Compositionally and functionally distinct editosomes in Trypanosoma brucei

Uridylate insertion/deletion RNA editing in Trypanosoma brucei mitochondria is catalyzed by a multiprotein complex, the approximately 20S editosome. Editosomes purified via three related tagged RNase III proteins, KREN1 (KREPB1/TbMP90), KREPB2 (TbMP67), and KREN2 (KREPB3/TbMP61), had very similar but nonidentical protein compositions, and only the tagged member of these three RNase III proteins was identified in each respective complex. Three new editosome proteins were also identified in these complexes. Each tagged complex catalyzed both precleaved insertion and deletion editing in vitro. However, KREN1 complexes cleaved deletion but not insertion editing sites in vitro, and, conversely, KREN2 complexes cleaved insertion but not deletion editing sites. These specific nuclease activities were abolished by mutations in the putative RNase III catalytic domain of the respective proteins. Thus editosomes appear to be heterogeneous in composition with KREN1 complexes catalyzing cleavage of deletion sites and KREN2 complexes cleaving insertion sites while both can catalyze the U addition, U removal, and ligation steps of editing.     

Responses of citrus plants to ozone: leaf biochemistry, antioxidant mechanisms and lipid peroxidation.

The effects of ozone upon 3-year-old trees of Clementina mandarin (Citrus clementina Hort. ex Tan.) cv. Marisol exposed for 12 months to ambient (10 nl l(-1)) and high (30 and 65 nl l(-1)) concentrations in open top chambers (OTCs) were investigated. The data showed that in leaves, ozone reduced total chlorophylls, carotenoid and carbohydrate concentration, and increased 1-aminocyclopropane-1-carboxylic acid (ACC) content and ethylene production. In treated plants, the ascorbate leaf pool was decreased, while lipid peroxidation and solute leakage were significantly higher than in ozone-free controls. The data indicated that ozone triggered protective mechanisms against oxidative stress in citrus.     

Offspring spatial patterns in Picconia excelsa (Oleaceae) in the Canarian laurel forest

We studied the spatial patterns of seedlings and seeds in isolated Picconia excelsa (Oleaceae) trees in the laurel forest of Anaga, Tenerife (Canary Islands). By finding isolated trees we assessed the correlation of seed and seedling bank traits and parent trees by removing the confounding effects of proximity (<100 m radius) of conspecific fruiting trees. We counted all the seedlings per age (height) class within its parental range, and sampled the seed number along transects departing from beneath the parent canopy at regular intervals. We mapped all seedlings per age class and plotted seed and seedling profiles in relation to distance to parent trees. Older Picconia seedlings tended to clump significantly further from parent trees than younger seedlings, which clumped just beneath the parents. We found significant differences among distances to parent tree in numbers of seedlings per age class. The seedling bank area was significantly correlated with maximum distance of seedlings to parent trees. The majority of seeds were deposited within the first 4 m below the parent crown. Seedlings amount at further distances from the trees is larger than seeds/fruits as counted on the ground. Our results suggest that disseminated, older seedlings have occupied germination sites far from the parent tree because there is probably lower seedling–seedling and parent–seedling competition for resources, and perhaps no intraspecific allelopathy and predation/disease.     

Refractoriness of urethral striated muscle contractility to nitric oxide-dependent cyclic GMP production

The purpose of this study was to investigate the role of cyclic GMP (cGMP) in the effects of nitric oxide (NO) on urethral striated muscle and its involvement in contractile function. The localization of cGMP, neuronal NO synthase (nNOS), vimentin, and neuronal markers was assessed by immunofluorescence in the sheep and rat urethra and the expression of nNOS was determined in Western blots. Nerve-mediated contractile responses to electrical field stimulation (EFS) were recorded in the sheep urethra. The scant nitrergic innervation of the striated muscle layer suggests that autonomic control of its activity is unlikely. The striated fiber itself may be the source of high levels NO produced by sarcolemmal and/or cytosolic μ or α variant of nNOS. This endogenous NO may provoke high basal production of soluble guanylate cyclase (GC) dependent cGMP, mainly in non-NO producing muscle fibers, which is not further enhanced by NO donors. cGMP co-localizes with neurofilament and PGP 9.5 at muscle endplates. Modulators of the cGMP pathway did not affect nerve-mediated contractile activity induced by EFS, suggesting that cGMP is not a significant mediator of neuromuscular transmission. In addition, NO donors did increase the accumulation of cGMP in dense networks of vimentin immunoreactive interstitial cells of Cajal (ICC), whose function is not yet known. These data suggest that there is a strong but non-regulated production of cGMP under resting conditions, which does not seem to affect contractile function. Modulation of cholinergic neurotransmission by NO through cGMP-independent mechanisms cannot be discarded.     

Pathways to extinction: beyond the error threshold

Since the introduction of the quasispecies and the error catastrophe concepts for molecular evolution by Eigen and their subsequent application to viral populations, increased mutagenesis has become a common strategy to cause the extinction of viral infectivity. Nevertheless, the high complexity of virus populations has shown that viral extinction can occur through several other pathways apart from crossing an error threshold. Increases in the mutation rate enhance the appearance of defective forms and promote the selection of mechanisms that are able to counteract the accelerated appearance of mutations. Current models of viral evolution take into account more realistic scenarios that consider compensatory and lethal mutations, a highly redundant genotype-to-phenotype map, rough fitness landscapes relating phenotype and fitness, and where phenotype is described as a set of interdependent traits. Further, viral populations cannot be understood without specifying the characteristics of the environment where they evolve and adapt. Altogether, it turns out that the pathways through which viral quasispecies go extinct are multiple and diverse.     

Dietary Exposure to Metals by Individuals Living Near a Hazardous Waste Incinerator in Catalonia, Spain: Temporal Trend

The concentrations of arsenic (As), beryllium (Be), cadmium (Cd), chromium (Cr), mercury (Hg), manganese (Mn), nickel (Ni), lead (Pb), tin (Sn), thallium (Tl), and vanadium (V) were determined in a number of foodstuffs purchased during 2006 in various localities of Tarragona County (Catalonia, Spain), near a hazardous waste incinerator, which has been operating since 1999. The dietary intake of the above elements by the population of the area was subsequently estimated. The results were compared with those obtained in previous surveys (baseline 1997 and 2002) performed in the same area. The levels of Be, Tl, and V were under their respective limits of detection in all samples. The estimated dietary intakes of the elements analyzed in the current survey by an adult man of 70 kg were the following: 351?μg/day for As, 4.6?μg/day for Cd, 57.5?μg/day for Cr, 7.1?μg/day for Hg, 2,229?μg/day for Mn, 78.0?μg/day for Ni, 39.9?μg/day for Pb, and 37.9?μg/day for Sn. On average, fish and seafood were the main contributor to the total dietary intake of As, Cd, and Hg and one of the most important for Pb. In general terms, the differences on metal exposure through dietary intake between the present and our two previous surveys were not particularly relevant. The current intake of the most toxic elements (As, Cd, Hg, and Pb) remains under the respective Provisional Tolerable Weekly Intakes established by the Food and Agriculture Organization/World Health Organization.     

The pcsA gene from Streptomyces diastaticus var. 108 encodes a polyene carboxamide synthase with broad substrate specificity for polyene amides biosynthesis

Two structurally related polyene macrolides are produced by Streptomyces diastaticus var. 108: rimocidin (3a) and CE-108 (2a). Both bioactive metabolites are biosynthesized from the same pathway through type I polyketide synthases by choosing a starter unit either acetate or butyrate, resulting in 2a or 3a formation, respectively. Two additional polyene amides, CE-108B (2b) and rimocidin B (3b), are also produced “in vivo” when this strain was genetically modified by transformation with engineered SCP2*-derived vectors carrying the ermE gene. The two polyene amides, 2b and 3b, showed improved pharmacological properties, and are generated by a tailoring activity involved in the conversion of the exocyclic carboxylic group of 2a and 3a into their amide derivatives. The improvement on some biological properties of the resulting polyenes, compared with that of the parental compounds, encourages our interest for isolating the tailoring gene responsible for the polyene carboxamide biosynthesis, aimed to use it as tool for generating new bioactive compounds. In this work, we describe the isolation from S. diastaticus var. 108 the corresponding gene, pcsA, encoding a polyene carboxamide synthase, belonging to the Class II glutamine amidotransferases and responsible for “in vivo” and “in vitro” formation of CE-108B (2b) and rimocidin B (3b). The fermentation broth from S. diastaticus var. 108 engineered with the appropriate pcsA gene construction, showed the polyene amides to be the major bioactive compounds.     

Young Children’s Drawings of Plant Life: A Study Concerning the Use of Colours and its Relationship with Age

This study examines the drawings that elementary grade school children make on the subject of plant life. More specifically, the pictorial elements drawn by children are analysed together with their colour choices and the size of coloured surfaces. Furthermore, the results are put into perspective with the age of the children in the sample. The results indicate that there are significant differences in terms of both the content of the drawings and how colours are used. These findings are linked to the process of the comprehension of the plant world that, according to several studies, initially begins in elementary education. The conclusions highlight the convenience of considering the study of young children’s drawings and colour choices in order to achieve a clearer picture regarding the initial steps of the process of the understanding of biological phenomena in early childhood.     

Distinct platelet-activating factor binding sites in synaptic endings and in intracellular membranes of rat cerebral cortex

The binding of 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine (AGEPC or PAF, platelet-activating factor) to synaptic plasma membranes, microsomal membranes, and other rat cerebral cortex subcellular fractions was studied. Using several PAF-binding antagonists, three distinct sites were identified. Two of them were in intracellular membranes (microsomes) and one in synaptic plasma membranes. Microsomal membranes were prepared after obtaining a 43,500 x g pellet from the postmitochondrial supernatant and subsequent centrifugation at 105,000 x g of the resulting supernatant. Most plasma membrane markers were retained in the 43,500 x g pellet (Sun, G.Y., Huang, H.-M., Kelleher, J.A., Stubbs, E.B., Sun, A. Y. (1988) Neurochem. Int. 12, 69-77). Microsomes were purified by density-gradient centrifugation and marker enzymes showed relatively very low contamination by plasma membrane markers. Myelin and mitochondria were devoid of specific PAF binding. A site displaying the highest PAF-binding affinity reported to date in all cells and membranes (KD = 22.5 +/- 1.7 pM and Bmax 8.75 = fmol/mg protein), was found in the microsomal fraction. There was a second binding site in microsomal fractions (KD = 25.0 +/- 0.8 nM and Bmax = 0.96 pmol/mg protein. Ca2+ decreases PAF affinity for the microsomal binding sites. The third binding site displays relatively low specific PAF binding and is present in synaptosomal plasma membranes. Moreover, displacement curves by a wide variety of PAF antagonists indicated different affinities for each of the binding sites described here. These results indicate that PAF-binding sites are heterogeneous in rat cerebral cortex, and they imply that the microsomal membrane sites may be involved, at least in part, in intracellular events such as gene expression.     

Regulation of plasma sex-steroid-binding protein in adult cynomolgus monkeys (Macaca fascicularis) during different reproductive states

Levels of the sex-steroid binding protein (mSBP) have been characterized in Cynomolgus monkeys, during different stages of reproduction and under hormonal treatments, by an immunoassay allowing a specific and accurate measurement of the protein itself. Using an antiserum specific for native human SBP, we have determined the mSBP level by electroimmunodiffusion. This method correlates closely with the binding capacity measured by a steady-state polyacrylamide electrophoresis. The levels are lower in males (M = 88 +/- 8.6 nmoles/1, n = 6) than in females in the follicular phase (M = 123 +/- 4.4 nmoles/1, n = 5, p less than 0.001) and in castrated males (M = 172 +/- 13.3 nmoles/1, n = 3, p less than 0.001). During gestation, the SBP level decreases (M = 69 +/- 16.3 nmoles/1, n = 7, in the latter part of pregnancy), but during lactation, it is similar to follicular values. Estrogen treatment fails to increase SBP levels in castrated animals, but the values are reduced by testosterone treatment. Since these results are different from those observed in women, we question the validity of using monkeys as models for understanding the mechanisms controlling the concentration of SBP in human blood.