6-Amino-4-oxo-1,3-diphenyl-2-thioxo-1,2,3,4-tetrahydropyrimidine-5-carbonyl derivatives as a new class of potent inhibitors of Interleukin-8-induced neutrophil chemotaxis

A series of 6-amino-4-oxo-1,3-diphenyl-2-thioxo-1,2,3,4-tetrahydropyrimidine-5-carbonyl derivatives was synthesized. The compounds demonstrated to be novel, potent and selective inhibitors of Interleukin-8-induced human neutrophil chemotaxis. A SAR study was performed by varying the carbonyl function at position 5 and the chain linked to the amino group at position 6 of the scaffold. All the compounds of the series displayed inhibitory activity at nano- or picomolar concentrations against Interleukin-8-driven migration and no activity against fMLP- and C5a-induced chemotaxis. The binding tests of selected compounds on CXCR1 and CXCR2 receptors were negative. The most potent derivative showed in vivo efficacy in a mouse model of Zymosan-induced peritonitis.     

Can different species of medicinal leeches (Hirudo spp.) interbreed?

Abstract. Since the 18th century, the medicinal leech Hirudo medicinalis has been thought to comprise a single species with several different color morphs, but recently some of these color morphs have been assigned to separate species based on morphology, geographical distribution, and molecular sequence data. This research was aimed at testing the ability of three of these species, H. medicinalis, Hirudo verbana, and Hirudo orientalis, to interbreed. We found that in the laboratory, all three species were able to mate with each other and produce hybrid offspring. This suggests that the reproductive isolation is not strong among these species of the genus Hirudo. However, fewer offspring were produced from interspecific crosses compared with intraspecific crosses. This decrease of fecundity (and in some cases, offspring viability) indicates some degree of reproductive isolation between H. medicinalis, H. verbana, and H. orientalis.     

Pre-steady-state kinetic study of substrate specificity of Escherichia coli formamidopyrimidine--DNA glycosylase

Formamidopyrimidine-DNA glycosylase (Fpg) is responsible for removal of 8-oxoguanine (8-oxoG) and other oxidized purine lesions from DNA and can also excise some oxidatively modified pyrimidines [such as dihydrouracil (DHU)]. Fpg is also specific for a base opposite the lesion, efficiently excising 8-oxoG paired with C but not with A. We have applied stopped-flow kinetics using intrinsic tryptophan fluorescence of the enzyme and fluorescence of 2-aminopurine-labeled DNA to analyze the conformational dynamics of Escherichia coli Fpg during processing of good substrates (8-oxoG.C), poor substrates (8-oxoG.A), and substrates of unclear specificity (such as DHU and 8-oxoG opposite T or G). The analysis of fluorescence traces allows us to conclude that when the enzyme encounters its true substrate, 8-oxoG.C, the complex enters the productive catalytic reaction after approximately 50 ms, partitioning the substrate away from the competing dissociation process, while poor substrates linger in the initial encounter complex for longer. Several intermediate ES complexes were attributed to different structures that exist along the reaction pathway. A likely sequence of events is that the damaged base is first destabilized by the enzyme binding and then everted from DNA, followed by insertion of several amino acid residues into DNA and isomerization of the enzyme into a pre-excision complex. We conclude that rejection of the incorrect substrates occurs mostly at the early stage of formation of the pre-eversion recognition complex, supporting the role of indirect readout in damage recognition.     

Chestnut cultivar diversification process in the Iberian Peninsula, Canary Islands, and Azores

This is a large-scale molecular study based on simple sequence repeat (SSR) loci of the diversification process in chestnut cultivars from Portugal and Spain, from the northern Iberian Peninsula to the Canary Islands and the Azores. A total of 593 grafted chestnut trees (Castanea sativa Mill.) were analysed with 10 SSRs: 292 from Portugal and 301 from Spain. Some of the trees studied were more than 300 years old. Accessions were analysed using a model-based Bayesian procedure to assess the geographical structure and to assign individuals to reconstructed populations based on the SSR genotypes. We found 356 different genotypes with a mean value of clonality of 33% owing to grafting. Mutations accounted for 6%, with hybridization being the main diversification process that can explain the great diversity found. Ten main cultivar groups were detected: four in northern Spain, five in the centre of the Iberian Peninsula, and one in southern Spain related to the centre of the Iberian Peninsula. This work demonstrated that cultivar origin and the diversification process was a combination of clonal propagation of selected seedlings, hybridization, and mutations, which allowed high levels of diversity to be maintained with respect to selected clones for fruit production. Furthermore, seedlings and graft sticks facilitated the transport to new destinations in the colonization process, transporting sometimes more than 3000 km if we consider the Azores and the Canary Islands.     

Mycoplasma and herpesvirus PCR detection in tortoises with rhinitis-stomatitis complex in Spain

Chelonid herpesvirus (ChHV) and mycoplasmal infections cause similar clinical signs in terrestrial tortoises and may be the most important causative agents of rhinitis-stomatitis complex, a common disease in captive tortoises worldwide. Currently, diagnosis of ChHV and Mycoplasma spp. infections is most often based on serologic testing. However, serologic results only detect past exposure, and the specificity of these tests can be reduced due to antigenic cross-reactions with other pathogens. Molecular-based techniques could help to define the causative agent and to better manage infected tortoises. Using polymerase chain reaction, we analyzed 63 tortoises (59 spur-thighed tortoise, Testudo graeca; three Greek tortoise, Testudo ibera; and one Russian tortoise, Agryonemys horsfieldii) with clinical signs of rhinitis-stomatitis complex to identify the causative agent. Molecular evidence of ChHV type I (24%), type II (3%), and Mycoplasma agassizii (6%) infections, as well as coinfection of Mycoplasma-ChHV and both types of ChHV, were detected. Both ChHV and M. agassizii are considered pathogenic in captive tortoises and both are a threat to wild populations. However, neither agent was detected from most of the symptomatic tortoises we evaluated, indicating that other agents could be involved in the rhinitis-stomatitis complex.     

Synthetic prions generated in vitro are similar to a newly identified subpopulation of PrPSc from sporadic Creutzfeldt-Jakob Disease

In recent studies, the amyloid form of recombinant prion protein (PrP) encompassing residues 89-230 (rPrP 89-230) produced in vitro induced transmissible prion disease in mice. These studies showed that unlike "classical" PrP(Sc) produced in vivo, the amyloid fibrils generated in vitro were more proteinase-K sensitive. Here we demonstrate that the amyloid form contains a proteinase K-resistant core composed only of residues 152/153-230 and 162-230. The PK-resistant fragments of the amyloid form are similar to those observed upon PK digestion of a minor subpopulation of PrP(Sc) recently identified in patients with sporadic Creutzfeldt-Jakob disease (CJD). Remarkably, this core is sufficient for self-propagating activity in vitro and preserves a beta-sheet-rich fibrillar structure. Full-length recombinant PrP 23-230, however, generates two subpopulations of amyloid in vitro: One is similar to the minor subpopulation of PrP(Sc), and the other to classical PrP(Sc). Since no cellular factors or templates were used for generation of the amyloid fibrils in vitro, we speculate that formation of the subpopulation of PrP(Sc) with a short PK-resistant C-terminal region reflects an intrinsic property of PrP rather than the influence of cellular environments and/or cofactors. Our work significantly increases our understanding of the biochemical nature of prion infectious agents and provides a fundamental insight into the mechanisms of prions biogenesis.     

MicroRNA‐guided gene expression in prostate cancer: Literature and database overview

Insight into the aberrant expression of microRNAs (miRNAs) and the genes that they regulate during the progression of cancer in general and prostate cancer (PCa) in particular is one of the most important issues in current molecular biomedicine and allows for the discovery of therapeutic or diagnostic miRNA targets. The present study aimed to analyze the available data regarding the direct or indirect effects of miRNAs on the expression of the mRNAs involved in carcinogenesis and to enable updating and optimizing the selection of the corresponding targets. The present review focuses on the data related to the genes with miRNA‐dependent expression during the development of PCa. The data used in this review have been extracted from research papers and the databases STRING, PANTHER and TargetScan, with a special focus on the genes directly associated with cell transformation and the maintenance of the transformed genotype, as well as tumor invasion and spread. The search for miRNA markers of PCa and therapeutically active molecules should rely on bioinformatics resources, such as data from recent experimental studies, as well as meta‐analysis and cross‐analysis of the data on the state of the tumor, patient status, histological/immunohistological data and data on mRNA–miRNA coexpression.     

Biological activities of human interferon and 2′–5′ oligoadenylates in plants

Exogenous human interferon 2 (IFN) and 2–5 oligoadenylates (2–5A) have been shown to cause at least a dual physiological effect in tobacco and wheat: (i) increased cytokinin activity and (ii) induced synthesis of numerous proteins, among which members of two groups of stress proteins have been identified, namely pathogenesis-related (PR) and heat shock (HS) proteins. These effects were observed only by low concentrations of these substances: IFN at 0.1–1 u/ml and 2–5A at 1–10 nM.     

Type III secretion and effectors shape the survival and growth pattern of Pseudomonas syringae on leaf surfaces

The bacterium Pseudomonas syringae pv syringae B728a (PsyB728a) uses a type III secretion system (T3SS) to inject effector proteins into plant cells, a process that modulates the susceptibility of different plants to infection. Analysis of GREEN FLUORESCENT PROTEIN-expressing PsyB728a after spray inoculation without additives under moderate relative humidity conditions permitted (1) a detailed analysis of this strain's survival and growth pattern on host (Nicotiana benthamiana) and nonhost (tomato [Solanum lycopersicum]) leaf surfaces, (2) an assessment of the role of plant defenses in affecting PsyB728a leaf surface (epiphytic) growth, and (3) the contribution of the T3SS and specific effectors to PsyB728a epiphytic survival and growth. On host leaf surfaces, PsyB728a cells initially persist without growing, and show an increased population only after 48 h, unless plants are pretreated with the defense-inducing chemical benzothiazole. During the persistence period, some PsyB728a cells induce a T3SS reporter, whereas a T3SS-deficient mutant shows reduced survival. By 72 h, rare invasion by PsyB728a to the mesophyll region of host leaves occurs, but endophytic and epiphytic bacterial growths are not correlated. The effectors HopZ3 and HopAA1 delay the onset of epiphytic growth of PsyB728a on N. benthamiana, whereas they promote epiphytic survival/growth on tomato. These effectors localize to distinct sites in plant cells and likely have different mechanisms of action. HopZ3 may enzymatically modify host targets, as it requires residues important for the catalytic activity of other proteins in its family of proteases. Thus, the T3SS, HopAA1, HopZ3, and plant defenses strongly influence epiphytic survival and/or growth of PsyB728a.