New insights into the past and recent evolutionary history of the Corsican mouflon (Ovis gmelini musimon) to inform its conservation

Human-mediated species dispersal across the Mediterranean stretches back at least 10,000 years and has left an indelible stamp on present-day biodiversity. Believed to be a descendant of the Asiatic mouflon (Ovis gmelini gmelinii), the Corsican mouflon (O. g. musimon) was translocated during the Neolithic as ancestral livestock by humans migrating from the Fertile Crescent to the Western Mediterranean. Today, two geographically limited and disconnected populations can be found in Corsica. Whether they originated from distinct founders or one ancestral population that later split remains unknown, although such information is pivotal for the species’ management on the island. We genotyped 109 and 176 individuals at the Cytochrome-b gene and 16 loci of the microsatellite DNA, respectively, to gain insights into the natural history of the Corsican mouflon. We found evidence confirming that the Asiatic was the ancestor of the Corsican mouflon, which should thus be unvaryingly referred to as O. g. musimon, i.e. as a subspecies of the Asiatic mouflon. Haplotype divergence dating and the investigation of genetic structure highlighted a strong and ancient genetic differentiation between the two Corsican populations. Approximate Bayesian Computation pointed to the introduction of a single group of founders as the most reliable scenario for the origin of the entire Corsican population. Later, this ancestral stock would have decreased in number, facing genetic bottlenecks and eventually resulting in two divergent demes. Splitting most likely occurred several hundred years ago. Their shared past notwithstanding, we discuss whether the two relic Corsican mouflon populations should be now considered as distinct management units.

     

Transient Neuronal Populations Are Required to Guide Callosal Axons: A Role for Semaphorin 3C

The corpus callosum (CC) is the main pathway responsible for interhemispheric communication. CC agenesis is associated with numerous human pathologies, suggesting that a range of developmental defects can result in abnormalities in this structure. Midline glial cells are known to play a role in CC development, but we here show that two transient populations of midline neurons also make major contributions to the formation of this commissure. We report that these two neuronal populations enter the CC midline prior to the arrival of callosal pioneer axons. Using a combination of mutant analysis and in vitro assays, we demonstrate that CC neurons are necessary for normal callosal axon navigation. They exert an attractive influence on callosal axons, in part via Semaphorin 3C and its receptor Neuropilin-1. By revealing a novel and essential role for these neuronal populations in the pathfinding of a major cerebral commissure, our study brings new perspectives to pathophysiological mechanisms altering CC formation.     

The NADH oxidase of Streptococcus pneumoniae: its involvement in competence and virulence

A soluble flavoprotein that reoxidizes NADH and reduces molecular oxygen to water was purified from the facultative anaerobic human pathogen Streptococcus pneumoniae. The nucleotide sequence of nox, the gene which encodes it, has been determined and was characterized at the functional and physiological level. Several nox mutants were obtained by insertion, nonsense or missense mutation. In extracts from these strains, no NADH oxidase activity could be measured, suggesting that a single enzyme encoded by nox, having a C44 in its active site, was utilizing O2 to oxidize NADH in S. pneumoniae. The growth rate and yield of the NADH oxidase-deficient strains were not changed under aerobic or anaerobic conditions, but the efficiency of development of competence for genetic transformation during growth was markedly altered. Conditions that triggered competence induction did not affect the amount of Nox, as measured using Western blotting, indicating that nox does not belong to the competence-regulated genetic network. The decrease in competence efficiency due to the nox mutations was similar to that due to the absence of oxygen in the nox+ strain, suggesting that input of oxygen into the metabolism via NADH oxidase was important for controlling competence development throughout growth. This was not related to regulation of nox expression by O2. Interestingly, the virulence and persistence in mice of a blood isolate was attenuated by a nox insertion mutation. Global cellular responses of S. pneumoniae, such as competence for genetic exchange or virulence in a mammalian host, could thus be modulated by oxygen via the NADH oxidase activity of the bacteria, although the bacterial energetic metabolism is essentially anaerobic. The enzymatic activity of the NADH oxidase coded by nox was probably involved in transducing the external signal, corresponding to O2 availability, to the cell metabolism and physiology; thus, this enzyme may function as an oxygen sensor. This work establishes, for the first time, the role of O2 in the regulation of pneumococcal transformability and virulence.     

Biasing a Monte Carlo chain growth method with Ramachandran's plot: Application to twenty-L-alanine

In this paper, we explore the possibility of using experimental observations in the Monte Carlo chain growth method that we have previously developed. In this method, the macromolecule (peptide, protein, nucleic acid, etc.) is grown atom-by-atom (or residue-by-residue, etc.) and partial chains are replicated according to their Boltzmann weights. Once the molecule completed, we are left with a Boltzmann-distributed ensemble of configurations. For long molecules, an efficient sampling of the (extremely large) phase space is difficult for obvious reasons (existence of many local minima, limited computer memory, etc.). In the case in which one is mainly interested in the low energy conformations, we have incorporated in the growth scheme experimental observations taken from the Protein Data Banks. More precisely, we have considered the case of twenty-L -alanine and we have used the (experimental) Ramachandran's plot for this residue. The biased growth procedure goes as follows: (a) each time one adds along the main backbone chain, either a carbon atom belonging to a carbonyl group, or a nitrogen atom, its dihedral angle (?) or (ψ) is drawn with a probability law that reflects the experimental Ramachandran (?,ψ) plot; (b) the bias introduced in this way is canceled through an extra term in the energy (replication energy = true energy + bias energy); (c) the configurations, generated at T = 1000 K, are then energy minimized. We have worked with an all-atom CHARMM force field, and Ramachandran's plot for the alanine was modeled through three angular zones (α-helix, β-sheet, coil). In our calculations, the probabilities of the α (p_α) and β (p_β) regions have been varied in large proportions (p_α between 0.64 and 0.19, the “experimental” value being 0.59). The results, based on 35 “unbiased” and 25 “biased” (or “guided”) distinct minimized configurations clearly demonstrate the efficiency of the method. The low energy configurations, for all tested values of p_α, have a total (or almost total) a helix content. The unbiased configurations have much higher energies (in general, even higher than the left-handed helix). Note that the method is not “α helix in-α helix out,” since working at T = 300 K with the experimental (p_α = 0.59) value yields configurations partially frozen in a C alanine dipeptide type of local minima. © 1993 John Wiley & Sons, Inc.     

Test of the extended two-state model for the kinetic intermediates observed in the folding transition of ribonuclease A

A test has been made of the proposal that: (a) the extended two-state model describes the kinetic intermediates seen in the folding transition of RNAase A, i.e. that the only species present in folding experiments are the native protein and multiple forms of the completely unfolded protein; and (b) that the interconversion between the two known unfolded forms of RNAase A (the U₁

U₂ reaction) is described solely by the cis-trans isomerization of the proline residues. The test is to measure the rate of the U₁

U₂ reaction in a wide range of refolding conditions and to compare these data with the kinetic properties of proline isomerization.The main results are as follows. (1) The activation enthalpy of the U₁

U₂ reaction in refolding conditions (pH 6, 20 ° to 40 °C) is less than 5 kcal/mol. This is much too small to be explained as proline isomerization. (2) Both the rate and the activation enthalpy change sharply at guanidine hydrochloride concentrations below 2 m. There appear to be two pathways for the U₁

U₂ reaction in refolding conditions, and the slower pathway is favored by adding guanidine hydrochloride. (3) The rate and activation enthalpy for proline isomerization in l-alanyl-l-proline are unaffected by 2 m-guanidine hydrochloride.The results show that the proline isomerization hypothesis and the extended two-state model cannot both be correct for RNAase A. They suggest that partial folding occurs rapidly in refolding conditions and that the extended two-state model is invalid. They leave open the question of whether or not proline isomerization is the rate-limiting step in the U₁

U₂ reaction.Another possible source of slow configurational reactions in the unfolded state is mentioned. The three major, overlapping, disulfide-bonded loops of RNAase A can exist in two isomeric configurations. Interconversion of these isomers requires pulling one loop, or one end of the polypeptide chain, through a second loop and this is likely to be a slow process.In some conditions, heat-unfolded but not guanidine-unfolded RNAase A shows a second slow-refolding process. It may result from aggregates of the heatunfolded protein which are formed and broken up slowly. Conditions are given for eliminating this reaction.     

Vitamin D-dependent calcium-binding protein. Changes during gestation, prenatal and postnatal development in rats

During the perinatal period, calcium metabolism is stressed. As intestinal Ca-binding protein is considered as a molecular expression of the hormonal effect of 1,25-dihydroxycholecalciferol (1,25(OH)2D3), Ca-binding protin measurements may document the vitamin D roles during this period. We describe the variations of Ca-binding protein concentrations in the rat during the last 5 days of gestation, in the maternal duodenum, placentas, fetal membranes and fetal intestines. We also report intestinal Ca-binding protein changes from birth until weaning. The evolution of the maternal intestinal Ca-binding protein, which increases on day 19.5 of gestation, is consistent with that of calcium intestinal absorption and may be explained by increased 1,25(OH)2D3 production. Placental Ca-binding protein rises from day 17.5 until the end of gestation, and may be related to the profile of calcium transfer from mother to fetuses. It is noteworthy that the placental Ca-binding protein is predominantly found in the fetal part of the organ where materno-fetal exchanges occur. The yolk sac synthesizes substantial amounts of Ca-binding protein. In the fetal membranes, Ca-binding protein plateaus from day 17.5 until day 20.5 and decreases on day 21.5. The Ca-binding protein presence in the fetal placenta and in the yolk sac may suggest that these tissues are also targets for vitamin D. In the fetus the intestinal Ca-binding protein s is detected as early as day 17.5 of gestation and increases markedly during the last day of gestation. From birth and during the first 3 weeks of postnatal life, the intestinal Ca-binding protein concentration does not change. It undergoes a sharp rise just at the time of weaning. We have also shown that the specific distribution of Ca-binding protein along the intestine is acquired during intrauterine life and does not change with sucking or weaning. The two main changes of intestinal Ca-binding protein, observed just before birth and at weaning, may reflect the intestinal maturation and/or variations in vitamin D metabolism.