms of the classic molecules that in adult muscle ensure excitation–contraction coupling. It remains to be seen, however, whether the mobilization of the internal pool of Ca²⁺is triggered by the type of mechanism already described in skeletal muscle.     

Reevaluating the classification of Halobacteroides and Haloanaerobacter species based on sequence comparisons of the 16S ribosomal RNA gene

Abstract The 16S rRNA gene (rDNA) sequence analysis of four halophilic anaerobes: Halobacteroides halobius, H. lacunaris, Haloanaerobacter (Hb.) chitinovorans and H. acetoethylicus confirmed that they were all members of the family Haloanaerobiaceae. H. lacunaris and H. halobius were found to be more closely related to each other and were distantly related to Sporohalobacter lortetti and the members of the genera Haloanaerobium and Halothermothrix . These data are in agreement with their assignment to the genus Halobacteroides . Further analysis indicated that Hb. chitinovorans was closely affiliated to members of the genus Halobacteroides , and therefore we propose to transfer it to the genus Halobacteroides as H. chitinovorans comb. nov. This transfer would invalidate the genus Haloanaerobacter , as Hb. chitinovorans is the only member of this genus. The 16S rDNA sequence analysis of H. acetoethylicum indicated that it was very closely related to members of the genus Haloanaerobium , viz. Haloanaerobium (Ha.) praevalens, Ha. salsugo , and Ha. alcaliphilum , and hence we propose to transfer it to the genus Haloanaerobium as Ha. acetoethylicus comb. nov.     

Peptidomic profiling of human cerebrospinal fluid identifies YPRPIHPA as a novel substrate for prolylcarboxypeptidase

Prolylcarboxypeptidase (PRCP) is a serine protease that catalyzes the cleavage of C‐terminal amino acids linked to proline in peptides. It is ubiquitously expressed and is involved in regulating blood pressure, proliferation, inflammation, angiogenesis, and weight maintenance. To identify the candidate proximal target engagement markers for PRCP inhibition in the central nervous system, we profiled the peptidome of human cerebrospinal fluid to look for PRCP substrates using a MS‐based in vitro substrate profiling assay. These experiments identified a single peptide, with the sequence YPRPIHPA, as a novel substrate for PRCP in human cerebrospinal fluid. The peptide YPRPIHPA is from the extracellular portion of human endothelin B receptor‐like protein 2.     

土壤微生物削弱了水生-陆地系统补贴对植物生长的正向影响

土壤微生物削弱了水生-陆地系统补贴对植物生长的正向影响 水生-陆地系统补贴形成的联结作用在构建群落和调节生态系统功能方面发挥重要作用。在营养贫瘠的生态系统中(例如密歇根湖周围的淡水沙丘),水生-陆生系统补贴显得尤为重要。春季成年蠓在密歇根湖涌出,成群交配,然后死亡。蠓尸体在植物的基部形成土丘状,通过输入营养提高植物的生产力。然而,水生-陆地系统补贴对植物生产力的影响可能取决于其他生物的交互作用,特别是土壤微生物可能通过促进养分转化为植物可利用的形式或与植物竞争养分而发挥关键作用。在温室实验中,我们检验了湖生蠓(Chironomidae)的尸体和土壤微生物如何独立和相互影响一种常见沙丘草(沙拂子茅,Calamovilfa longifolia)的生长表现。为确定蠓是否影响土壤非生物特性,我们检验了添加蠓如何影响土壤养分和土壤湿度。研究结果显示,蠓极大地增加了植物生物量,但其效应的大小受土壤微生物的影响。在没有土壤微生物的情况下,添加蠓的植物生物量比没有添加的高7倍,而在有土壤微生物的情况下,植物生物量提高了3倍。蠓对植物生长的促进作用可能由于它们向土壤中输入养分所导致,因为与沙丘土壤相比,蠓的氮、磷、钾含量分别高100倍、10倍和150倍。我们的研究结果表明,土壤微生物可能与植物竞争这些养分。总之,我们发现蠓是重要的水生-陆地系统补贴,对密歇根湖沿岸植物生产力产生强烈和正向的影响,但水生-陆地系统补贴作用必须在生态群落内发生的复杂相互作用的背景下考虑。     

Toxoplasma gondii: comparison of a rhoptry-ELISA with IFAT and MAT for antibody detection in sera of experimentally infected pigs

Indirect ELISA and IFAT have been reported to be more sensitive and specific than agglutination tests. However, MAT is cheaper, easier than the others and does not need special equipment. The purpose of this study was to compare an enzyme linked immunosorbent assay using crude rhoptries of Toxoplasma gondii as coating wells (r-ELISA) with indirect fluorescence antibody test (IFAT) and modified agglutination test (MAT) to detect anti-T. gondii antibodies in sera of experimentally infected pigs. Ten mixed breed pigs between 6.5 and 7.5 weeks old were used. All pigs were negative for the presence of T. gondii antibodies by IFAT (titre < 16), r-ELISA (OD < 0.295) and MAT (titre < 16). Animals received 7x10(7) viable tachyzoites of the RH strain by intramuscular (IM) route at day 0. Serum samples were collected at days -6, 0, 7, 14, 21, 28, 35, 42, 50, and 57. IFAT detected anti-T. gondii antibodies earlier than r-ELISA and MAT. The average of antibody levels was higher at day 35 in IFAT (Log10=2.9) and in MAT (Log10 = 3.5), and at day 42 in r-ELISA (OD = 0.797). The antibody levels remained high through the 57th day after inoculation in MAT, and there was a decrease tendency in r-ELISA and IFAT. IFAT was used as "gold standard" and r-ELISA demonstrated a higher prevalence (73.3%), sensitivity (94.3%), negative predictive value (83.3%), and accuracy (95.6%) than MAT. Kappa agreements among tests were calculated, and the best results were shown by r-ELISAxIFAT (kappa = 0.88, p < 0.001). Cross-reaction with Sarcocystis miescheriana was investigated in r-ELISA and OD mean was 0.163 +/- 0.035 (n = 65). Additionally, none of the animals inoculated with Sarcocystis reacted positively in r-ELISA. Our results indicate that r-ELISA could be a good method for serological detection of T. gondii infection in pigs.     

Heterogeneous synaptic covering and differential charge transfer sensitivity among the dendrites of a reconstructed abducens motor neurone: Correlations between electron microscopic and computer simulation data

Ultrastructural studies on the synaptology of dendritic arborizations of motoneurones have been problematic because dendrites are very thin in relation to their great length, and most of the studies on this topic have therefore dealt with only small parts of the dendritic tree. Here we compared the ultrastructural characteristics of the axon terminals distributed along the various dendrites of a single motoneurone. For this purpose, the light microscopic 3D reconstruction of the dendritic arborization of an intracellularly labelled abducens motoneurone was combined with an electron microscopic analysis of its synaptic contacts. Dendritic profiles were randomly sampled along the various dendrites and the axon terminals they received were classified on the basis of their ultrastructural features and their GABA-immunoreactivity. It emerged that the various dendrites differed according to the type and local arrangement of their synaptic inputs. Our second aim was to incorporate the morphological data obtained into a model giving the charge transfer effectiveness T(x) of the dendritic sites. The sensitivity S(x) of T(x) to changes in the membrane resistivity (Rm) simulating various levels of tonic synaptic activity was calculated. It turned out that both the proximal and distal regions of the dendritic arborization have a dense synaptic covering and a weak sensitivity to changes in the Rm, whereas the intermediate dendrites have a sparse synaptic covering and a high sensitivity to changes in tonic synaptic activity. This pattern of organisation might mediate the “gating” of a population of synapses covering some dendritic regions in a state-dependent fashion.     

Isolation and Characterization of Mercury Resistant Bacillus sp. from Soils with an Extensive History as Substrates for Mercury Extraction in Mexico

Metal phytoextraction assisted by bacteria plays an important role in bioremediation systems. In this work, mercury-resistant bacterial strains were isolated from soils with high levels of mercury (San Joaquin, Queretaro State, Mexico) and identified as Bacillus sp. based on the 16S rDNA gene sequence analysis. The bacterial strains were found to exhibit different multiple mercury-resistance and carbon source utilization characteristics. The mercury reduction ability was tested through a volatilization assay. The bacterial isolates were also evaluated for their ability to promote growth and mercury uptake in tomato plants. In a roll towel assay, the maximum vigor index of tomato plants was obtained with the inoculation of Bacillus sp. A2, A12, B11, B15 and C1, while in a pot assay, the maximum vigor index was obtained with the inoculation of Bacillus sp. A6, A7 and B20, compared with un-inoculated controls in the presence of HgCl₂. Maximum Hg accumulation in the roots and shoots of tomato plants was obtained only with Bacillus sp. A7 in the roll towel assay, whereas in the pot assay, maximum accumulation was obtained with Bacillus sp. A12 compared with un-inoculated controls. Our results show that mercury accumulation in tissue is enhanced by these plant growth promoting bacterial strains, which recommends their possible use as microbe-assisted phytoremediation systems in mercury-polluted soils.