Epithelial cell spreading induced by hepatocyte growth factor influences paxillin protein synthesis and posttranslational modification

Superficial wounds in the gastrointestinal tract rapidly reseal by coordinated epithelial cell migration facilitated by cytokines such as hepatocyte growth factor (HGF)/scatter factor released in the wound vicinity. However, the mechanisms by which HGF promotes physiological and pathophysiologic epithelial migration are incompletely understood. Using in vitro models of polarized T84 and Caco-2 intestinal epithelia, we report that HGF promoted epithelial spreading and RhoA GTPase activation in a time-dependent manner. Inducible expression of enhanced green fluorescent protein-tagged dominant-negative RhoA significantly attenuated HGF-induced spreading. HGF expanded a zone of partially flattened cells behind the wound edge containing basal F-actin fibers aligned in the direction of spreading. Concomitantly, plaques positive for the focal adhesion protein paxillin were enhanced. HGF induced an increase in the translation of paxillin and, to a lesser extent, beta1-integrin. This was independent of cell-matrix adhesion through beta1-integrin. Subcellular fractionation revealed increased cosedimentation of paxillin with plasma membrane-containing fractions following HGF stimulation, without corresponding enhancements in paxillin coassociation with beta1 integrin or actin. Tyrosine phosphorylation of paxillin was reduced by HGF and was sensitive to the Src kinase inhibitor PP2. With these taken together, we propose that HGF upregulates a free cytosolic pool of paxillin that is unaffiliated with either the cytoskeleton or focal cell-matrix contacts. Thus early spreading responses to HGF may partly relate to increased paxillin availability for incorporation into, and turnover within, dynamic cytoskeletal/membrane complexes whose rapid and transient adhesion to the matrix drives migration.     

Legacy Effects of Oil Road Reclamation on Soil Biology and Plant Community Composition

Following the unprecedented oil drilling presently occurring in western North Dakota, thousands of kilometers of oil roads must be reclaimed to an acceptable post‐extraction condition. This study assessed the soil biological and plant communities of nine decommissioned oil roads reclaimed during three periods between 1983 and 2002 in the Little Missouri National Grasslands of western North Dakota. We hypothesized that time‐since‐reclamation would positively affect soil biological and plant communities and, consequently, success of reclamation at older sites. To assess this hypothesis, we measured soil enzyme activity, soil microbial community composition, plant community composition, and soil physical and chemical properties along a gradient extending from road‐center to adjacent native prairie for the nine roads. Time‐since‐reclamation did not affect soil and plant properties measured, indicating that older reclamations are not more similar to native prairie than reclamations occurring more recently. A strong gradient between samples from road‐center and native prairie was identified with univariate and ordination analyses, indicating that soil and plant communities of reclaimed oil roads do not resemble those of the surrounding prairie. Soil organic matter (SOM) was identified as the most significantly affected soil property, being 30% lower on reclaimed roads than prairie. The relationship between SOM, microbial community, and plant community suggests that incorporating additional SOM could hasten reclamation as a result of improving the physical environment for plants and providing a labile carbon and energy source for the soil microbial community which, in turn, will enhance the nutrient and physical conditions for plant growth.     

Sensitivity of tropical forests to climate change in the humid tropics of north Queensland

An analysis using an artificial neural network model suggests that the tropical forests of north Queensland are highly sensitive to climate change within the range that is likely to occur in the next 50–100 years. The distribution and extent of environments suitable for 15 structural forest types were estimated, using the model, in 10 climate scenarios that include warming up to 1°C and altered precipitation from –10% to +20%. Large changes in the distribution of forest environments are predicted with even minor climate change. Increased precipitation favours some rainforest types, whereas decreased rainfall increases the area suitable for forests dominated by sclerophyllous genera such as Eucalyptus and Allocasuarina. Rainforest environments respond differentially to increased temperature. The area of lowland mesophyll vine forest environments increases with warming, whereas upland complex notophyll vine forest environments respond either positively or negatively to temperature, depending on precipitation. Highland rainforest environments (simple notophyll and simple microphyll vine fern forests and thickets), the habitat for many of the region’s endemic vertebrates, decrease by 50% with only a 1°C warming. Estimates of the stress to present forests resulting from spatial shifts of forest environments (assuming no change in the present forest distributions) indicate that several forest types would be highly stressed by a 1°C warming and most are sensitive to any change in rainfall. Most forests will experience climates in the near future that are more appropriate to some other structural forest type. Thus, the propensity for ecological change in the region is high and, in the long term, significant shifts in the extent and spatial distribution of forests are likely. A detailed spatial analysis of the sensitivity to climate change indicates that the strongest effects of climate change will be experienced at boundaries between forest classes and in ecotonal communities between rainforest and open woodland.     

(Homo)glutathione deficiency impairs root-knot nematode development in Medicago truncatula

Root-knot nematodes (RKN) are obligatory plant parasitic worms that establish and maintain an intimate relationship with their host plants. During a compatible interaction, RKN induce the redifferentiation of root cells into multinucleate and hypertrophied giant cells essential for nematode growth and reproduction. These metabolically active feeding cells constitute the exclusive source of nutrients for the nematode. Detailed analysis of glutathione (GSH) and homoglutathione (hGSH) metabolism demonstrated the importance of these compounds for the success of nematode infection in Medicago truncatula. We reported quantification of GSH and hGSH and gene expression analysis showing that (h)GSH metabolism in neoformed gall organs differs from that in uninfected roots. Depletion of (h)GSH content impaired nematode egg mass formation and modified the sex ratio. In addition, gene expression and metabolomic analyses showed a substantial modification of starch and γ-aminobutyrate metabolism and of malate and glucose content in (h)GSH-depleted galls. Interestingly, these modifications did not occur in (h)GSH-depleted roots. These various results suggest that (h)GSH have a key role in the regulation of giant cell metabolism. The discovery of these specific plant regulatory elements could lead to the development of new pest management strategies against nematodes.     

Unwanted caesarean sections among public and private patients in Brazil: prospective study

ObjectiveTo assess and compare the preferences of pregnant women in the public and private sector regarding delivery in Brazil.DesignFace to face structured interviews with women who were interviewed early in pregnancy, about one month before the due date, and about one month post partum.SettingFour cities in Brazil.Participants1612 pregnant women: 1093 public patients and 519 private patients.Results1136 women completed all three interviews; 476 women were lost to follow up (376 public patients and 100 private patients). Despite large differences in the rates of caesarean section in the two sectors (222/717 (31%) among public patients and 302/419 (72%) among private patients) there were no significant differences in preferences between the two groups. In both antenatal interviews, 70-80% in both sectors said they would prefer to deliver vaginally. In a large proportion of cases (237/502) caesarean delivery was decided on before admission: 48/207 (23%) in women in the public sector and 189/295 (64%) in women in the private sector.ConclusionsThe large difference in the rates of caesarean sections in women in the public and private sectors is due to more unwanted caesarean sections among private patients rather than to a difference in preferences for delivery. High or rising rates of caesarean sections do not necessarily reflect demand for surgical delivery.

What is already known on this topic

In Brazil, one quarter of all women deliver in the private sectorThe rate of caesarean deliveries in the private sector is extremely high (70%) and more than twice that in the public sector, where rates have recently fallen due to a new policyPrevious studies in which women were interviewed after birth showed that a substantial proportion of private patients who have caesarean sections would have preferred normal delivery

What this study adds

In two antenatal interviews, preferences regarding type of delivery were nearly identical among public and private patients and strongly favoured vaginal birthsContrary to popular belief, middle and upper class women in Brazil do not want to deliver by caesarean section     

Protein synthesis and morphogenesis are not tightly linked during embryogenesis in Fucus

Fertilized eggs of the brown alga Fucus have long been used as model organisms for investigating the early events in the establishment of polarity and subsequent embryogenesis since large numbers of zygotes can easily be obtained. We have analyzed protein synthesis in eggs and embryos during the first day of development using two-dimensional gels and found that synthesis of 12 of the 60 most prominent proteins changed either qualitatively or quantitatively. Actin and beta-tubulin were identified by immunoblotting; synthesis of these cytoskeletal proteins was initiated at different times during the first 12 hr of development. Unique, reproducible patterns of protein synthesis observed during development in the light permitted accurate staging of developing embryos. Inhibitors such as cytochalasin and sucrose, however, blocked morphogenesis without affecting protein synthesis, and, conversely, growth in the dark delayed protein synthesis but had very little effect on the timing of morphogenesis. The data are consistent with morphogenesis and protein synthesis being relatively independent during early embryogenesis. Actinomycin D added soon after fertilization had no effect on protein synthesis 1 day later, indicating that the proteins analyzed were encoded by maternal mRNA stored in the egg.     

Model-independent analysis of the orientation of fluorescent probes with restricted mobility in muscle fibers

The orientation of proteins in ordered biological samples can be investigated using steady-state polarized fluorescence from probes conjugated to the protein. A general limitation of this approach is that the probes typically exhibit rapid orientational motion ("wobble") with respect to the protein backbone. Here we present a method for characterizing the extent of this wobble and for removing its effects from the available information about the static orientational distribution of the probes. The analysis depends on four assumptions: 1) the probe wobble is fast compared with the nanosecond time scale of its excited-state decay; 2) the orientational distributions of the absorption and emission transition dipole moments are cylindrically symmetrical about a common axis c fixed in the protein; 3) protein motions are negligible during the excited-state decay; 4) the distribution of c is cylindrically symmetrical about the director of the experimental sample. In a muscle fiber, the director is the fiber axis, F. All of the information on the orientational order of the probe that is available from measurements of linearly polarized fluorescence is contained in five independent polarized fluorescence intensities measured with excitation and emission polarizers parallel or perpendicular to F and with the propagation axis of the detected fluorescence parallel or perpendicular to that of the excitation. The analysis then yields the average second-rank and fourth-rank order parameters ( and ) of the angular distribution of c relative to F, and and , the average second-rank order parameters of the angular distribution for wobble of the absorption and emission transition dipole moments relative to c. The method can also be applied to other cylindrically ordered systems such as oriented lipid bilayer membranes and to processes slower than fluorescence that may be observed using longer-lived optically excited states.     

Optimization of novel reversible Bruton’s tyrosine kinase inhibitors identified using Tethering-fragment-based screens

Since the approval of ibrutinib for the treatment of B-cell malignancies in 2012, numerous clinical trials have been reported using covalent inhibitors to target Bruton’s tyrosine kinase (BTK) for oncology indications. However, a formidable challenge for the pharmaceutical industry has been the identification of reversible, selective, potent molecules for inhibition of BTK. Herein, we report application of Tethering-fragment-based screens to identify low molecular weight fragments which were further optimized to improve on-target potency and ADME properties leading to the discovery of reversible, selective, potent BTK inhibitors suitable for pre-clinical proof-of-concept studies.     

Vocal communication as a function of differential rearing experiences inPan paniscus: A preliminary report

There is little evidence of vocal learning in nonhuman primates despite the well-documented abilities found in avian species. We describe the vocal repertoire of five bonobos (Pan paniscus), four of which live in a seminatural environment. The fifth subject, Kanzi, has been reared with humans during the course of language training. The data indicated that the four bonobos living in a seminatural environment exhibit a variety of species-typical vocalizations. In addition to producing all the species-typical vocalization, Kanzi produced four structurally unique vocalizations that were not heard among the other subjects. These data suggest that Kanzi has learned vocalizations that are novel due to his unique rearing experience. Discussion is focused on the flexibility of vocal communication and vocal comprehension inPan paniscus.