A general approach of using hair-tubes to monitor the European red squirrel: A method applicable at regional and national scales

Monitoring constitutes a key element in the management and conservation of many mammal species. We describe a technique to obtain population indices for red squirrels (Sciurus vulgaris) using hair-tubes and compare these indices to population estimates obtained by live trapping. Data were collected in seven study areas in the Western and Central Alps in Italy and compared with data previously collected in 11 sites in northern England. The aim was to test if hair-tube census could be used to derive a general predictive model allowing accurate predictions of squirrel numbers in different years, habitats and geographic regions. We used model equations developed from the proportion of hair-tubes visited to predict densities obtained from live-trapping. Hair-tube data gathered in the Central Alps correctly predicted squirrel densities in the Western Alps. A combined data set pooling the sites of these two regions based on the first three years successfully predicted the two successive years. In addition, a combined model derived from areas monitored for five years had a high predictive value locally (89%) and internationally (73%) when applied to the English data set. We therefore believe that the predictive model developed in this study could be of general value and be used to monitor squirrel populations in European low density conifer habitats (0.1–0.5 squirrels/ha). The approach may also be suitable for many tree squirrel populations in North America and other arboreal rodents that occur at similar densities.     

Agroecology: the key role of arbuscular mycorrhizas in ecosystem services

The beneficial effects of arbuscular mycorrhizal (AM) fungi on plant performance and soil health are essential for the sustainable management of agricultural ecosystems. Nevertheless, since the ‘first green revolution’, less attention has been given to beneficial soil microorganisms in general and to AM fungi in particular. Human society benefits from a multitude of resources and processes from natural and managed ecosystems, to which AM make a crucial contribution. These resources and processes, which are called ecosystem services, include products like food and processes like nutrient transfer. Many people have been under the illusion that these ecosystem services are free, invulnerable and infinitely available; taken for granted as public benefits, they lack a formal market and are traditionally absent from society’s balance sheet. In 1997, a team of researchers from the USA, Argentina and the Netherlands put an average price tag of US $33 trillion a year on these fundamental ecosystem services. The present review highlights the key role that the AM symbiosis can play as an ecosystem service provider to guarantee plant productivity and quality in emerging systems of sustainable agriculture. The appropriate management of ecosystem services rendered by AM will impact on natural resource conservation and utilisation with an obvious net gain for human society.     

Dispersal and habitat cuing of Eurasian red squirrels in fragmented habitats

Animal dispersal and subsequent settlement is a key process in the life history of many organisms, when individuals use demographic and environmental cues to target post-dispersal habitats where fitness will be highest. To investigate the hypothesis that environmental disturbance (habitat fragmentation) may alter these cues, we compared dispersal patterns of 60 red squirrels (Sciurus vulgaris) in three study sites that differ in habitat composition and fragmentation. We determined dispersal distances, pre- and post-dispersal habitat types and survival using a combination of capture–mark–recapture, radio-tracking and genetic parentage assignment. Most (75%) squirrels emigrated from the natal home range with mean dispersal distance of 1,014 ± 925 m (range 51–4,118 m). There were no sex-related differences in dispersal patterns and no differences in average dispersal distance, and the proportion of dispersers did not differ between sites. In one of the sites, dispersers settled in patches where density was lower than in the natal patch. In the least fragmented site, 90% of animals settled in the natal habitat type (habitat cuing) against 44–54% in the more strongly fragmented sites. Overall, more squirrels settled in the natal habitat type than expected based on habitat availability, but this was mainly due to individuals remaining within the natal wood. In the highly fragmented landscape, habitat cuing among emigrants did not occur more frequently than expected. We concluded that increased habitat fragmentation seemed to reduce reliable cues for habitat choice, but that dispersing squirrels settled in patches with lower densities of same-sex animals than at the natal home range or patch, independent of degree of fragmentation.     

Evolutionary lability of a complex life cycle in the aphid genus <Emphasis Type="Italic">Brachycaudus</Emphasis>

Background  

Most aphid species complete their life cycle on the same set of host-plant species, but some (heteroecious species) alternate between different hosts, migrating from primary (woody) to secondary (herbaceous) host plants. The evolutionary processes behind the evolution of this complex life cycle have often been debated. One widely accepted scenario is that heteroecy evolved from monoecy on woody host plants. Several shifts towards monoecy on herbaceous plants have subsequently occurred and resulted in the radiation of aphids. Host alternation would have persisted in some cases due to developmental constraints preventing aphids from shifting their entire life cycle to herbaceous hosts (which are thought to be more favourable). According to this scenario, if aphids lose their primary host during evolution they should not regain it. The genus Brachycaudus includes species with all the types of life cycle (monoecy on woody plants, heteroecy, monoecy on herbs). We used this genus to test hypotheses concerning the evolution of life cycles in aphids.     

T cell polarity at the immunological synapse is required for CD154-dependent IL-12 secretion by dendritic cells

Ag-specific interaction between T lymphocytes and dendritic cells (DCs) leads to both T cell and DC activation. CD154 (CD40 ligand)/CD40 interactions have been shown to play a major, although not exclusive, role in this functional cross-talk. Interactions between T cells and DCs are structured by an immunological synapse (IS), characterized by polarization of the T cell microtubule cytoskeleton toward the interacting DCs. Yet the role T cell polarization may play in T cell-induced DC activation is mostly unknown. In this study, we address the role of T cell polarity in CD154-dependent activation of DCs in a human model, using two different tools to block T cell polarity (i.e., a microtubule depolymerizing drug and an inhibitor of atypical protein kinase C). We show that CD154 is recruited and concentrated at the IS formed between human primary T cells and autologous DCs and that this recruitment requires T cell polarity at the IS. Moreover, we show that T cell polarization at the IS controls T cell-dependent CD154-CD40 signaling in DCs as well as CD154-dependent IL-12 secretion by DCs. This study shows that T cell polarity at the IS plays a key role in CD154/CD40-dependent cross-talk between CD4(+) T cells and DCs.     

Screening for subtelomeric rearrangements using genetic markers in 70 patients with unexplained mental retardation

Cryptic unbalanced rearrangements involving chromosome ends are a significant cause of idiopathic mental retardation. The most frequently used technique to screen for these subtle rearrangements is Multiprobe fluorescence in situ hybridization (FISH). As this is a labor-intensive technique, we used microsatellite genotyping to detect possible subtelomeric rearrangements in a study population. Out of the 70 patients we screened, three chromosomal rearrangements were detected: a deletion of marker D2S2986, a deletion of marker D7S594 and a deletion of marker D19S424. However, none of these aberrations appeared to be disease causing.     

Roles of stem cell factor/c-Kit and effects of Glivec/STI571 in human uveal melanoma cell tumorigenesis

The B-Raf(V599E)-mediated constitutive activation of ERK1/2 is involved in establishing the transformed phenotype of some uveal melanoma cells (Calipel, A., Lefevre, G., Pouponnot, C., Mouriaux, F., Eychene, A., and Mascarelli, F. (2003) J. Biol. Chem. 278, 42409-42418). We have shown that stem cell factor (SCF) is involved in the proliferation of normal uveal melanocytes and that c-Kit is expressed in 75% of primary uveal melanomas. This suggests that the acquisition of autonomous growth during melanoma progression may involve the SCF/c-Kit axis. We used six human uveal melanoma tumor-derived cell lines and normal uveal melanocytes to characterize the SCF/c-Kit system and to assess its specific role in transformation. We investigated the possible roles of activating mutations in c-KIT, the overexpression of this gene, and ligand-dependent c-Kit overactivation in uveal melanoma cell tumorigenesis. Four cell lines (92.1, SP6.5, Mel270, and TP31) expressed both SCF and c-Kit, and none harbored the c-KIT mutations in exons 9, 11, 13, and 17 that have been shown to induce SCF-independent c-Kit activation. Melanoma cell proliferation was strongly inhibited by small interfering RNA-mediated depletion of c-Kit in these cells, despite the presence of (V599E)B-Raf in SP6.5 and TP31 cells. We characterized the signaling pathways involved in SCF/c-Kit-mediated cell growth and survival in normal and tumoral melanocytes and found that constitutive ERK1/2 activation played a key role in both the SCF/c-Kit autocrine loop and the gain of function of (V599E)B-Raf for melanoma cell proliferation and transformation. We also provide the first evidence that Glivec/STI571, a c-Kit tyrosine kinase inhibitor, could be used to treat uveal melanomas.     

Analysis of human cytochrome P450 2C8 substrate specificity using a substrate pharmacophore and site-directed mutants

The structural determinants of substrate specificity of human liver cytochrome P450 2C8 (CYP2C8) were investigated using site-directed mutants chosen on the basis of a preliminary substrate pharmacophore and a three-dimensional (3D) model. Analysis of the structural features common to CYP2C8 substrates exhibiting a micromolar K(m) led to a substrate pharmacophore in which the site of oxidation by CYP2C8 is 12.9, 8.6, 4.4, and 3.9 A from features that could establish ionic or hydrogen bonds, and hydrophobic interactions with protein amino acid residues. Comparison of this pharmacophore with a 3D model of CYP2C8 constructed using the X-ray structure of CYP2C5 suggested potential CYP2C8 amino acid residues that could be involved in substrate recognition. Twenty CYP2C8 site-directed mutants were constructed and expressed in yeast to compare their catalytic activities using five CYP2C8 substrates that exhibit different structures and sizes [paclitaxel, fluvastatin, retinoic acid, a sulfaphenazole derivative (DMZ), and diclofenac]. Mutation of arginine 241 had marked effects on the hydroxylation of anionic substrates of CYP2C8 such as retinoic acid and fluvastatin. Serine 100 appears to be involved in hydrogen bonding interactions with a polar site of the CYP2C8 substrate pharmacophore, as shown by the 3-4-fold increase in the K(m) of paclitaxel and DMZ hydroxylation after the S100A mutation. Residues 114, 201, and 205 are predicted to be in close contact with substrates, and their mutations lead either to favorable hydrophobic interactions or to steric clashes with substrates. For instance, the S114F mutant was unable to catalyze the 6alpha-hydroxylation of paclitaxel. The S114F and F205A mutants were the best catalysts for retinoic acid and paclitaxel (or fluvastatin) hydroxylation, respectively, with k(cat)/K(m) values 5 and 2.1 (or 2.4) times higher, respectively, than those found for CYP2C8. Preliminary experiments of docking of the substrate into the experimentally determined X-ray structure of substrate-free CYP2C8, which became available quite recently [Schoch, G. A., et al. (2004) J. Biol. Chem. 279, 9497], were consistent with key roles for S100, S114, and F205 residues in substrate binding. The results suggest that the effects of mutation of arginine 241 on anionic substrate hydroxylation could be indirect and result from alterations of the packing of helix G with helix B'.     

Inactivation and intracellular retention of the human I183N mutated melanocortin 3 receptor associated with obesity

Melanocortins are known to be involved in the regulation of feeding behavior. These hormones mediate their effects through G protein-coupled receptors (GPCRs) by stimulating adenylate cyclase. The melanocortin 3 receptor (MC3R) in the melanocortin receptor (MCR) family has been identified as a neural receptor subtype mainly expressed in the brain in mammals. Until now, only one heterozygous mutation (I183N) has been identified in the coding region of this receptor in two obese patients of the same family. In this study, we reported the functional characterization of the I183N mutated MC3R compared with that of the wild-type MC3R after transfection in HEK293 cells. Our results showed that the I183N mutation totally abolished the activity of the mutated receptor to generate intracellular cAMP. Furthermore, confocal microscopy observation revealed that the mutation induced an intracellular retention of the mutated receptor. Moreover, we demonstrated for the first time by co-transfection studies that the mutated receptor could reduce the wild-type receptor activity through a dominant negative effect.