Identification and function of exchange proteins activated directly by cyclic AMP (Epac) in mammalian spermatozoa

The role of cAMP in spermatic functions was classically thought to be mediated exclusively through the activation of Protein Kinase A (PKA). However, it has recently been shown that cAMP also exerts its effects through a PKA-independent pathway activating a family of proteins known as Epac proteins. Therefore, many of the spermatic functions thought to be regulated by cAMP through the activation of PKA are again under study. We aimed to identify and to investigate the role of Epac proteins in spermatozoa using a specific permeable analog (8-Br-2'-O-Me-cAMP). Also, we aimed to study its relationship with E-cadherin, an adhesion protein involved in fertility. Our results demonstrate the presence and sub-cellular distribution of Epac 1 and Epac 2 in mammalian spermatozoa. Capacitation and the acrosome reaction induced a change in the localization of Epac proteins in sperm. Moreover, incubation with 8-Br-2'-O-Me-cAMP prompted an increase in Rap1 activation, in the scrambling of plasma membrane phospholipids (necessary for the capacitation process), the acrosome reaction, motility, and calcium mobilization, when spermatozoa were incubated in acrosome reaction conditions. Finally, the activation of Epac proteins induced a change in the distribution of E-cadherin. Therefore, the increase in the acrosome reaction, together with the increase in calcium (which is known to be essential for fertilization) and the Epac nteraction with E-cadherin, might indicate that Epac proteins have an important role in gamete recognition and fertilization.     

Epididymis response partly compensates for spermatozoa oxidative defects in snGPx4 and GPx5 double mutant mice

We report here that spermatozoa of mice lacking both the sperm nucleus glutathione peroxidase 4 (snGPx4) and the epididymal glutathione peroxidase 5 (GPx5) activities display sperm nucleus structural abnormalities including delayed and defective nuclear compaction, nuclear instability and DNA damage. We show that to counteract the GPx activity losses, the epididymis of the double KO animals mounted an antioxydant response resulting in a strong increase in the global H(2)O(2)-scavenger activity especially in the cauda epididymis. Quantitative RT-PCR data show that together with the up-regulation of epididymal scavengers (of the thioredoxin/peroxiredoxin system as well as glutathione-S-transferases) the epididymis of double mutant animals increased the expression of several disulfide isomerases in an attempt to recover normal disulfide-bridging activity. Despite these compensatory mechanisms cauda-stored spermatozoa of double mutant animals show high levels of DNA oxidation, increased fragmentation and greater susceptibility to nuclear decondensation. Nevertheless, the enzymatic epididymal salvage response is sufficient to maintain full fertility of double KO males whatever their age, crossed with young WT female mice.     

Insecticide resistance in the dengue vector Aedes aegypti from Martinique: distribution, mechanisms and relations with environmental factors

Dengue is an important mosquito borne viral disease in Martinique Island (French West Indies). The viruses responsible for dengue are transmitted by Aedes aegypti, an indoor day-biting mosquito. The most effective proven method for disease prevention has been by vector control by various chemical or biological means. Unfortunately insecticide resistance has already been observed on the Island and recently showed to significantly reduce the efficacy of vector control interventions. In this study, we investigated the distribution of resistance and the underlying mechanisms in nine Ae. aegypti populations. Statistical multifactorial approach was used to investigate the correlations between insecticide resistance levels, associated mechanisms and environmental factors characterizing the mosquito populations. Bioassays revealed high levels of resistance to temephos and deltamethrin and susceptibility to Bti in the 9 populations tested. Biochemical assays showed elevated detoxification enzyme activities of monooxygenases, carboxylesterases and glutathione S-tranferases in most of the populations. Molecular screening for common insecticide target-site mutations, revealed the presence of the "knock-down resistance" V1016I Kdr mutation at high frequency (>87%). Real time quantitative RT-PCR showed the potential involvement of several candidate detoxification genes in insecticide resistance. Principal Component Analysis (PCA) performed with variables characterizing Ae. aegypti from Martinique permitted to underline potential links existing between resistance distribution and other variables such as agriculture practices, vector control interventions and urbanization. Insecticide resistance is widespread but not homogeneously distributed across Martinique. The influence of environmental and operational factors on the evolution of the resistance and mechanisms are discussed.     

Down-regulation of GABA(A) receptor via promiscuity with the vasoactive peptide urotensin II receptor. Potential involvement in astrocyte plasticity

GABA(A) receptor (GABA(A)R) expression level is inversely correlated with the proliferation rate of astrocytes after stroke or during malignancy of astrocytoma, leading to the hypothesis that GABA(A)R expression/activation may work as a cell proliferation repressor. A number of vasoactive peptides exhibit the potential to modulate astrocyte proliferation, and the question whether these mechanisms may imply alteration in GABA(A)R-mediated functions and/or plasma membrane densities is open. The peptide urotensin II (UII) activates a G protein-coupled receptor named UT, and mediates potent vasoconstriction or vasodilation in mammalian vasculature. We have previously demonstrated that UII activates a PLC/PIPs/Ca(2+) transduction pathway, via both G(q) and G(i/o) proteins and stimulates astrocyte proliferation in culture. It was also shown that UT/G(q)/IP(3) coupling is regulated by the GABA(A)R in rat cultured astrocytes. Here we report that UT and GABA(A)R are co-expressed in cerebellar glial cells from rat brain slices, in human native astrocytes and in glioma cell line, and that UII inhibited the GABAergic activity in rat cultured astrocytes. In CHO cell line co-expressing human UT and combinations of GABA(A)R subunits, UII markedly depressed the GABA current (β(3)γ(2)>α(2)β(3)γ(2)>α(2)β(1)γ(2)). This effect, characterized by a fast short-term inhibition followed by drastic and irreversible run-down, is not relayed by G proteins. The run-down partially involves Ca(2+) and phosphorylation processes, requires dynamin, and results from GABA(A)R internalization. Thus, activation of the vasoactive G protein-coupled receptor UT triggers functional inhibition and endocytosis of GABA(A)R in CHO and human astrocytes, via its receptor C-terminus. This UII-induced disappearance of the repressor activity of GABA(A)R, may play a key role in the initiation of astrocyte proliferation.     

Adaptive value of phenological traits in stressful environments: predictions based on seed production and laboratory natural selection

Phenological traits often show variation within and among natural populations of annual plants. Nevertheless, the adaptive value of post-anthesis traits is seldom tested. In this study, we estimated the adaptive values of pre- and post-anthesis traits in two stressful environments (water stress and interspecific competition), using the selfing annual species Arabidopsis thaliana. By estimating seed production and by performing laboratory natural selection (LNS), we assessed the strength and nature (directional, disruptive and stabilizing) of selection acting on phenological traits in A. thaliana under the two tested stress conditions, each with four intensities. Both the type of stress and its intensity affected the strength and nature of selection, as did genetic constraints among phenological traits. Under water stress, both experimental approaches demonstrated directional selection for a shorter life cycle, although bolting time imposes a genetic constraint on the length of the interval between bolting and anthesis. Under interspecific competition, results from the two experimental approaches showed discrepancies. Estimation of seed production predicted directional selection toward early pre-anthesis traits and long post-anthesis periods. In contrast, the LNS approach suggested neutrality for all phenological traits. This study opens questions on adaptation in complex natural environment where many selective pressures act simultaneously.     

Differential contribution of the repeats to heparin binding of HBHA, a major adhesin of Mycobacterium tuberculosis

Background

Tuberculosis remains one of the most important causes of global mortality and morbidity, and the molecular mechanisms of the pathogenesis are still incompletely understood. Only few virulence factors of the causative agent Mycobacterium tuberculosis are known. One of them is the heparin-binding haemagglutinin (HBHA), an important adhesin for epithelial cells and an extrapulmonary dissemination factor. HBHA mediates mycobacterial adherence to epithelial cells via the interactions of its C-terminal, lysine rich repeat domain with sulfated glycoconjugates on the surface of epithelial cells.

Methodology/Principal Findings

Using defined heparin sulfate (HS) analogs, we determined the minimal heparin fragment length for HBHA binding and structural adaptations of the HBHA heparin-binding domain (HBD) upon binding to heparin. The NMR studies show significant shifts of all residues in the HBD upon interaction with heparin, with stronger shifts in the last repeats compared to the upstream repeats, and indicated that the HS fragments with 14 sugar units cover the entire C-terminal lysine-rich domain of HBHA. The differential implication of the repeats is determined by the relative position of prolines and lysines within each repeat, and may contribute to binding specificity. GAG binding induces a non-homogeneous structural rearrangement in the HBD, with stabilization of a nascent α-helix only in the last penta-repeats.

Conclusion/Significance

Mycobacterial HBHA undergoes structural adaptation upon interaction with GAGs, which is likely involved in binding specificities of the adhesin, and mycobacterial pathogens may use HBD polymorphisms for host or organ specificity. Further studies will aim at decoding the complementarity between HBD repeats and HS sequence.     

On the influence of prior information evaluated by fully Bayesian criteria in a personalized whole-brain model of epilepsy spread

Individualized anatomical information has been used as prior knowledge in Bayesian inference paradigms of whole-brain network models. However, the actual sensitivity to such personalized information in priors is still unknown. In this study, we introduce the use of fully Bayesian information criteria and leave-one-out cross-validation technique on the subject-specific information to assess different epileptogenicity hypotheses regarding the location of pathological brain areas based on a priori knowledge from dynamical system properties. The Bayesian Virtual Epileptic Patient (BVEP) model, which relies on the fusion of structural data of individuals, a generative model of epileptiform discharges, and a self-tuning Monte Carlo sampling algorithm, is used to infer the spatial map of epileptogenicity across different brain areas. Our results indicate that measuring the out-of-sample prediction accuracy of the BVEP model with informative priors enables reliable and efficient evaluation of potential hypotheses regarding the degree of epileptogenicity across different brain regions. In contrast, while using uninformative priors, the information criteria are unable to provide strong evidence about the epileptogenicity of brain areas. We also show that the fully Bayesian criteria correctly assess different hypotheses about both structural and functional components of whole-brain models that differ across individuals. The fully Bayesian information-theory based approach used in this study suggests a patient-specific strategy for epileptogenicity hypothesis testing in generative brain network models of epilepsy to improve surgical outcomes.     

Towards identifying genes underlying ecologically relevant traits in Arabidopsis thaliana

A major challenge in evolutionary biology and plant breeding is to identify the genetic basis of complex quantitative traits, including those that contribute to adaptive variation. Here we review the development of new methods and resources to fine-map intraspecific genetic variation that underlies natural phenotypic variation in plants. In particular, the analysis of 107 quantitative traits reported in the first genome-wide association mapping study in Arabidopsis thaliana sets the stage for an exciting time in our understanding of plant adaptation. We also argue for the need to place phenotype-genotype association studies in an ecological context if one is to predict the evolutionary trajectories of plant species.