Endoplasmic reticulum distribution during bovine oocyte activation is regulated by protein kinase C via actin filaments

Fertilization-induced [Ca²⁺]_i oscillations generally depend on the release of calcium ions from the endoplasmic reticulum (ER). Since ER is the main store of calcium ions, it plays an important role in oocyte fertilization. However, the mechanism of ER organization at oocyte activation is unknown. Here, we show that protein kinase C (PKC) is involved in ER distribution during bovine oocyte activation, but not involved in cell cycle resumption and spindle organization. Actin filaments were affected by PKC pharmacological inhibition. In addition, similar to PKC results, the actin-depolymerizing drug cytochalasin B affected the ER distribution during oocyte activation. Specifically, we have demonstrated that ER organization during bovine oocyte activation is regulated by PKC possibly through its action on actin filaments regulation. Taken together, the results presented here provide further information on the pathway involved in the regulation of ER organization during oocyte activation and new insight into the functional role of PKC and actin filaments during this process.     

Improvement of homologous GH10 xylanase production by deletion of genes with predicted function in the Aspergillus nidulans secretion pathway

Filamentous fungi are important cell factories for large-scale enzyme production. However, production levels are often low, and this limitation has stimulated research focusing on the manipulation of genes with predicted function in the protein secretory pathway. This pathway is the major route for the delivery of proteins to the cell exterior, and a positive relationship between the production of recombinant enzymes and the unfolded protein response (UPR) pathway has been observed. In this study, Aspergillus nidulans was exposed to UPR-inducing chemicals and differentially expressed genes were identified by RNA-seq. Twelve target genes were deleted in A. nidulans recombinant strains producing homologous and heterologous GH10 xylanases. The knockout of pbnA (glycosyltransferase), ydjA (Hsp40 co-chaperone), trxA (thioredoxin) and cypA (cyclophilin) improved the production of the homologous xylanase by 78, 171, 105 and 125% respectively. Interestingly, these deletions decreased the overall protein secretion, suggesting that the production of the homologous xylanase was specifically altered. However, the production of the heterologous xylanase and the secretion of total proteins were not altered by deleting the same genes. Considering the results, this approach demonstrated the possibility of rationally increase the production of a homologous enzyme, indicating that trxA, cypA, ydjA and pbnA are involved in protein production by A. nidulans.     

Effect of hydrophobicity degree of polymer particles on lipase immobilization and on biocatalyst performance

Abstract

Many studies describe the advantages of using hydrophobic particles on lipase immobilisation. However, many of these works neglect the effect of other variables of the supports, such as specific area and porosity, on the biocatalyst performance, and do not evaluate the influence of the hydrophobicity level of the particles on the biocatalysts’ activity as a single variable. Thus, the focus of the present work was to evaluate the effect of the hydrophobicity degree of polymeric particles on the biocatalysts’ activities, mitigating the influence of other variables. The study was divided into two steps. Firstly, distinct particles, exhibiting different composition and hydrophobicity levels, were used for the immobilization of a commercial lipase B from Candida antarctica (CAL-B). Then, distinct core-shell polymeric particles presenting different functional compounds on the surface were produced, using as comonomers styrene, divinylbenzene, 1-octene, vinylbenzoate and cardanol. Such particles were subsequently used for CAL-B immobilisation and the performance of the biocatalysts was evaluated on hydrolysis (using p-nitrophenyl laurate, as substrate) and esterification (using ethanol and oleic acid, as substrate) reactions. Based on the screening step, it was observed that for non-porous particles the correlation coefficients between the hydrophobicity level of the supports and the biocatalysts performance, for both hydrolysis and esterification reactions, were very low (0.32 and 0.45, respectively). It highlights that there was no significant correlation between these variables and that, probably, the chemical composition of the polymeric chains affects more significantly the biocatalyst performance. Then, analysing the subsequent stage, it was observed that small changes in the surface composition of the core-shell particles result in significant changes on the textural properties of the supports (specific area ranging from 1.2?m².g^?1 to 18.3?m².g^?1; and contact angles ranging from 71° (hydrophilic particles) to 92° (hydrophobic supports) when polymer films were put in contact with water). Such particles were also employed on CAL-B immobilization and it was noticed that higher correlation coefficients were achieved for hydrolysis (ρ?=?0.53) and esterification (ρ?=?0.74) reactions. Therefore, it is shown that the hydrophobicity degree of such supports starts to affect more effectively the biocatalysts performance when other textural features of the supports become more significant, such as specific area and porosity.     

Climatic factors and population density of Lutzomyia longipalpis (Lutz & Neiva, 1912) in an urban endemic area of visceral leishmaniasis in midwest Brazil

The life cycle of vectors and the reservoirs that participate in the chain of infectious diseases have a strong relationship with the environmental dynamics of the ecosystems in which they live. Oscillations in population abundance and seasonality of insects can be explained by factors inherent in each region and time period. Therefore, knowledge of the relationship and influence of environmental factors on the population of Lutzomyia longipalpis is necessary because of the high incidence of visceral leishmaniasis (VL) in Brazil. This study evaluates the influence of abiotic variables on the population density and seasonal behavior of L. longipalpis in an urban endemic area of VL in Brazil. The sand fly captures were performed every two months between November, 2009 and November, 2010 in the peridomicile of 13 randomly selected residences. We captured 1,367 specimens of L. longipalpis, and the ratio of male/female flies was 2.86:1. The comparison of the total male specimens in the two seasons showed a statistical difference in the wet season, but there was no significant difference when considering the total females. With respect to climatic variables, a significant negative association was observed only with wind speed. During periods of high wind speeds, the population density of this vector decreased. The presence of L. longipalpis was found in all months of the study with bimodal behavior and population peaks during the wet season.     

Cooperative foraging in neotropical pseudoscorpions: effects of prey changes on behavioral adjustments of colonies

The pseudoscorpion Paratemnoides nidificator is a generalist predator that captures large arthropods that live on tree trunks. Few pseudoscorpions species show some degree of sociality. We investigated how colonies of the pseudoscorpion P. nidificator adjust their cooperative capture behavior under a situation of changing prey types as a simulation of variation in prey availability. We hypothesized that colonies would be more efficient at prey capture under repeated exposure to the same prey, and that the change in the availability of prey would be followed by new behavioral adjustments to adequately exploit the new prey. Eight experimental colonies housed in the laboratory received repetitions of three different ant species as prey. The number of pseudoscorpions attacking the prey, the number of behavioral acts, and the time expended subduing prey were evaluated as measures of prey capture performance, in relation to repetitive exposure to the same prey and also in relation to prey type changes. However, only individuals’ recruitment significantly responded to prey type exposure. Prey capture behavior was heterogeneous among colonies, resulting in highly variable behavioral responses. Colonies showed a tendency toward increasing capture success through repeated prey type exposure. However, 50% of the colonies were unable to capture the new prey type and died of starvation. Although it is a generalist predator, prey capture behavior could depend on different coordination components for subduing and handling large prey. Therefore, changes in prey availability could cause the attenuation of a cooperative relationship in some colonies, making them more prone to failure during capture.     

Ecological speciation in Gambusia fishes

Although theory indicates that natural selection can facilitate speciation as a by-product, demonstrating ongoing speciation via this by-product mechanism in nature has proven difficult. We examined morphological, molecular, and behavioral data to investigate ecology's role in incipient speciation for a post-Pleistocene radiation of Bahamas mosquitofish (Gambusia hubbsi) inhabiting blue holes. We show that adaptation to divergent predator regimes is driving ecological speciation as a by-product. Divergence in body shape, coupled with assortative mating for body shape, produces reproductive isolation that is twice as strong between populations inhabiting different predator regimes than between populations that evolved in similar ecological environments. Gathering analogous data on reproductive isolation at the interspecific level in the genus, we find that this mechanism of speciation may have been historically prevalent in Gambusia. These results suggest that speciation in nature can result as a by-product of divergence in ecologically important traits, producing interspecific patterns that persist long after speciation events have completed.     

Variability in tolerance to UV-B radiation among Beauveria spp. isolates

Solar radiation, particularly the UV-B component, negatively affects survival of entomopathogenic fungi in the field. In an effort to identify Beauveria spp. isolates with promise for use in biological control settings with high insolation, we examined 53 Beauveria bassiana isolates, 7 isolates of 4 other Beauveria spp. and Engyodontium albus (=Beauveria alba). The origins of these fungi varied widely as to host/substrate and country, but approximately 30% of these isolates were B. bassiana from ticks in Brazil. A preliminary trial with three B. bassiana isolates (Bb 19, CG 310 and CG 481) at several UV-B dosages indicated that 2h of weighted UV-B irradiance at 978mWm(-2) (providing a total dose of 7.04kJm(-2)) allowed separation of isolates into low, medium or high UV-B tolerance. This dose, therefore, was selected as a single dose to compare UV-B tolerances of all 60 Beauveria spp. isolates. There was high variability in tolerance to UV-B radiation among the B. bassiana isolates, ranging from virtually zero tolerance (e.g., Bb 03) to almost 80% tolerance (e.g., CG 228). In addition, surviving B. bassiana conidia demonstrated delayed germination; and this is likely to reduce virulence. Conidia of the other species were markedly more sensitive to UV-B, with E. albus (UFPE 3138) being the least UV-B tolerant. Among B. bassiana isolates originating from 0 degrees to 22 degrees latitudes, those from lower latitudes demonstrated statistically significant greater UV-B tolerances than those isolates from higher latitudes. Isolates from above 22 degrees , however, were unaffected by latitude of origin. A similar analysis based on host type did not indicate a correlation between original host and UV-B tolerance. The identification in this study of several B. bassiana isolates with relatively high UV-B tolerance will guide the selection of isolates for future arthropod microbial control experiments.     

Visible light during mycelial growth and conidiation of Metarhizium robertsii produces conidia with increased stress tolerance

Light conditions during mycelial growth are known to influence fungi in many ways. The effect of visible-light exposure during mycelial growth was investigated on conidial tolerance to UVB irradiation and wet heat of Metarhizium robertsii, an insect-pathogenic fungus. Two nutrient media and two light regimens were compared. Conidia were produced on (A) potato dextrose agar plus yeast extract medium (PDAY) (A1) under dark conditions or (A2) under continuous visible light (provided by two fluorescent lamps with intensity 5.4 W m(-2)). For comparison, the fungus was also produced on (B) minimal medium (MM) under continuous-dark incubation, which is known to produce conidia with increased tolerance to heat and UVB radiation. The UVB tolerances of conidia produced on PDAY under continuous visible light were twofold higher than conidia produced on PDAY medium under dark conditions, and this elevated UVB tolerance was similar to that of conidia produced on MM in the dark. The heat tolerance of conidia produced under continuous light was, however, similar to that of conidia produced on MM or PDAY in the dark. Conidial yield on PDAY medium was equivalent when the fungus was grown either under continuous-dark or under continuous-light conditions.