Psoralen-sensitive mutant pso9-1 of Saccharomyces cerevisiae contains a mutant allele of the DNA damage checkpoint gene MEC3

Complementation analysis of the pso9-1 yeast mutant strain sensitive to photoactivated mono- and bifunctional psoralens, UV-light 254 nm, and nitrosoguanidine, with pso1 to pso8 mutants, confirmed that it contains a novel pso mutation. Molecular cloning via the reverse genetics complementation approach using a yeast genomic library suggested pso9-1 to be a mutant allele of the DNA damage checkpoint control gene MEC3. Non-complementation of several sensitivity phenotypes in pso9-1/mec3Delta diploids confirmed allelism. The pso9-1 mutant allele contains a -1 frameshift mutation (deletion of one A) at nucleotide position 802 (802delA), resulting in nine different amino acid residues from that point and a premature termination. This mutation affected the binding properties of Pso9-1p, abolishing its interactions with both Rad17p and Ddc1p. Further interaction assays employing mec3 constructions lacking the last 25 and 75 amino acid carboxyl termini were also not able to maintain stable interactions. Moreover, the pso9-1 mutant strain could no longer sense DNA damage since it continued in the cell cycle after 8-MOP + UVA treatment. Taken together, these observations allowed us to propose a model for checkpoint activation generated by photo-induced adducts.     

Diversification of plant species in a subtropical region of eastern South American highlands: a phylogeographic perspective on native Petunia (Solanaceae)

In the Southern and Southeastern Brazilian highlands, a clade of seven species of Petunia that are endemic to the region (P. altiplana, P. bonjardinensis, P. guarapuavensis, P. mantiqueirensis, P. reitzii, P. saxicola and P. scheideana) exists in association with grassland formations. These formations are isolated in high-altitude regions, being surrounded by forested areas, and experienced contraction-expansion cycles associated with the glacial cycles of the Pleistocene. To understand the evolutionary history of this group, the divergence of which is probably linked to these past shifts in habitat, we analysed the sequences of the plastidial intergenic spacers trnH-psbA and trnS-trnG from populations throughout the known distributions of all seven species. The common ancestor of this highland clade started to differentiate ～0.9 million years (Myr) ago, which corresponds to a high diversification rate of 2.06 species per Myr in the intervening period. The high level of haplotype sharing among several species in the clade and the absence of reciprocal monophyly suggest the persistence of ancestral polymorphisms during speciation events and/or past hybridization, because no hybrid was found. Four of the five species displayed very low genetic diversity and possessed either one or two haplotypes, which is consistent with long-term isolation in restricted areas. The three more diverse species displayed significant population structure, and P. altiplana showed a clear signs of population growth during the last glacial period. These results suggest that diversification occurred as a result of expansion of the ancestral species of the clade during glacial periods followed by fragmentation and isolation during retraction in interglacial periods.     

Cross-amplification and characterization of 13 tetranucleotide microsatellites in multiple species of Neotropical canids

We tested 47 tetranucleotide microsatellite loci developed for the domestic dog in four species of Neotropical canids, aiming to produce a standardized set that could be successfully used even in noninvasive samples across this group. We identified 13 suitable loci, nine of which constitute a standardized set for all species. Considering only the ideal panel of nine loci, the mean expected heterozygosity (averaged across species) per locus ranged from 0.58 to 0.92 (overall mean 0.76), and the maximum probability of identity value was 1.3 × 10^?9. This set of loci has a great potential for application in evolutionary, ecological and conservation studies.     

Mycoflora and mycotoxins natural occurrence in corn from entre rios province, Argentina

Corn samples were collected in 1999 from three departments of Entre Réos province, Argentina, and were surveyed for mould contamination and natural occurrence ofFusarium mycotoxins, ochratoxin A and aflatoxins.Fusarium verticillioides was the most prevalent fungal species recorded at all departments. Zearalenone, deoxynivalenol and ochratoxin A were not found in any samples. Only one of the 52 corn samples analysed was contaminated with aflatoxin B₁ (17 μg/kg). Fumonisin B₁ was found in 58 % of samples (range of positive samples: 47– 3,347 μg/kg), fumonisin B₂ in 33.0 % (range of positive samples: 23–537 μg/kg) and fumonisin B₃ in 25.0 % (range of positive samples: 24–287 μg/kg) of them. This is the first report on the natural occurrence of mycotoxins in corn from Entre Ríos province, Argentina. Levels of fumonisins were lower than detected in other Argentinian provinces.     

Molecular action of methotrexate in inflammatory diseases

Despite the recent introduction of biological response modifiers and potent new small-molecule antirheumatic drugs, the efficacy of methotrexate is nearly unsurpassed in the treatment of inflammatory arthritis. Although methotrexate was first introduced as an antiproliferative agent that inhibits the synthesis of purines and pyrimidines for the therapy of malignancies, it is now clear that many of the anti-inflammatory effects of methotrexate are mediated by adenosine. This nucleoside, acting at one or more of its receptors, is a potent endogenous anti-inflammatory mediator. In confirmation of this mechanism of action, recent studies in both animals and patients suggest that adenosine-receptor antagonists, among which is caffeine, reverse or prevent the anti-inflammatory effects of methotrexate.     

Mitochondrial population genomics supports a single pre-Clovis origin with a coastal route for the peopling of the Americas

It is well accepted that the Americas were the last continents reached by modern humans, most likely through Beringia. However, the precise time and mode of the colonization of the New World remain hotly disputed issues. Native American populations exhibit almost exclusively five mitochondrial DNA (mtDNA) haplogroups (A-D and X). Haplogroups A-D are also frequent in Asia, suggesting a northeastern Asian origin of these lineages. However, the differential pattern of distribution and frequency of haplogroup X led some to suggest that it may represent an independent migration to the Americas. Here we show, by using 86 complete mitochondrial genomes, that all Native American haplogroups, including haplogroup X, were part of a single founding population, thereby refuting multiple-migration models. A detailed demographic history of the mtDNA sequences estimated with a Bayesian coalescent method indicates a complex model for the peopling of the Americas, in which the initial differentiation from Asian populations ended with a moderate bottleneck in Beringia during the last glacial maximum (LGM), around approximately 23,000 to approximately 19,000 years ago. Toward the end of the LGM, a strong population expansion started approximately 18,000 and finished approximately 15,000 years ago. These results support a pre-Clovis occupation of the New World, suggesting a rapid settlement of the continent along a Pacific coastal route.     

Production of Short Side Chain-Poly[Hydroxyalkanoate] by a Newly Isolated Ralstonia Pickettii Strain

In order to obtain better bacterial species or strains for production of short side chain-poly[hydroxyalkanoate](ssc-PHA) from cheap carbon sources, a bioprospecting programme was performed in a subtropical rainforest soil. From 398 bacterial isolates, one produced high amounts of ssc-PHA when grown on sugarcane molasses or sucrose as detected by spectrophotometric scanning and gas chromatography coupled to mass spectrometry. Also, the GC—MS analysis indicated that the polymer was composed basically of poly[3-hydroxybutyrate](PHB). Phylogenetic studies using 16S rDNA analysis showed that the isolated bacterium belonged to the Ralstonia pickettii species and had a high identity/similarity with 16S rDNA obtained from total DNA of uncultured strains of soils and with unidentified bacteria at species level. The new strain was named R. pickettii 61A6. Spectrofluorometric analysis showed that the best rates of ssc-PHA accumulation within the cells occurred in 10%(w/v) sucrose and in 5%(w/v) sugarcane molasses at the stationary phase, with a yield of 231 and 357 mg/l of ssc-PHA per g dry cell weight, respectively.     

Lifelong exposure to dietary fish oil alters macrophage responses in Walker 256 tumor-bearing rats

Supplementation of the diet with fish oil (FO) decreases growth of the Walker 256 tumor and decreases the cachexia associated with tumor-bearing. The mechanisms by which FO inhibits tumor growth and cachexia are unknown. Macrophages are very important in host defence against tumors since they produce several anti-tumor agents which in turn have been shown to be modified by dietary FO, but rarely in the setting of tumor bearing and never in relation to lifelong exposure. In this study, we compared the effects of supplementation of the diet of pregnant and lactating rats and subsequent supplementation of the offspring with coconut fat or FO on macrophage activities involved in anti-tumor defence. FO supplementation was able to induce an increase in phagocytosis, in O2-, H2O2, nitric oxide, and TNF-alpha production by macrophages and in lysosomal volume in non-tumor-bearing rats. However, phagocytosis, production of O2- and H2O2 and lysosomal volume were not affected by the FO diet when rats were bearing tumors, although nitric oxide production was higher in these animals. It appears that tumor bearing activates the innate immune system and that dietary FO has little effect on innate immunity in the presence of Walker 256 tumors. Thus, it is still unclear how FO decreases the growth of Walker 256 tumors and the associated cachexia.     

Diversity and Age of the Four Major mtDNA Haplogroups, and Their Implications for the Peopling of the New World

Despite considerable investigation, two main questions on the origin of Native Americans remain the topic of intense debate-namely, the number and time of the migration(s) into the Americas. Using the 720 available Amerindian mtDNA control-region sequences, we reanalyzed the nucleotide diversity found within each of the four major mtDNA haplogroups (A-D) thought to have been present in the colonization of the New World. We first verified whether the within-haplogroup sequence diversity could be used as a measure of the haplogroup's age. The pattern of shared polymorphism, the mismatch distribution, the phylogenetic trees, the value of Tajima's D, and the computer simulations all suggested that the four haplogroups underwent a bottleneck followed by a large population expansion. The four haplogroup diversities were very similar to each other, offering a strong support for their single origin. They suggested that the beginning of the Native Americans' ancestral-population differentiation occurred approximately 30,000-40,000 years before the present (ybp), with a 95%-confidence-interval lower bound of approximately 25,000 ybp. These values are in good agreement with the New World-settlement model that we have presented elsewhere, extending the results initially found for haplogroup A to the three other major groups of mtDNA sequences found in the Americas. These results put the peopling of the Americas clearly in an early, pre-Clovis time frame.