Further characterization of bovine pancreatic lipase.

1. The amino acid composition of bovine pancreatic lipase is very similar to that of porcine lipase. 2. Bovine lipase possesses a residue of lysine at the N-terminal position and a half cystine or a cysteine at the C-terminal position. 3. Bovine lipase contains two free sulfhydryl groups of different reactivities to 5, 5'-dithiobis-(2-nitrobenzoic) acid. One of these groups is buried in the native conformation of the enzyme and is fully titrated in 1.5 M urea when reaction is performed in the presense of 1 mM EDTA.     

Base Flipping in Tn10 Transposition: An Active Flip and Capture Mechanism

The bacterial Tn5 and Tn10 transposases have a single active site that cuts both strands of DNA at their respective transposon ends. This is achieved using a hairpin intermediate that requires the DNA to change conformation during the reaction. In Tn5 these changes are controlled in part by a flipped nucleoside that is stacked on a tryptophan residue in a hydrophobic pocket of the transposase. Here we have investigated the base flipping mechanism in Tn10 transposition. As in Tn5 transposition, we find that base flipping takes place after the first nick and is required for efficient hairpin formation and resolution. Experiments with an abasic substrate show that the role of base flipping in hairpin formation is to remove the base from the DNA helix. Specific interactions between the flipped base and the stacking tryptophan residue are required for hairpin resolution later in the reaction. We show that base flipping in Tn10 transposition is not a passive reaction in which a spontaneously flipped base is captured and retained by the protein. Rather, it is driven in part by a methionine probe residue that helps to force the flipped base from the base stack. Overall, it appears that base flipping in Tn10 transposition is similar to that in Tn5 transposition.     

Co-culture of human fibroblasts,smooth muscle and endothelial cells promotes osteopontin induction in hypoxia

Arteriovenous fistulas (AVFs) are the preferred vascular access for haemodialysis of patients suffering from end-stage renal disease, a worldwide public health problem. However, they are prone to a high rate of failure due to neointimal hyperplasia and stenosis. This study aimed to determine if osteopontin (OPN) was induced in hypoxia and if OPN could be responsible for driving AVF failure. Identification of new factors that participate in remodelling of AVFs is a challenge. Three cell lines representing the cells of the three layers of the walls of arteries and veins, fibroblasts, smooth muscle cells and endothelial cells, were tested in mono- and co-culture in vitro for OPN expression and secretion in normoxia compared to hypoxia after silencing the hypoxia-inducible factors (HIF-1α, HIF-2α and HIF-1/2α) with siRNA or after treatment with an inhibitor of NF-kB. None of the cells in mono-culture showed OPN induction in hypoxia, whereas cells in co-culture secreted OPN in hypoxia. The changes in oxygenation that occur during AVF maturation up-regulate secretion of OPN through cell-cell interactions between the different cell layers that form AVF, and in turn, these promote endothelial cell proliferation and could participate in neointimal hyperplasia.     

Climate‐associated genetic variation in Fagus sylvatica and potential responses to climate change in the French Alps

Local adaptation patterns have been found in many plants and animals, highlighting the genetic heterogeneity of species along their range of distribution. In the next decades, global warming is predicted to induce a change in the selective pressures that drive this adaptive variation, forcing a reshuffling of the underlying adaptive allele distributions. For species with low dispersion capacity and long generation time such as trees, the rapidity of the change could impede the migration of beneficial alleles and lower their capacity to track the changing environment. Identifying the main selective pressures driving the adaptive genetic variation is thus necessary when investigating species capacity to respond to global warming. In this study, we investigate the adaptive landscape of Fagus sylvatica along a gradient of populations in the French Alps. Using a double‐digest restriction‐site‐associated DNA (ddRAD) sequencing approach, we identified 7,000 SNPs from 570 individuals across 36 different sites. A redundancy analysis (RDA)‐derived method allowed us to identify several SNPs that were strongly associated with climatic gradients; moreover, we defined the primary selective gradients along the natural populations of F. sylvatica in the Alps. Strong effects of elevation and humidity, which contrast north‐western and south‐eastern site, were found and were believed to be important drivers of genetic adaptation. Finally, simulations of future genetic landscapes that used these findings allowed identifying populations at risk for F. sylvatica in the Alps, which could be helpful for future management plans.