The Caenorhabditis elegans homologue of the proto-oncogene ect-2 positively regulates RAS signalling during vulval development

Guanine nucleotide exchange factors (GEFs) regulate the activity of small GTP-binding proteins in a variety of biological processes. We have identified a gain-of-function mutation in the Caenorhabditis elegans GEF ect-2, the homologue of the mammalian ect2 proto-oncogene that has an essential role during cytokinesis. Here, we report that, in addition to its known function during mitosis, ECT-2 promotes the specification of the primary vulval cell fate by activating RAS/mitogen-activated protein kinase (MAPK) signalling before the end of the S-phase. Epistasis analysis indicates that ECT-2 crosstalks to the canonical RAS/MAPK cascade upstream of the RAS GEF SOS-1 by means of a RHO-1 signalling pathway. Our results raise the possibility that the transforming activity of the mammalian ect-2 oncogene could be due to hyperactivation of the RAS/MAPK pathway.     

Inverse electrostatic effect: electrostatic repulsion in the unfolded state stabilizes a leucine zipper

The pH-dependent stability of a protein is strongly affected by electrostatic interactions between ionizable residues in the folded as well as unfolded state. Here we characterize the individual contributions of charged Glu and His residues to stability and determine the NMR structure of the designed, heterodimeric leucine zipper AB consisting of an acidic A chain and a basic B chain. Thermodynamic parameters are compared with those of the homologous leucine zipper AB(SS) in which the A and B chains are disulfide-linked. NMR structures of AB based on (1)H NMR data collected at 600 MHz converge, and formation of the same six interchain salt bridges found previously in disulfide-linked AB(SS) [Marti, D. N., and Bosshard, H. R. (2003) J. Mol. Biol. 330, 621-637] is indicated. While the structures of AB and AB(SS) are very similar, their pH-dependent relative stabilities are strikingly different. The stability of AB peaks at pH approximately 4.5 and is higher at pH 8 than at pH 2. In contrast, AB(SS) is most stable at acidic pH where no interhelical salt bridges are formed. The different energetic contributions of charged Glu and His residues to stability of the two coiled coil structures were evaluated from pK(a) shifts induced by folding. The six charged Glu residues involved in salt bridges stabilize leucine zipper AB by 4.5 kJ/mol yet destabilize disulfide-linked AB(SS) by -1.1 kJ/mol. Two non-ion-paired Glu charges destabilize AB by only -1.8 kJ/mol but AB(SS) by -5.6 kJ/mol. The higher relative stability of AB at neutral pH is not caused by more favorable electrostatic interactions in the folded leucine zipper. It is due mainly to unfavorable electrostatic interactions in the unfolded A and B chains and may therefore be called an inverse electrostatic effect. This study illustrates the importance of residual interactions in the unfolded state and how the energetics of the unfolded state affect the stability of the folded protein.     

Dioxygen affinity in heme proteins investigated by computer simulation

We present an investigation of the molecular basis of the modulation of oxygen affinity in heme proteins using computer simulation. QM-MM calculations are applied to explore distal and proximal effects on O(2) binding to the heme, while classical molecular dynamics simulations are employed to investigate ligand migration across the polypeptide to the active site. Trends in binding energies and in the kinetic constants are illustrated through a number of selected examples highlighting the virtues and the limitations of the applied methodologies. These examples cover a wide range of O(2)-affinities, and include: the truncated-N and truncated-O hemoglobins from Mycobacterium tuberculosis, the mammalian muscular O(2) storage protein: myoglobin, the hemoglobin from the parasitic nematode Ascaris lumbricoides, the oxygen transporter in the root of leguminous plants: leghemoglobin, the Cerebratulus lacteus nerve tissue hemoglobin, and the Alcaligenes xyloxidans cytochrome c'.     

Genetic diversity among horse populations with a special focus on the Franches-Montagnes breed

Genetic characterization helps to assure breed integrity and to assign individuals to defined populations. The objective of this study was to characterize genetic diversity in six horse breeds and to analyse the population structure of the Franches-Montagnes breed, especially with regard to the degree of introgression with Warmblood. A total of 402 alleles from 50 microsatellite loci were used. The average number of alleles per locus was significantly lower in Thoroughbreds and Arabians. Average heterozygosities between breeds ranged from 0.61 to 0.72. The overall average of the coefficient of gene differentiation because of breed differences was 0.100, with a range of 0.036-0.263. No significant correlation was found between this parameter and the number of alleles per locus. An increase in the number of homozygous loci with increasing inbreeding could not be shown for the Franches-Montagnes horses. The proportion of shared alleles, combined with the neighbour-joining method, defined clusters for Icelandic Horse, Comtois, Arabians and Franches-Montagnes. A more disparate clustering could be seen for European Warmbloods and Thoroughbreds, presumably from frequent grading-up of Warmbloods with Thoroughbreds. Grading-up effects were also observed when Bayesian and Monte Carlo resampling approaches were used for individual assignment to a given population. Individual breed assignments to defined reference populations will be very difficult when introgression has occurred. The Bayesian approach within the Franches-Montagnes breed differentiated individuals with varied proportions of Warmblood.     

Marine reserves demonstrate trophic interactions across habitats

Several infaunal bivalve taxa show patterns of decreased biomass in areas with higher densities of adjacent reef-associated predators (the snapper, Pagrus auratus and rock lobster, Jasus edwardsii). A caging experiment was used to test the hypothesis that patterns observed were caused by predation, using plots seeded with a known initial density of the bivalve Dosinia subrosea to estimate survivorship. The caging experiment was replicated at several sites inside and outside two highly protected marine reserves: predators are significantly more abundant inside these reserves. Survivorship in fully caged, partially caged and open plots were then compared at sites having either low (non reserve) or high (reserve) predator density. The highest rates of survivorship of the bivalve were found in caged plots inside reserves and in all treatments outside reserves. However, inside reserves, open and partially caged treatments exhibited low survivorship. It was possible to specifically attribute much of this mortality to predation by large rock lobsters, due to distinctive marks on the valves of dead D. subrosea. This suggests that predation by large rock lobster could indeed account for the distributional patterns previously documented for certain bivalve populations. Our results illustrate that protection afforded by marine reserves is necessary to investigate how depletion through fishing pressure can change the role of upper-level predators and trophic processes between habitats. Electronic Supplementary Material Supplementary material is available for this article at An erratum to this article can be found at     

Differential Effects of Lovastatin on Cisplatin Responses in Normal Human Mesothelial Cells versus Cancer Cells: Implication for Therapy

The cancer killing efficacy of standard chemotherapeutic agents such as cisplatin (CDDP) is limited by their side effects to normal tissues. Therefore, research efforts optimizing the safety and efficacy of those agents are clinically relevant. We did screen for agents that specifically protect normal human mesothelial cells against CDDP without reducing the cancer cell killing efficacy. Lovastatin was identified from the screen. Lovastatin at a pharmacologically relevant concentration strongly arrested the proliferation of normal cells, whereas cancer cells were less affected. CDDP-induced DNA damage response was not activated and normal cells showed enhanced tolerance to CDDP when normal cells were treated with the combination of CDDP and lovastatin. We demonstrate that interfering with protein geranylgeranylation is involved in the lovastatin-mediated CDDP protective effect in normal cells. In contrast to normal cells, in cancer cells lovastatin did not change the CDDP-induced response, and cancer cells were not protected by lovastatin. Furthermore, lovastatin at the pharmacological relevant concentration per se induced DNA damage, oxidative stress and autophagy in cancer cells but not in normal mesothelial cells. Therefore, our data suggest that lovastatin has a potential to improve the therapeutic index of cisplatin-based therapy.     

1H, 15N and 13C chemical shift assignments of the BA42 protein of the psychrophilic bacteria Bizionia argentinensis sp. nov

BA42 is a protein belonging to the psychrophilic bacteria Bizionia argentinensis sp. nov. Bioinformatics analysis showed that it presents significant sequence identity with a Pfam A family, DUF 477, found both in eukarya and eubacteria but of unknown function in all these organisms. Here, we report the NMR spectra assignment of this 145 amino acid protein. These data will allow performing NMR structural studies with the aim of using the three-dimensional structure as relevant information in order to determine the function of this family of proteins.