Phylogeography of the Guinea multimammate mouse (Mastomys erythroleucus): a case study for Sahelian species in West Africa

Aim To investigate the phylogeographical structure of the Guinea multimammate mouse, Mastomys erythroleucus (Temminck, 1853), a widespread murid rodent in sub‐Saharan (Sahel and Sudan) savannas, for a better understanding of the impacts of geographical and historical factors on the evolutionary history of this species, in the context of the growing database of phylogeographical studies of African savanna mammal species. Location Sahel and Sudan savannas, Africa. Methods We sequenced the whole cytochrome b gene in 211 individuals from 59 localities distributed from Senegal to Ethiopia. Sequence data were analysed using both phylogenetic (several rooted tree‐construction methods, median‐joining networks) and population genetic methods (spatial analyses of molecular variance, mismatch distributions). Results Haplotypes were distributed into four major monophyletic groups corresponding to distinct geographical regions across a west–east axis. Diversification events were estimated to have occurred between 1.16 and 0.18 Ma. Main conclusions Vicariance events related to the fragmentation of savanna habitats during the Pleistocene era may explain the phylogeographical patterns observed. Genetic structure was consistent with a role of major Sahelian rivers as significant barriers to west–east dispersal. Recent demographic expansions probably occurred during arid phases of the Holocene with the southward expansion of savannas.     

Stable transmission of targeted gene modification using single-stranded oligonucleotides with flanking LNAs

Targeted mutagenesis directed by oligonucleotides (ONs) is a promising method for manipulating the genome in higher eukaryotes. In this study, we have compared gene editing by different ONs on two new target sequences, the eBFP and the rd1 mutant photoreceptor βPDE cDNAs, which were integrated as single copy transgenes at the same genomic site in 293T cells. Interestingly, antisense ONs were superior to sense ONs for one target only, showing that target sequence can by itself impart strand-bias in gene editing. The most efficient ONs were short 25 nt ONs with flanking locked nucleic acids (LNAs), a chemistry that had only been tested for targeted nucleotide mutagenesis in yeast, and 25 nt ONs with phosphorothioate linkages. We showed that LNA-modified ONs mediate dose-dependent target modification and analyzed the importance of LNA position and content. Importantly, when using ONs with flanking LNAs, targeted gene modification was stably transmitted during cell division, which allowed reliable cloning of modified cells, a feature essential for further applications in functional genomics and gene therapy. Finally, we showed that ONs with flanking LNAs aimed at correcting the rd1 stop mutation could promote survival of photoreceptors in retinas of rd1 mutant mice, suggesting that they are also active in vivo.     

Supported membrane composition analysis by secondary ion mass spectrometry with high lateral resolution

The lateral organization of lipid components within membranes is usually investigated with fluorescence microscopy, which, though highly sensitive, introduces bulky fluorophores that might alter the behavior of the components they label. Secondary ion mass spectroscopy performed with a NanoSIMS 50 instrument also provides high lateral resolution and sensitivity, and many species can be observed in parallel without the use of bulky labels. A tightly focused beam (approximately 100 nm) of Cs ions is scanned across a sample, and up to five of the resulting small negative secondary ions can be simultaneously analyzed by a high-resolution mass spectrometer. Thin layers of (15)N- and (19)F-labeled proteins were microcontact-printed on an oxidized silicon substrate and imaged using the NanoSIMS 50, demonstrating the sensitivity and selectivity of this approach. Supported lipid bilayers were assembled on an oxidized silicon substrate, then flash-frozen and freeze-dried to preserve their lateral organization. Lipid bilayers were analyzed with the NanoSIMS 50, where the identity of each specific lipid was determined through detection of its unique secondary ions, including (12)C(1)H(-), (12)C(2)H(-), (13)C(-), (12)C(14)N(-), and (12)C(15)N(-). Steps toward obtaining quantitative composition analysis of lipid membranes that varied spatially in isotopic composition are presented. This approach has the potential to provide a composition-specific analysis of membrane organization that compliments other imaging modalities.     

Interaction between the ADH1C polymorphism and maternal alcohol intake in the risk of nonsyndromic oral clefts: an evaluation of the contribution of child and maternal genotypes

BACKGROUND: Maternal alcohol consumption has been associated with an increased risk of nonsyndromic oral clefts in some studies. Study of gene-environment interaction may provide insight into the reasons for their discrepancies observed. We focused on a polymorphism of the ADH1C gene (third gene of the class I alcohol dehydrogenase family), involved in the metabolism of ethanol and other alcohols. METHODS: Data come from a French case-control study (1998-2001), which tested the association between maternal alcohol consumption during the first trimester of pregnancy and the risk of nonsyndromic oral clefts (240 cases, 236 controls). A case-parent study design looked at the association with an ADH1C polymorphism (Ile349Val site) and potential gene-environment interaction effects. A log-linear model was used to distinguish the direct effect of the child's genotype from the maternally mediated effects. RESULTS: An increased risk of nonsyndromic oral clefts was observed for women who reported drinking alcohol during the first trimester, compared with women who did not. The mutated ADH1C allele carried by the child seemed to have a protective effect against the risk of oral clefts (RRone copy, 0.71; 95% confidence interval [CI], 0.50-1.02; RRtwo copies, 0.63; 95% CI, 0.3-1.3). The maternal genotype played a less important role than the child's, and its action remains unclear. No significant evidence of interaction effects between the ADH1C genotype and maternal alcohol consumption was observed. CONCLUSIONS: Because the ADH1C gene is involved in the metabolic pathways of many alcohols, we propose several hypotheses about the causal pathway, including ethanol oxidation activity and, more probably, retinol oxidation.     

Portal sensing of intestinal gluconeogenesis is a mechanistic link in the diminution of food intake induced by diet protein

Protein feeding is known to decrease hunger and subsequent food intake in animals and humans. It has also been suggested that glucose appearance into portal vein, as occurring during meal assimilation, may induce comparable effects. Here, we connect these previous observations by reporting that intestinal gluconeogenesis (i.e., de novo synthesis of glucose) is induced during the postabsorptive time (following food digestion) in rats specifically fed on protein-enriched diet. This results in glucose release into portal blood, counterbalancing the lowering of glycemia resulting from intestinal glucose utilization. Comparable infusions into the portal vein of control postabsorptive rats (fed on starch-enriched diet) decrease food consumption and activate the hypothalamic nuclei regulating food intake. Similar hypothalamic activation occurs on protein feeding. All these effects are absent after denervation of the portal vein. Thus, portal sensing of intestinal gluconeogenesis may be a novel mechanism connecting the macronutrient composition of diet to food intake.     

Bone marrow-derived dendritic cells reverse the anergic state of CD4+CD25+ T cells without reversing their suppressive function

Dendritic cells (DC) are potent inducers of immunity to foreign Ags, but also contribute to self-tolerance by induction of regulatory T cells or deletion/anergy of self-reactive T cells. In this study, we have studied the capacity of DC to activate naturally occurring CD4+CD25+ regulatory T cells as well as the ability of CD4+CD25+ T cells to suppress the DC-mediated activation of CD4+CD25- T cells. Mature bone marrow-derived dendritic cells, but not splenic DC, were able to induce the proliferation of CD4+CD25+ T cells in the presence of a polyclonal stimulus and in the absence of exogenous IL-2. The DC-induced proliferative response of the CD4+CD25+ T cells was partially dependent on IL-2 produced by a small number of contaminating CD25+ effector cells. Because bone marrow-derived dendritic cells induce proliferation of both CD4+CD25+ and CD4+CD25- T cells in vitro, it was impossible to assay the suppressive function of the CD4+CD25+ T cells using [3H]TdR uptake or CFSE dilution. We therefore measured IL-2 production in cocultures of CD4+CD25+ and CD4+CD25- T cells using the IL-2 secretion assay. Surprisingly, CD4+CD25+ T cells markedly suppressed IL-2 secretion by the CD4+CD25- T cells without inhibiting their proliferation. Collectively, these results suggest that Ag presentation by DC can induce the expansion of CD4+CD25+ T cells while simultaneously activating their ability to suppress cytokine secretion by effector T cells.