Biological parameters and damage by Macrosoma tipulata Hübner (Lepidoptera: Hedylidae), in cupuaçu tree [Theobroma grandiflorum (Wild ex Spreng Schum)] in Amazonas, Brazil

Biological aspects of the caterpillar Macrosoma tipulata Hübner and its damage to young leaves of Theobroma grandiflorum (Wild ex Spreng Schum) were studied under laboratory conditions of 24.5 +/- 0.7 masculineC, UR 61.6 +/- 9.1% and 12h photophase. The larval stage presented five instars with a total duration of 15.9 +/- 4.31 days, and survival of 98%. The pupal period stage was 7.0 +/- 0.56 days for males and 6.9 +/- 0.56 days for females, with 99% survival for both sex. The pupal mean length was 18.9 +/- 0.89 mm for males and 19.1 +/- 1.13 mm for females. The mean longevity of the adult was 10.3 +/- 2.25 days. The vertical distribution of damage was relatively uniform, with a tendency of greater defoliation in the middle of the treetop, but without significant difference among the three positions.     

Rab27a controls HIV-1 assembly by regulating plasma membrane levels of phosphatidylinositol 4,5-bisphosphate

During the late stages of the HIV-1 replication cycle, the viral polyprotein Pr55^Gag is recruited to the plasma membrane (PM), where it binds phosphatidylinositol 4,5-bisphosphate (PI(4,5)P₂) and directs HIV-1 assembly. We show that Rab27a controls the trafficking of late endosomes carrying phosphatidylinositol 4-kinase type 2 α (PI4KIIα) toward the PM of CD4⁺ T cells. Hence, Rab27a promotes high levels of PM phosphatidylinositol 4-phosphate and the localized production of PI(4,5)P₂, therefore controlling Pr55^Gag membrane association. Rab27a also controls PI(4,5)P₂ levels at the virus-containing compartments of macrophages. By screening Rab27a effectors, we identified that Slp2a, Slp3, and Slac2b are required for the association of Pr55^Gag with the PM and that Slp2a cooperates with Rab27a in the recruitment of PI4KIIα to the PM. We conclude that by directing the trafficking of PI4KIIα-positive endosomes toward the PM, Rab27a controls PI(4,5)P₂ production and, consequently, HIV-1 replication.     

The removal of a disulfide bridge in CotA-laccase changes the slower motion dynamics involved in copper binding but has no effect on the thermodynamic stability

The contribution of the disulfide bridge in CotA-laccase from Bacillus subtilis is assessed with respect to the enzyme’s functional and structural properties. The removal of the disulfide bond by site-directed mutagenesis, creating the C322A mutant, does not affect the spectroscopic or catalytic properties and, surprisingly, neither the long-term nor the thermodynamic stability parameters of the enzyme. Furthermore, the crystal structure of the C322A mutant indicates that the overall structure is essentially the same as that of the wild type, with only slight alterations evident in the immediate proximity of the mutation. In the mutant enzyme, the loop containing the C322 residue becomes less ordered, suggesting perturbations to the substrate binding pocket. Despite the wild type and the C322A mutant showing similar thermodynamic stability in equilibrium, the holo or apo forms of the mutant unfold at faster rates than the wild-type enzyme. The picosecond to nanosecond time range dynamics of the mutant enzyme was not affected as shown by acrylamide collisional fluorescence quenching analysis. Interestingly, copper uptake or copper release as measured by the stopped-flow technique also occurs more rapidly in the C322A mutant than in the wild-type enzyme. Overall the structural and kinetic data presented here suggest that the disulfide bridge in CotA-laccase contributes to the conformational dynamics of the protein on the microsecond to millisecond timescale, with implications for the rates of copper incorporation into and release from the catalytic centres.     

Biocellulose membranes as supports for dermal release of lidocaine

Biocellulose (BC) is a highly pure form of cellulose, produced in the form of a swollen membrane, with several applications in the biomedical area. In this study, the behavior of BC membranes as systems for topical delivery of lidocaine was evaluated. The BC-lidocaine membranes were prepared and characterized in terms of structural and morphological properties. A uniform distribution of the drug inside the BC membranes was observed. In vitro diffusion studies with Franz cells were conducted using human epidermal membranes and showed that the permeation rate of the drug in BC membranes was slightly slower than that obtained with the conventional systems, which was attributed to the establishment of interactions between the lidocaine molecules and the BC membrane, as evidenced by FTIR and NMR analysis. These results indicate that this methodology can be successfully applied for the dermal administration of lidocaine regarding the release profile and ease of application.     

Reactive Oxygen Species and Cellular Interactions Between Mycosphaerella fijiensis and Banana

Globally, the banana plant (Musa spp) is the fourth most important crop after rice, wheat and corn (based on production in tons). It is cultivated in more than 100 tropical and subtropical countries, mainly by small producers and is a fundamental food source for millions of people. Black leaf streak disease (BLSD), caused by Mycosphaerella fijiensis Morelet (sexual phase) or Paracercospora fijiensis (Morelet) Deighton (asexual phase), is the main disease affecting the world??s banana culture. This disease has a wide geographical distribution accounting for losses exceeding 50% of global banana production. We conducted a comparative histocytological study on the kinetics of the infection process using three banana genotypes with phenotypes that differ in resistance to BLSD: Grand Naine (Susceptible), Pisang Madu (Partially Resistant) and Calcutta 4 (Resistant). Experiments were conducted under controlled conditions with the objective of characterizing the cellular interaction processes between M. fijiensis and Musa acuminata. Conidia germination occurred 24 hours after inoculation. Germination rates were high (97%) and there were no significant differences between the three genotypes (P?>?0.147). The Peroxidase enzyme and H₂O₂ were associated with a hypersensitivity-like reaction in the resistant genotype Calcutta 4, indicating a possible role of the enzyme or its product as defense mechanisms against M. fijiensis in banana plants.     

Human NK cells differ more in their KIR2DL1-dependent thresholds for HLA-Cw6-mediated inhibition than in their maximal killing capacity

In this study we have addressed the question of how activation and inhibition of human NK cells is regulated by the expression level of MHC class I protein on target cells. Using target cell transfectants sorted to stably express different levels of the MHC class I protein HLA-Cw6, we show that induction of degranulation and that of IFN-γ secretion are not correlated. In contrast, the inhibition of these two processes by MHC class-I occurs at the same level of class I MHC protein. Primary human NK cell clones were found to differ in the amount of target MHC class I protein required for their inhibition, rather than in their maximum killing capacity. Importantly, we show that KIR2DL1 expression determines the thresholds (in terms of MHC I protein levels) required for NK cell inhibition, while the expression of other receptors such as LIR1 is less important. Furthermore, using mathematical models to explore the dynamics of target cell killing, we found that the observed delay in target cell killing is exhibited by a model in which NK cells require some activation or priming, such that each cell can lyse a target cell only after being activated by a first encounter with the same or a different target cell, but not by models which lack this feature.