Exploring the role of microRNAs in axolotl regeneration

The axolotl, Ambystoma mexicanum, is used extensively for research in developmental biology, particularly for its ability to regenerate and restore lost organs, including in the nervous system, to full functionality. Regeneration in mammals typically depends on the healing process and scar formation with limited replacement of lost tissue. Other organisms, such as spiny mice (Acomys cahirinus), salamanders, and zebrafish, are able to regenerate some damaged body components. Blastema is a tissue that is formed after tissue injury in such organisms and is composed of progenitor cells or dedifferentiated cells that differentiate into various cell types during regeneration. Thus, identifying the molecules responsible for initiation of blastema formation is an important aspect for understanding regeneration. Introns, a major source of noncoding RNAs (ncRNAs), have characteristic sizes in the axolotl, particularly in genes associated with development. These ncRNAs, particularly microRNAs (miRNAs), exhibit dynamic regulation during regeneration. These miRNAs play an essential role in timing and control of gene expression to order and organize processes necessary for blastema creation. Master keys or molecules that underlie the remarkable regenerative abilities of the axolotl remain to be fully explored and exploited. Further and ongoing research on regeneration promises new knowledge that may allow improved repair and renewal of human tissues.     

Potent inhibitors of malarial P. Falciparum protein kinase G: Improving the cell activity of a series of imidazopyridines

Development of a class of bicyclic inhibitors of the Plasmodium falciparum cyclic GMP-dependent protein kinase (PfPKG), starting from known compounds with activity against a related parasite PKG orthologue, is reported. Examination of key sub-structural elements led to new compounds with good levels of inhibitory activity against the recombinant kinase and in vitro activity against the parasite. Key examples were shown to possess encouraging in vitro ADME properties, and computational analysis provided valuable insight into the origins of the observed activity profiles.     

Spatial distribution of leaf nitrogen and photosynthetic capacity within the foliage of individual trees: disentangling the effects of local light quality,leaf irradiance,and transpiration

There is presently no consensus about the factor(s) driving photosynthetic acclimation and the intra-canopy distribution of leaf characteristics under natural conditions. The impact was tested of local (i) light quality (red/far red ratio), (ii) leaf irradiance (PPFD(i)), and (iii) transpiration rate (E) on total non-structural carbohydrates per leaf area (TNC(a)), TNC-free leaf mass-to-area ratio (LMA), total leaf nitrogen per leaf area (N(a)), photosynthetic capacity (maximum carboxylation rate and light-saturated electron transport rate), and leaf N partitioning between carboxylation and bioenergetics within the foliage of young walnut trees grown outdoors. Light environment (quantity and quality) was controlled by placing individual branches under neutral or green screens during spring growth, and air vapour pressure deficit (VPD) was prescribed and leaf transpiration and photosynthesis measured at branch level by a branch bag technique. Under similar levels of leaf irradiance, low air vapour pressure deficit decreased transpiration rate but did not influence leaf characteristics. Close linear relationships were detected between leaf irradiance and leaf N(a), LMA or photosynthetic capacity, and low R/FR ratio decreased leaf N(a), LMA and photosynthetic capacity. Irradiance and R/FR also influenced the partitioning of leaf nitrogen into carboxylation and electron light transport. Thus, local light level and quality are the major factors driving photosynthetic acclimation and intra-canopy distribution of leaf characteristics, whereas local transpiration rate is of less importance.     

RNase P: role of distinct protein cofactors in tRNA substrate recognition and RNA-based catalysis

The Escherichia coli ribonuclease P (RNase P) has a protein component, termed C5, which acts as a cofactor for the catalytic M1 RNA subunit that processes the 5′ leader sequence of precursor tRNA. Rpp29, a conserved protein subunit of human RNase P, can substitute for C5 protein in reconstitution assays of M1 RNA activity. To better understand the role of the former protein, we compare the mode of action of Rpp29 to that of the C5 protein in activation of M1 RNA. Enzyme kinetic analyses reveal that complexes of M1 RNA–Rpp29 and M1 RNA–C5 exhibit comparable binding affinities to precursor tRNA but different catalytic efficiencies. High concentrations of substrate impede the activity of the former complex. Rpp29 itself exhibits high affinity in substrate binding, which seems to reduce the catalytic efficiency of the reconstituted ribonucleoprotein. Rpp29 has a conserved C-terminal domain with an Sm-like fold that mediates interaction with M1 RNA and precursor tRNA and can activate M1 RNA. The results suggest that distinct protein folds in two unrelated protein cofactors can facilitate transition from RNA- to ribonucleoprotein-based catalysis by RNase P.     

Assessment of DNA damage in glue sniffers by use of the alkaline comet assay

Toluene is used widely, not only in industry, but also in households where toluene exposure and abuse can occur. To estimate the genotoxic risk of toluene exposure, DNA damage was determined in peripheral lymphocytes of 20 glue sniffers and 20 age-matched controls by use of the alkaline comet assay. Urinary hippuric acid and o-cresol excretion rates, which are used as a marker for toluene exposure, were also measured in sniffers and compared with historical control values. The increase in genetic damage in sniffers was statistically significant as compared to control subjects (P<0.0001). The mean values of the hippuric acid and o-cresol excretion rate for glue sniffers was 73- and 1582-fold higher, respectively, than in controls and confirms the putative exposure. Education of the general public and efforts to keep adolescents away from volatile solvent-based products, which may lead to a desire of sniffing in the future, would be advisable.     

Auto-align - multi-modality fluorescence microscopy image co-registration

Multi-modality microscopes incorporate multiple microscopy techniques into one module, imaging through a common objective lens. Simultaneous or consecutive image acquisition of a single specimen, using multiple techniques, increases the amount of measurable information available. In order to benefit from each modality, it is necessary to accurately co-register data sets. Intrinsic differences in the image formation process employed by each modality result in images which possess different characteristics. In addition, as a result of using different measurement devices, images often differ in size and can suffer relative geometrical deformations including rotation, scale and translation, making registration a complex problem. Current methods generally rely on manual input and are therefore subject to human error. Here, we present an automated image registration tool for fluorescence microscopy. We show that it successfully registers images obtained via total internal reflection fluorescence (TIRF), or epi-fluorescence, and confocal microscopy. Furthermore, we provide several other applications including channel merging following image acquisition through an emission beam splitter, and lateral stage drift correction. We also discuss areas of membrane trafficking which could benefit from application of Auto-Align. Auto-Align is an essential item in the advanced microscopist's toolbox which can create a synergy of single or multi-modality image data.     

Acute Schistosoma mansoni infection increases susceptibility to systemic SHIV clade C infection in rhesus macaques after mucosal virus exposure

Background

Individuals living in sub-Saharan Africa represent 10% of the world''s population but almost 2/3 of all HIV-1/AIDS cases. The disproportionate HIV-1 infection rates in this region may be linked to helminthic parasite infections that affect many individuals in the developing world. However, the hypothesis that parasite infection increases an individual''s susceptibility to HIV-1 has never been prospectively tested in a relevant in vivo model.

Methodology/Principal Findings

We measured whether pre-existing infection of rhesus monkeys with a parasitic worm would facilitate systemic infection after mucosal AIDS virus exposure. Two groups of animals, one consisting of normal monkeys and the other harboring Schistosoma mansoni, were challenged intrarectally with decreasing doses of R5-tropic clade C simian-human immunodeficiency virus (SHIV-C). Systemic infection occurred in parasitized monkeys at viral doses that remained sub-infectious in normal hosts. In fact, the 50% animal infectious (AID₅₀) SHIV-C dose was 17-fold lower in parasitized animals compared to controls (P<0.001). Coinfected animals also had significantly higher peak viral RNA loads than controls (P<0.001), as well as increased viral replication in CD4⁺ central memory cells (P = 0.03).

Conclusions/Significance

Our data provide the first direct evidence that acute schistosomiasis significantly increases the risk of de novo AIDS virus acquisition, and the magnitude of the effect suggests that control of helminth infections may be a useful public health intervention to help decrease the spread of HIV-1.