Adjustments of water use efficiency by stomatal regulation during drought and recovery in the drought-adapted Vitis hybrid Richter-110 (V. berlandieri x V. rupestris)

The hybrid Richter-110 (Vitis berlandieri x Vitis rupestris) (R-110) has the reputation of being a genotype strongly adapted to drought. A study was performed with plants of R-110 subjected to water withholding followed by re-watering. The goal was to analyze how stomatal conductance (g(s)) is regulated with respect to different physiological variables under water stress and recovery, as well as how water stress affects adjustments of water use efficiency (WUE) at the leaf level. Water stress induced a substantial stomatal closure and an increase in WUE, which persisted many days after re-watering. The g(s) during water stress was mainly related to the content of ABA in the xylem and partly related to plant hydraulic conductivity but not to leaf water potential. By contrast, low g(s) during re-watering did not correlate with ABA contents and was only related to a sustained decreased hydraulic conductivity. In addition to a complex physiological regulation of stomatal closure, g(s) and rate of transpiration (E) were strongly affected by leaf-to-air vapor pressure deficit (VPD) in a way dependent of the treatment. Interestingly, E increased with increasing VPD in control plants, but decreased with increasing VPD in severely stressed plants. All together, the fine stomatal regulation in R-110 resulted in very high WUE at the leaf level. This genotype is revealed to be very interesting for further studies on the physiological mechanisms leading to regulation of stomatal responsiveness and WUE in response to drought.     

Backbone dynamics and hydrogen exchange of Pseudomonas aeruginosa ferricytochrome c 551

A model-free analysis of Pseudomonas aeruginosa ferricytochrome c(551) dynamics based on (15)N R(1), (15)N R(2), and [(1)H]-(15)N heteronuclear nuclear Overhauser effect data is reported. The protein backbone is highly rigid (< S(2)>=0.924+/-0.005) and displays little variation in picosecond-nanosecond time scale dynamics over the structure. The loop structure containing the axial methionine ligand (loop 3) displays anomalous rigidity, which is attributed to its high proline content. Also reported are protection factors calculated from hydrogen-exchange rates. These data reveal that loop 3 residues, including the axial methionine, are protected from exchange as a result of long-range hydrogen-bonding interactions. These results are contrasted with data reported for Saccharomyces cerevisiae iso-1-ferricytochrome c, which displays higher overall flexibility (< S(2)>=0.80+/-0.07), greater variation of dynamics as a function of structure, and low protection factors for loop 3. This analysis reveals that heme proteins with similar functions and topologies may display diverse dynamical properties.     

Genetic Variability in European Populations of an Invasive American Crayfish: Preliminary Results

Since 1973, the red swamp crayfish, Procambarus clarkii, native to south-central United States and northeastern Mexico, has spread throughout Europe. Here, we surveyed the genetic variability of five European populations of the species using RAPD markers. Genetic variation was found to be so high as to uniquely fingerprint most of the surveyed individuals. Analysis of molecular variance (AMOVA) of the RAPD markers showed that 1) a large part of the genetic variation can be attributed to the differentiation among localities, and 2) the differentiation was mainly due to the separation of the samples from Louisiana with respect to the European set. A hypothesis emerged in which subsequent introductions of crayfish from different sources were performed. This hypothesis might explain the high genetic diversity observed within each population and the genetic differentiation among populations, as the result, respectively, of the introduction of different sets of crayfish and the casual bias of introductions. Although preliminary, our results suggest that RAPDs could be helpful in providing information about human-mediated introduced populations.     

The Tumor Suppressor Functions of p27kip1 Include Control of the Mesenchymal/Amoeboid Transition

In many human cancers, p27 downregulation correlates with a worse prognosis, suggesting that p27 levels could represent an important determinant in cell transformation and cancer development. Using a mouse model system based on v-src-induced transformation, we show here that p27 absence is always linked to a more aggressive phenotype. When cultured in three-dimensional contexts, v-src-transformed p27-null fibroblasts undergo a morphological switch from an elongated to a rounded cell shape, accompanied by amoeboid-like morphology and motility. Importantly, the acquisition of the amoeboid motility is associated with a greater ability to move and colonize distant sites in vivo. The reintroduction of different p27 mutants in v-src-transformed p27-null cells demonstrates that the control of cell proliferation and motility represents two distinct functions of p27, both necessary for it to fully act as a tumor suppressor. Thus, we highlight here a new p27 function in driving cell plasticity that is associated with its C-terminal portion and does not depend on the control of cyclin-dependent kinase activity.Dissemination of tumor cells is strictly linked to their ability to attach to and move within the extracellular matrix (ECM) in a three-dimensional (3D) environment. The use of 3D experimental model systems revealed that a higher complexity in cell migration and adaptation responses exists in the 3D model than in the classical 2D model (10, 16, 41, 49). A striking example is given by the fact that only in 3D could individually migrating cells use different mechanisms such as mesenchymal and amoeboid motility (16, 17). The relative slow mesenchymal migration is characterized by a fibroblast-like spindle shape and is dependent on integrin-mediated adhesion and on protease function (16). The amoeboid motility can in some cases represent a less adhesive, integrin-independent type of movement. Cells use a propulsive mechanism and are highly deformable, and rather than degrade the matrix, they are able to squeeze through it (16). As a result, the cells that use the amoeboid motility can potentially move faster than cells that use a mesenchymal strategy. Mesenchymal and amoeboid movements are also characterized by a different involvement of small GTPases of the Rho family. A high RhoA activity is associated mainly with the amoeboid motility, while the mesenchymal migration needs a high Rac activity at the leading edge to promote the extension of cellular protrusions (41, 48). Under certain circumstances, cancer cells can undergo conversion from a mesenchymal toward an amoeboid motility, an event referred as mesenchymal-amoeboid transition (MAT) (50). MAT represents a putative escape mechanism in tumor cell dissemination that could be induced by inhibition of pericellular proteolysis (50) or by increased membrane-associated RhoA activity (18, 40).Key mediators of cell motility through ECM substrates are the members of the Src family kinases. The prototype of Src family kinases, c-Src (14), is activated following cell-ECM adhesion and contributes to regulate the focal adhesion turnover and the cytoskeletal modifications necessary for normal cell adhesion and motility (52). The c-Src gene is the proto-oncogene of the transforming gene v-src of Rous sarcoma virus, and its elevated protein level and activity have been found in many human tumors (20, 28, 27, 34). Despite the accumulation of information and new molecular understanding of how Src is controlled, there is still an incomplete picture about its role in the generation of the malignant phenotype. v-Src shows higher levels of the kinase activity and transforming ability than c-Src (14, 15, 52). It induces normal cells to acquire a variety of transformed features, including alteration of morphology and increase of invasion ability due to its role in focal adhesion remodeling (7, 9, 13).Many data suggest that there is a close relationship between cell-ECM interaction and the proliferation and movements in both normal and tumor cells (5, 38, 43). Accordingly, Src activation may influence not only cell motility but also cell cycle progression by targeting the cell cycle inhibitor p27^kip1 to proteasomal degradation (22, 39). Recent evidences indicated that p27^kip1 (hereafter called p27) can also regulate cell migration, even though its role still remains controversial since it has been reported to either block or stimulate cell movements (1, 4, 11, 19, 21, 23, 29, 45).Based on these notions, we tested the possible contribution of p27 to the growth and motility phenotypes induced by v-src transformation, with special regard to those cellular invasive features that can be observed in 3D environments. By studying in vitro and in vivo the behavior of wild-type (WT) and p27-null fibroblasts transformed with v-src, we highlight a new role for p27 in the regulation of cellular plasticity that can ultimately drive tumor cell shape, motility, and invasion.     

Phloem sap of tomato plants contains a DIR1 putative ortholog

Arabidopsis thaliana defective in induced resistance 1 (At-DIR1) has been characterized as a protein responsible for the generation or transmission of the still unknown signal involved in systemic acquired resistance. This acidic apoplastic protein is a member of the family of lipid transfer proteins and was detected in vascular fluids. To our knowledge, no DIR1-like protein has been described in other plant species. Hence, we have performed data mining to identify a putative ortholog of DIR1 in tomato. This strategy allowed the detection of a few gene products displaying sequence similarity to At-DIR1 whose structural features were further analysed in silico. The best match (unigene SGN-327306) encoded a protein with an acidic pI, a peculiar characteristic of DIR1 among lipid transfer proteins, and was hence selected as a putative tomato ortholog of At-DIR1. This sequence, named Le-DIR1, served for the design of a specific antigenic peptide and the generation of polyclonal antibodies. The antiserum anti-Le-DIR1 recognized a peptide of the expected size (7kDa) in phloem sap of tomato plants, hence confirming the existence of the predicted protein in vascular fluids. This result supports the notion of the existence of common systemic acquired resistance (SAR) signaling molecules in different species.     

Primate cathelicidin orthologues display different structures and membrane interactions

The human cathelicidin LL-37 displays both direct antibacterial activities and the capacity to modulate host-cell activities. These depend on structural characteristics that are subject to positive selection for variation, as observed in a previous analysis of the CAMP gene (encoding LL-37) in primates. The altered balance between cationic and anionic residues in different primate orthologues affects intramolecular salt-bridging and influences the stability of the helical conformation and tendency to aggregate in solution of the peptide. In the present study, we have analysed the effects of these structural variations on membrane interactions for human LL-37, rhesus RL-37 and orang-utan LL-37, using several complementary biophysical and biochemical methods. CD and ATR (attenuated total reflection)-FTIR (Fourier-transform IR) spectroscopy on model membranes indicate that RL-37, which is monomeric and unstructured in bulk solution [F-form (free form)], and human LL-37, which is partly structured and probably aggregated [A-form (aggregated form)], bind biological membranes in different manners. RL-37 may insert more deeply into the lipid bilayer than LL-37, which remains aggregated. AFM (atomic force microscopy) performed on the same supported bilayer as used for ATR-FTIR measurements suggests a carpet-like mode of permeabilization for RL37 and formation of more defined worm-holes for LL-37. Comparison of data from the biological activity on bacterial cells with permeabilization of model membranes indicates that the structure/aggregation state also affects the trajectory of the peptides from bulk solution through the outer cell-wall layers to the membrane. The results of the present study suggest that F-form cathelicidin orthologues may have evolved to have primarily a direct antimicrobial defensive capacity, whereas the A-forms have somewhat sacrificed this to gain host-cell modulating functions.     

<Emphasis Type="Italic">N</Emphasis>-vanillylnonanamide tested as a non-toxic antifoulant,applied to surfaces in a polyurethane coating

The potential on N-vanillylnonanamide (NVN) in preventing the attachment of Pseudomonas stutzeri and a Bacillus cereus-group strain was investigated. NVN up to 852 μM was not toxic, nor was it an energy source for either organism. Microbial attachment assays were carried out on glass and polylysine slides. with NVN being dispersed in or applied to the surfaces using a polyurethane coating. NVN at 205 μM inhibited Bacillus adhesion on glass slides by 48% and the percentage did not significantly increase at 852 μM. NVN blended into or sprayed onto the coating at 205 μmol/kg did not prevent adhesion. The compound is therefore not useful as an antifouling product under the tested coating conditions.     

Species identity, diversity and microbial carbon flow in reassembling macrobenthic communities

Intense disturbance may locally destroy patches of habitat and shape the landscape into a mosaic of reassembling communities. The development of ecosystem properties during such community reassembly is poorly understood. In intertidal bare sediments, trophic relations between microphytobenthos or heterotrophic bacteria and macrofauna invertebrates may guarantee fundamental ecosystem properties such as carbon flow through the food web. We studied the dynamic relation between reassembling macrofauna communities and such microbial carbon flow during recovery after severe disturbance. We deliberately induced prolonged hypoxia in winter and early summer and allowed recolonisation for periods of two and five months. Carbon flow was quantified from basal resources (microphytobenthos and bacteria) to intermediate consumers using ¹³C as a tracer. Within the period of study (5 months), microbial carbon flow fully recovered, although macrofauna diversity was still very low compared to the natural communities (ranging from 6 to 17 species). More than 90% of microbial carbon flow to macrofauna was due to the consumers that recolonised within two months. Two of these species were dominant contributors to microphytobenthos carbon transfer to fauna. Furthermore, at an early stage of reassembly, this ecosystem property was remarkably similar when disturbance took place at different times of the year (winter or early summer), although there were differences in assemblage composition and functional diversity. We conclude that species assemblages and ecosystem function developed relatively independently in this benthic system. We discuss which ecological factors may have caused such non-parallel development of macrofaunal communities and carbon flow.