Overlapping functions of argonaute proteins in patterning and morphogenesis of Drosophila embryos

Argonaute proteins are essential components of the molecular machinery that drives RNA silencing. In Drosophila, different members of the Argonaute family of proteins have been assigned to distinct RNA silencing pathways. While Ago1 is required for microRNA function, Ago2 is a crucial component of the RNA-induced silencing complex in siRNA-triggered RNA interference. Drosophila Ago2 contains an unusual amino-terminus with two types of imperfect glutamine-rich repeats (GRRs) of unknown function. Here we show that the GRRs of Ago2 are essential for the normal function of the protein. Alleles with reduced numbers of GRRs cause specific disruptions in two morphogenetic processes associated with the midblastula transition: membrane growth and microtubule-based organelle transport. These defects do not appear to result from disruption of siRNA-dependent processes but rather suggest an interference of the mutant Ago2 proteins in an Ago1-dependent pathway. Using loss-of-function alleles, we further demonstrate that Ago1 and Ago2 act in a partially redundant manner to control the expression of the segment-polarity gene wingless in the early embryo. Our findings argue against a strict separation of Ago1 and Ago2 functions and suggest that these proteins act in concert to control key steps of the midblastula transition and of segmental patterning.     

Studies on the CPA cysteine peptidase in the Leishmania infantum genome strain JPCM5

Background  

Visceral leishmaniasis caused by members of the Leishmania donovani complex is often fatal in the absence of treatment. Research has been hampered by the lack of good laboratory models and tools for genetic manipulation. In this study, we have characterised a L. infantum line (JPCM5) that was isolated from a naturally infected dog and then cloned. We found that JPCM5 has attributes that make it an excellent laboratory model; different stages of the parasite life cycle can be studied in vitro, it is accessible to genetic manipulation and it has retained its virulence. Furthermore, the L. infantum JPCM5 genome has now been fully sequenced.     

Polymer-supported ferrocenyl oxazolines for the catalyzed highly enantioselective phenyl transfer to aldehydes

A new chiral MeO-PEG-supported ferrocenyl oxazoline was synthesized and successfully employed in the enantioselective phenyl transfer to aldehydes. The products were obtained in high yields and with excellent enantioselectivities (up to 97% ee). Furthermore, the recovery of the ferrocene was facile, and catalyst efficiency was maintained in subsequent reactions.     

Susceptibility Screening of Hyphae-Forming Fungi with a New, Easy, and Fast Inoculum Preparation Method

In vitro susceptibility testing of clinically important fungi becomes more and more essential due to the rising number of fungal infections in patients with impaired immune system. Existing standardized microbroth dilution methods for in vitro testing of molds (CLSI, EUCAST) are not intended for routine testing. These methods are very time-consuming and dependent on sporulating of hyphomycetes. In this multicentre study, a new (independent of sporulation) inoculum preparation method (containing a mixture of vegetative cells, hyphae, and conidia) was evaluated. Minimal inhibitory concentrations (MIC) of amphotericin B, posaconazole, and voriconazole of 180 molds were determined with two different culture media (YST and RPMI 1640) according to the DIN (Deutsches Institut für Normung) microdilution assay. 24 and 48?h MIC of quality control strains, tested per each test run, prepared with the new inoculum method were in the range of DIN. YST and RPMI 1640 media showed similar MIC distributions for all molds tested. MIC readings at 48 versus 24?h yield 1 log₂ higher MIC values and more than 90?% of the MICs read at 24 and 48?h were within ±2 log₂ dilution. MIC end point reading (log_{2 MIC-RPMI 1640}?log_{2 MIC-YST}) of both media demonstrated a tendency to slightly lower MICs with RPMI 1640 medium. This study reports the results of a new, time–saving, and easy-to-perform method for inoculum preparation for routine susceptibility testing that can be applied for all types of spore-/non-spore and hyphae-forming fungi.     

The additional value of TGFβ1 and IL-7 to predict the course of prostate cancer progression

Background

Given the fact that prostate cancer incidence will increase in the coming years, new prognostic biomarkers are needed with regard to the biological aggressiveness of the prostate cancer diagnosed. Since cytokines have been associated with the biology of cancer and its prognosis, we determined whether transforming growth factor beta 1 (TGFβ1), interleukin-7 (IL-7) receptor and IL-7 levels add additional prognostic information with regard to prostate cancer-specific survival.

Materials and methods

Retrospective survival analysis of forty-four prostate cancer patients, that underwent radical prostatectomy, was performed (1989–2001). Age, Gleason score and pre-treatment PSA levels were collected. IL-7, IL-7 receptor and TGFβ1 levels in prostate cancer tissue were determined by quantitative real-time RT-PCR and their additional prognostic value analyzed with regard to prostate cancer survival. Hazard ratios and their confidence intervals were estimated, and Akaike’s information criterion was calculated for model comparison.

Results

The predictive ability of a model for prostate cancer survival more than doubled when TGFβ1 and IL-7 were added to a model containing only the Gleason score and pre-treatment PSA (AIC: 18.1 and AIC: 6.5, respectively).

Conclusion

IL-7 and TGFβ1 are promising markers to indicate those at risk for poor prostate cancer survival. This additional information may be of interest with regard to the biological aggressiveness of the diagnosed prostate cancer, especially for those patients screened for prostate cancer and their considered therapy.     

Increased hepatic fibrosis and JNK2-dependent liver injury in mice exhibiting hepatocyte-specific deletion of cFLIP

Chronic liver disease promotes hepatocellular injury involving apoptosis and triggers compensatory regeneration that leads to the activation of quiescent stellate cells in the liver. The deposition of extracellular matrix from activated myofibroblasts promotes hepatic fibrosis and the progression to cirrhosis with deleterious effects on liver physiology. The role of apoptosis signaling pathways in the development of fibrosis remains undefined. The aim of the current study was to determine the involvement of the caspase-8 homologue cellular FLICE-inhibitory protein (cFLIP) during the initiation and progression of fibrosis. Liver injury and fibrosis from carbon tetrachloride (CCl(4)) and thioacetamide (TAA) were examined in mice exhibiting a hepatocyte-specific deletion of cFLIP (flip(-/-)). Acute liver injury from CCl(4) and TAA were enhanced in flip(-/-) mice. This was accompanied by increased activation of caspase-3 and -9, pronounced phosphorylation of JNK, and decreased phosphorylation of Erk. Deletion of the cJun NH(2)-terminal kinase 2 (JNK2) in flip(-/-) mice protected from injury. Hepatic fibrosis was increased at baseline in 12-wk-old flip(-/-) mice, and progression of fibrosis from TAA was accelerated compared with the wild type. In conclusion, deletion of cFLIP in hepatocytes leads to increased fibrosis and accelerated fibrosis progression. This is accompanied by increased injury involving the activation of caspases and JNK2. Thus predisposition to liver injury involving increased hepatocellular apoptosis is a critical mediator of accelerated fibrogenesis, and prevention of liver injury will be a most important measure for patients with chronic liver disease.     

An alternative theoretical approach to escape decision-making: the role of visual cues

Escape enables prey to avoid an approaching predator. The escape decision-making process has traditionally been interpreted using theoretical models that consider ultimate explanations based on the cost/benefit paradigm. Ultimate approaches, however, suffer from inseparable extra-assumptions due to an inability to accurately parameterize the model's variables and their interactive relationships. In this study, we propose a mathematical model that uses intensity of predator-mediated visual stimuli as a basic cue for the escape response. We consider looming stimuli (i.e. expanding retinal image of the moving predator) as a cue to flight initiation distance (FID; distance at which escape begins) of incubating Mallards (Anas platyrhynchos). We then examine the relationship between FID, vegetation cover and directness of predator trajectory, and fit the resultant model to experimental data. As predicted by the model, vegetation concealment and directness of predator trajectory interact, with FID decreasing with increased concealment during a direct approach toward prey, but not during a tangential approach. Thus, we show that a simple proximate expectation, which involves only visual processing of a moving predator, may explain interactive effects of environmental and predator-induced variables on an escape response. We assume that our proximate approach, which offers a plausible and parsimonious explanation for variation in FID, may serve as an evolutionary background for traditional, ultimate explanations and should be incorporated into interpretation of escape behavior.     

Stigmastane derivatives from the roots of Vernonia guineensis and their antimicrobial activity

Chemical investigation of the roots of Vernonia guineensis (Asteraceae) afforded a new stigmastane derivative, vernoguinoside A (1) and the known vernoguinoside (2), stigmasterol 3-O-β-d-glucoside (3) and sitosterol 3-O-β-d-glucoside (4). Their structures were elucidated by spectroscopic analysis. Antimicrobial activities of 1–3 and CH₂Cl₂–MeOH (1:1) extract were evaluated against three bacteria species (Salmonella typhi, Staphylococcus aureus and Shigella flexneri) and three yeasts species (Candida albicans, Candida parapsilosis and Cryptococcus neoformans). Compounds 1 and 2 exhibited both antibacterial and antifungal activities that varied between the microbial species (MIC = 7.81–125 μg/mL) while S. flexneri and C. albicans were sensitive to all the tested compounds.