Plankton relationships under small water level fluctuations in a subtropical reservoir

In reservoirs, variations in water level may affect plankton biomass and species composition. Studies on the effect of water-level fluctuations are scarce and restricted to Europe and Australia. In the Río Tercero Reservoir (Argentina), the management policies of a nuclear-power plant require a minimum depth of 650 m. During periods of excessive rainfall, however, the input is such that the excess passes over the spillway, thus causing a high turnover of water. Phytoplankton, zooplankton, and physicochemical variables were monitored over 2 years at three sampling stations during a period with annual precipitation higher than the historical annual mean. Different hydrological situations occurred based on precipitation, spillway outflow, and water-renewal rate. At high renewal rates, phyto- and zoo-plankton diversities peaked. During high outflow periods phytoplankton biomass peaked through the contribution of Ceratium hirundinella. Once the spillway outflow ceased, stable conditions (low renewal rates) were achieved, thus allowing the onset of biological interactions. Maximum phytoplankton density (mainly Actinocyclus normanii) was reached at such times, and efficient grazers (Daphnia laevis) with long life cycles dominated in terms of biomass. The structure and dynamics of the plankton community could be altered by changes in hydrological conditions (renewal rate and spillway outflow) that act to compromise the apparent stability imposed by steady water levels. These variables must be considered to identify disturbance conditions and improve knowledge of reservoir environments, so as to implement appropriate management practices.     

A Single Mutation at the Sheet Switch Region Results in Conformational Changes Favoring λ6 Light-Chain Fibrillogenesis

Systemic amyloid light-chain (LC) amyloidosis is a disease process characterized by the pathological deposition of monoclonal LCs in tissue. All LC subtypes are capable of fibril formation although λ chains, particularly those belonging to the λ6 type, are overrepresented. Here, we report the thermodynamic and in vitro fibrillogenic properties of several mutants of the λ6 protein 6aJL2 in which Pro7 and/or His8 was substituted by Ser or Pro. The H8P and H8S mutants were almost as stable as the wild-type protein and were poorly fibrillogenic. In contrast, the P7S mutation decreased the thermodynamic stability of 6aJL2 and greatly enhanced its capacity to form amyloid-like fibrils in vitro. The crystal structure of the P7S mutant showed that the substitution induced both local and long-distance effects, such as the rearrangement of the V_L (variable region of the light chain)-V_L interface. This mutant crystallized in two orthorhombic polymorphs, P2₁2₁2₁ and C222₁. In the latter, a monomer that was not arranged in the typical Bence-Jones dimer was observed for the first time. Crystal-packing analysis of the C222₁ lattice showed the establishment of intermolecular β-β interactions that involved the N-terminus and β-strand B and that these could be relevant in the mechanism of LC fibril formation. Our results strongly suggest that Pro7 is a key residue in the conformation of the N-terminal sheet switch motif and, through long-distance interactions, is also critically involved in the contacts that stabilized the V_L interface in λ6 LCs.     

Identification of chalcones as in vivo liver monofunctional phase II enzymes inducers

Cancer preventive agents (CPA) are drugs able to suppress the carcinogen metabolic activation or block the formation of ultimate carcinogens. CPA could act through various molecular mechanisms, for example by interfering with the action of procarcinogen. This could be attained by increasing the phase II enzymes levels of quinone reductase (QR) and glutathione S-transferase (GST). New flavonoids, especially chalcones, have been identified as in vivo monofunctional phase II enzymes inducers. Oral administration of chalcone, 4, and both p-methoxy-substituted chalcones, 6 and 14, increased hepatic QR activity with concomitant decrease in CYP1A1 activity, a member of the most important group of phase I enzymes cytochrome P450. Among them, 4 also increased GST activity. While p-bromo-substituted chalcone 8 was the best inducer of QR it decreased hepatic GST expression and cytochrome P450, being the most effective decreasing cytochrome P450-expression. Thienyl-chalcone 20 being the bioisostere of chalcone 4 did not display the same in vivo profile in the phase I level modification. As chalcone 4 its bioisostere, chalcone 20, displayed low DNA strand breakage and absence of mutagenicity. Also, in our preliminary in vivo tumourigenesis/chemopreventive and acute-toxicity studies, chalcones 4, 6 and 8 showed the best behaviours as CPA justifying additional studies that are ongoing.     

Mucoidy,Quorum Sensing,Mismatch Repair and Antibiotic Resistance in Pseudomonas aeruginosa from Cystic Fibrosis Chronic Airways Infections

Survival of Pseudomonas aeruginosa in cystic fibrosis (CF) chronic infections is based on a genetic adaptation process consisting of mutations in specific genes, which can produce advantageous phenotypic switches and ensure its persistence in the lung. Among these, mutations inactivating the regulators MucA (alginate biosynthesis), LasR (quorum sensing) and MexZ (multidrug-efflux pump MexXY) are the most frequently observed, with those inactivating the DNA mismatch repair system (MRS) being also highly prevalent in P. aeruginosa CF isolates, leading to hypermutator phenotypes that could contribute to this adaptive mutagenesis by virtue of an increased mutation rate. Here, we characterized the mutations found in the mucA, lasR, mexZ and MRS genes in P. aeruginosa isolates obtained from Argentinean CF patients, and analyzed the potential association of mucA, lasR and mexZ mutagenesis with MRS-deficiency and antibiotic resistance. Thus, 38 isolates from 26 chronically infected CF patients were characterized for their phenotypic traits, PFGE genotypic patterns, mutations in the mucA, lasR, mexZ, mutS and mutL gene coding sequences and antibiotic resistance profiles. The most frequently mutated gene was mexZ (79%), followed by mucA (63%) and lasR (39%) as well as a high prevalence (42%) of hypermutators being observed due to loss-of-function mutations in mutL (60%) followed by mutS (40%). Interestingly, mutational spectra were particular to each gene, suggesting that several mechanisms are responsible for mutations during chronic infection. However, no link could be established between hypermutability and mutagenesis in mucA, lasR and mexZ, indicating that MRS-deficiency was not involved in the acquisition of these mutations. Finally, although inactivation of mucA, lasR and mexZ has been previously shown to confer resistance/tolerance to antibiotics, only mutations in MRS genes could be related to an antibiotic resistance increase. These results help to unravel the mutational dynamics that lead to the adaptation of P. aeruginosa to the CF lung.     

Towards the identification of late-embryogenic-abundant phosphoproteome in Arabidopsis by 2-DE and MS

Late-embryogenesis-abundant (LEA) proteins accumulate as plant seeds desiccate and also in vegetative organs during periods of stress. They are predicted to play a role in plant stress tolerance. In the present study, we have initiated the characterization of phosphorylated LEA proteins present in the Arabidopsis seed, using a strategy that combines the thermostability (solubility upon heating) of many LEA-type proteins with the use of phosphoaffinity chromatography to obtain an enriched subpopulation of phosphoproteins. The specificity and efficiency of the procedure was assessed by alkaline phosphatase treatment and by a specific stain for phosphoproteins, in addition to the immunodetection of AtRab18, a phosphorylated LEA protein present in the mature dry seed. The phosphoproteins were identified by MS either by PMF using MALDI-TOF MS after 2-DE separation, or by peptide sequencing using both capillary LC MS/MS (LC muESI-ITMS/MS) and nanoLC coupled to nanoESI-MS/MS (LC-nanoESI-Q-TOF-MS/MS). Several LEA-type and storage-like proteins were identified as components of the phosphoproteome of the Arabidopsis seed.     

Cytochalasin-D retards sperm incorporation deep into the egg cytoplasm but not membrane fusion with the egg plasma membrane

The fertilization process is impaired when spermatozoa are previously incubated with Cytochalasin-D (Cyt-D). Although this fact reveals the participation of polymerized actin in fertilization, the specific event obstructed by Cyt-D treatment has not been determined. To identify this event, we capacitated guinea pig spermatozoa in minimal capacitating medium with pyruvate and lactate (MCM-PL) with Cyt-D, to inseminate hamster zona pellucida (ZP)-free eggs. Cyt-D (70 microM) decreased F-actin relative concentration in capacitated spermatozoa to a larger extent than in spermatozoa incubated under control conditions. Cyt-D also cancelled the F-actin increase normally observed in acrosome-reacted cells, and decreased the number of these cells with normal F-actin localization at the equatorial zone. Insemination of eggs with Cyt-D treated spermatozoa did not change early fertilization events such as the egg cortical reaction (CR), membranes fusion, and egg F-actin new localization, but clearly retarded, by 16 hr, spermatozoa incorporation deep into the egg cytoplasm, and decondensation of egg metaphase II chromosomes. These results show that actin polymerization is necessary for spermatozoa incorporation deep into the egg cytoplasm, but not for plasma membrane fusion nor egg activation early steps.