Comparative Analysis of the Tyr-Kinases CapB1 and CapB2 Fused to Their Cognate Modulators CapA1 and CapA2 from Staphylococcus aureus

A particular class of tyrosine-kinases sharing no structural similarity with eukaryotic tyrosine-kinases has been evidenced in a large array of bacterial species. These bacterial tyrosine-kinases are able to autophosphorylate on a C-terminal tyrosine-rich motif. Their autophosphorylation has been shown to play a crucial role in the biosynthesis or export of capsular polysaccharide. The analysis of the first crystal structure of the staphylococcal tyrosine kinase CapB2 associated with the activating domain of the transmembrane modulator CapA1 had brought conclusive explanation for both the autophosphorylation and activation processes. In order to explain why CapA1 activates CapB2 more efficiently than its cognate transmembrane modulator CapA2, we solved the crystal structure of CapA2B2 and compared it with the previously published structure of CapA1B2. This structural analysis did not provide the expected clues about the activation discrepancy observed between the two modulators. Staphylococcus aureus also encodes for a CapB2 homologue named CapB1 displaying more than 70% sequence similarity and being surprisingly nearly unable to autophosphorylate. We solved the crystal structure of CapA1B1 and carefully compare it with the structure of CapA1B2. The active sites of both proteins are highly conserved and the biochemical characterization of mutant proteins engineered to test the importance of small structural discrepancies identified between the two structures did not explain the inactivity of CapB1. We thus tested if CapB1 could phosphorylate other protein substrates or hydrolyze ATP. However, no activity could be detected in our in vitro assays. Taken together, these data question about the biological role of the homologous protein pairs CapA1/CapB1 and CapA2/CapB2 and we discuss about several possible interpretations.     

Geldanamycin-Induced Osteosarcoma Cell Death Is Associated with Hyperacetylation and Loss of Mitochondrial Pool of Heat Shock Protein 60 (Hsp60)

Osteosarcoma is one of the most malignant tumors of childhood and adolescence that is often resistant to standard chemo- and radio-therapy. Geldanamycin and geldanamycin analogs have been recently studied as potential anticancer agents for osteosarcoma treatment. Here, for the first time, we have presented novel anticancer mechanisms of geldanamycin biological activity. Moreover, we demonstrated an association between the effects of geldanamycin on the major heat shock proteins (HSPs) and the overall survival of highly metastatic human osteosarcoma 143B cells. We demonstrated that the treatment of 143B cells with geldanamycin caused a subsequent upregulation of cytoplasmic Hsp90 and Hsp70 whose activity is at least partly responsible for cancer development and drug resistance. On the other hand, geldanamycin induced upregulation of Hsp60 gene expression, and a simultaneous loss of hyperacetylated Hsp60 mitochondrial protein pool resulting in decreased viability and augmented cancer cell death. Hyperacetylation of Hsp60 seems to be associated with anticancer activity of geldanamycin. In light of the fact that mitochondrial dysfunction plays a critical role in the apoptotic signaling pathway, the presented data may support a hypothesis that Hsp60 can be another functional part of mitochondria-related acetylome being a potential target for developing novel anticancer strategies.     

Modelling the potential distribution of Mesalina watsonana (Stoliczka, 1872) (Reptilia: Lacertidae) on the Iranian Plateau

The Persian Long-tailed Desert Lizard, Mesalina watsonana, is one of the most common and most widely distributed lizards on the Iranian Plateau extending from Iran to Pakistan and Afghanistan. The species is frequently encountered in various types of habitats. We collected over 600 distributional records from available literature, museum collections, and our own field work and used bioclimatic and land cover characteristics to develop a model of potential distribution for M. watsonana. According to the model, the most important factors limiting the distribution of M. watsonana are: precipitation in wettest quarter exceeding 250–300 mm, precipitation in coldest quarter lower than 40 mm and exceeding 250 mm, altitudes above 2500 m and slopes steeper than 10.5°. The model suggests that most of the Iranian Plateau is suitable for the species except for some isolated areas such as the Dasht-e Kavir and Dasht-e Lut deserts in Iran, Helmand basin in Afghanistan, the Karakum Desert in Turkmenistan, the western Chagai-Kharan deserts of Pakistani Balochistan, and Thar and Cholistan deserts in eastern Pakistan. The most important factor in these regions appears to be the extremely low rainfall during coldest quarter of the year. The outer boundary of the distribution of M. watsonana follows important biogeographic barriers that are also clearly delimited by climatic conditions.     

A Selective Method of Estimation of Bacitracin in the Presence of Other Antibiotics and Growth Factors

The plum cankerworm moth, Cystidia couaggaria couaggaria (Geometridae: Ennominae), is a defoliator of Chinese plum trees (Prunus mume). The pheromone components of the female were analyzed by gas chromatography (GC) with an electro-antennographic (EAG) detector and GC coupled with mass spectrometry. The crude pheromone extract included several EAG-active components, i.e., trienyl, dienyl, and saturated hydrocarbons, with a C₂₁–C₂₅ straight chain. The characteristic mass spectra indicated the unsaturated hydrocarbons to be (3Z,6Z,9Z)-3,6,9-trienes and (6Z,9Z)-6,9-dienes. In the fields, mixtures of the synthetic C₂₁ and C₂₃ trienes in a ratio of 2:3 and 1:4 successfully attracted males of this diurnal species during daytime. While the male antennae responded to the C₂₅ triene and saturated hydrocarbons, their synergistic effects were not observed on the male attraction in the fields. Addition of the C₂₁ diene interestingly inhibited the activity of the triene mixture. Males of Cystidia truncangulata, a sympatric diurnal congener of C. c. couaggaria, showed similar EAG responses to the unsaturated hydrocarbons, but no C. truncangulata males were attracted by the lures tested for C. c. couaggaria males, indicating that the identified hydrocarbons comprised the species-specific pheromone of C. c. couaggaria females.     

Dry calcareous grasslands from two neighboring biogeographic regions: relationship between plant traits and rarity

European dry grasslands formed of the Festuco-Brometea type are among the most diverse plant communities within agricultural landscapes. We examined floristic composition, functional trait structure and threatened species occurrence in grasslands of this type from in two distinct biogeographic regions, Dinaric (NW Balkan) and the Central European. In the Central European region this type of grassland is threatened by a decline in traditional extensive management. We tested, if differences in the level of threat between regions are matched by differences in community-weighted means for resource use strategy traits and occurrence of Red List (threatened) species. We then tested if threatened species differed in their traits from other species and if threatened species richness was related to CWM for resource use strategy traits. The communities from the Central European region had significantly higher SLA and lower LDMC, which perhaps reflects an increase in intensive agriculture promoting fast-growing species. Threatened species occurrence did not differ significantly between regions, but threatened species richness was significantly negatively correlated with CWM for height and SLA. This may suggest that threatened species are less likely to occur in high productivity communities where light competition is intense. This study provides initial evidence that changes in CWM for resource use strategy traits may provide a useful means for predicting threatened species loss in dry grassland ecosystems.     

Biological activity of Pd(II) complexes with mono- and disubstituted pyridines—Experimental and theoretical studies

Due to the similarity between Pd and Pt, the complexes of palladium(II) can be considered as potential anticancer agents. Activity of six PdCl₂(X₂Py)₂ complexes (Py = pyridine, and X = CH₃ or Cl) was measured by MTT test using MCF7, CCRF-SB, PC3 and human B-lymphoblastoid cell lines. We found that the effect of PdCl₂(X_nPy)₂ was cell-specific and time-dependent. Obtained results were discussed and compared with the activation parameters calculated for the hydrolysis of PdCl₂(X_nPy)₂, indicating a correlation between viability of MCF7 and CCRF-SB cells and rates of hydrolysis of the Pd(II) complexes.     

Preparation and Standardisation of Smoke-Water for Seed Germination and Plant Growth Stimulation

Smoke-water (SW) has a positive effect on seed germination in many plant species and its application in various fields of plant science has become popular. The method of preparing SW is relatively easy and inexpensive. However, many researchers working in the field of smoke biology are still unfamiliar with preparing SW and testing it for bioactivity. Thus, the aim of this study was to develop a low-cost efficient apparatus to produce SW using grasses, its standardisation and quantification of major active biomolecules. The prepared crude SW was diluted with distilled water (SW:DW) to 75:25, 50:50 and 25:75 (v/v) and these sub-dilutions were further diluted to 1:500; 1:1000; 1:1500; 1:2000; 1:2500; 1:3000 and 1:3500 (v/v). For the standardisation of SW, lettuce seeds (Lactuca sativa L. cv. Grand Rapids) were tested to determine the best working concentration. The ratio 1:2500 (v/v) from the sub-dilution 25:75 (v/v) of SW was the best for germination of lettuce seeds in the dark, achieving 91% germination against the water control that had only 7–10% germination. This is the first study which reports the levels of stimulatory (karrikinolide 1, KAR₁ and karrikinolide 2, KAR₂) and inhibitory (trimethylbutenolide, TMB) compounds present in SW using ultrahigh-performance liquid chromatography–electrospray positive ionisation tandem mass spectrometry (UHPLC-ESI(+)-MS/MS).

     

The Staphylococcus aureus ArlRS two-component system regulates virulence factor expression through MgrA

The Gram-positive bacterium, Staphylococcus aureus, is a versatile pathogen that can sense and adapt to a wide variety of environments within the human host, in part through its 16 two-component regulatory systems. The ArlRS two-component system has been shown to affect many cellular processes in S. aureus, including autolysis, biofilm formation, capsule synthesis and virulence. Yet the molecular details of this regulation remained largely unknown. We used RNA sequencing to identify the ArlRS regulon, and found 70% overlap with that of the global regulator MgrA. These genes included cell wall-anchored adhesins (ebh, sdrD), polysaccharide and capsule synthesis genes, cell wall remodeling genes (lytN, ddh), the urease operon, genes involved in metal transport (feoA, mntH, sirA), anaerobic metabolism genes (adhE, pflA, nrdDG) and a large number of virulence factors (lukSF, lukAB, nuc, gehB, norB, chs, scn and esxA). We show that ArlR directly activates expression of mgrA and identify a probable ArlR-binding site (TTTTCTCAT-N₄-TTTTAATAA). A highly similar sequence is also found in the spx P2 promoter, which was recently shown to be regulated by ArlRS. We also demonstrate that ArlS has kinase activity toward ArlR in vitro, although it has slower kinetics than other similar histidine kinases.     

Cardiac disease in mucopolysaccharidosis type I attributed to catecholaminergic and hemodynamic deficiencies

Cardiac dysfunction is a common cause of death among pediatric patients with mutations in the lysosomal hydrolase α-l-iduronidase (IDUA) gene, which causes mucopolysaccharidosis type I (MPS-I). The purpose of this study was to analyze adrenergic regulation of cardiac hemodynamic function in MPS-I. An analysis of murine heart function was performed using conductance micromanometry to assess in vivo cardiac hemodynamics. Although MPS-I (IDUA(-/-)) mice were able to maintain normal cardiac output and ejection fraction at baseline, this cohort had significantly compromised systolic and diastolic function compared with IDUA(+/-) control mice. During dobutamine infusion MPS-I mice did not significantly increase cardiac output from baseline, indicative of blunted cardiac reserve. Autonomic tone, measured functionally by β-blockade, indicated that MPS-I mice required catecholaminergic stimulation to maintain baseline hemodynamics. Survival analysis showed mortality only among MPS-I mice. Linear regression analysis revealed that heightened end-systolic volume in the resting heart is significantly correlated with susceptibility to mortality in MPS-I hearts. This study reveals that cardiac remodeling in the pathology of MPS-I involves heightened adrenergic tone at the expense of cardiac reserve with cardiac decompensation predicted on the basis of increased baseline systolic volumes.